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1
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Zencir S, Dilg D, Bruzzone M, Stutz F, Soudet J, Shore D, Albert B. A two-step regulatory mechanism dynamically controls histone H3 acetylation by SAGA complex at growth-related promoters. Nucleic Acids Res 2025; 53:gkaf276. [PMID: 40207626 PMCID: PMC11983098 DOI: 10.1093/nar/gkaf276] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/10/2024] [Revised: 03/03/2025] [Accepted: 03/31/2025] [Indexed: 04/11/2025] Open
Abstract
Acetylation of histone H3 at residue K9 (H3K9ac) is a dynamically regulated mark associated with transcriptionally active promoters in eukaryotes. However, our understanding of the relationship between H3K9ac and gene expression remains mostly correlative. In this study, we identify a large suite of growth-related (GR) genes in yeast that undergo a particularly strong down-regulation of both transcription and promoter-associated H3K9ac upon stress, and delineate the roles of transcriptional activators (TAs), repressors, SAGA (Spt-Ada-Gcn5 acetyltransferase) histone acetyltransferase, and RNA-polymerase II in this response. We demonstrate that H3K9 acetylation states are orchestrated by a two-step mechanism driven by the dynamic binding of transcriptional repressors (TRs) and activators, that is independent of transcription. In response to stress, promoter release of TAs at GR genes is a prerequisite for rapid reduction of H3K9ac, whereas binding of TRs is required to establish a hypo-acetylated, strongly repressed state.
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Affiliation(s)
- Sevil Zencir
- Department of Molecular and Cellular Biology, Université de Genève, 1211, Geneva, Switzerland
| | - Daniel Dilg
- Department of Molecular and Cellular Biology, Université de Genève, 1211, Geneva, Switzerland
| | - Maria Jessica Bruzzone
- Department of Molecular and Cellular Biology, Université de Genève, 1211, Geneva, Switzerland
| | - Françoise Stutz
- Department of Molecular and Cellular Biology, Université de Genève, 1211, Geneva, Switzerland
| | - Julien Soudet
- Department of Molecular and Cellular Biology, Université de Genève, 1211, Geneva, Switzerland
| | - David Shore
- Department of Molecular and Cellular Biology, Université de Genève, 1211, Geneva, Switzerland
| | - Benjamin Albert
- Department of Molecular and Cellular Biology, Université de Genève, 1211, Geneva, Switzerland
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Librais GMN, Jiang Y, Razzaq I, Brandl CJ, Shapiro RS, Lajoie P. Evolutionary diversity of the control of the azole response by Tra1 across yeast species. G3 (BETHESDA, MD.) 2024; 14:jkad250. [PMID: 37889998 PMCID: PMC10849324 DOI: 10.1093/g3journal/jkad250] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 09/04/2023] [Revised: 02/16/2023] [Accepted: 10/20/2023] [Indexed: 10/29/2023]
Abstract
Tra1 is an essential coactivator protein of the yeast SAGA and NuA4 acetyltransferase complexes that regulate gene expression through multiple mechanisms including the acetylation of histone proteins. Tra1 is a pseudokinase of the PIKK family characterized by a C-terminal PI3K domain with no known kinase activity. However, mutations of specific arginine residues to glutamine in the PI3K domains (an allele termed tra1Q3) result in reduced growth and increased sensitivity to multiple stresses. In the opportunistic fungal pathogen Candida albicans, the tra1Q3 allele reduces pathogenicity and increases sensitivity to the echinocandin antifungal drug caspofungin, which disrupts the fungal cell wall. Here, we found that compromised Tra1 function, in contrast to what is seen with caspofungin, increases tolerance to the azole class of antifungal drugs, which inhibits ergosterol synthesis. In C. albicans, tra1Q3 increases the expression of genes linked to azole resistance, such as ERG11 and CDR1. CDR1 encodes a multidrug ABC transporter associated with efflux of multiple xenobiotics, including azoles. Consequently, cells carrying tra1Q3 show reduced intracellular accumulation of fluconazole. In contrast, a tra1Q3 Saccharomyces cerevisiae strain displayed opposite phenotypes: decreased tolerance to azole, decreased expression of the efflux pump PDR5, and increased intracellular accumulation of fluconazole. Therefore, our data provide evidence that Tra1 differentially regulates the antifungal response across yeast species.
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Affiliation(s)
| | - Yuwei Jiang
- Department of Anatomy and Cell Biology, The University of Western Ontario, London, Ontario, N6A 5C1, Canada
| | - Iqra Razzaq
- Department of Molecular and Cellular Biology, University of Guelph, Guelph, Ontario N1G 2W1, Canada
| | - Christopher J Brandl
- Department of Biochemistry, The University of Western Ontario, London, Ontario, N6A 5C1, Canada
| | - Rebecca S Shapiro
- Department of Molecular and Cellular Biology, University of Guelph, Guelph, Ontario N1G 2W1, Canada
| | - Patrick Lajoie
- Department of Anatomy and Cell Biology, The University of Western Ontario, London, Ontario, N6A 5C1, Canada
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Huang YH, Lee YH, Lin CJ, Hsu LH, Chen YL. Deubiquitination module is critical for oxidative stress response and biofilm formation in Candida glabrata. Med Mycol 2023; 61:myad099. [PMID: 37844959 DOI: 10.1093/mmy/myad099] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/27/2023] [Revised: 09/02/2023] [Accepted: 10/11/2023] [Indexed: 10/18/2023] Open
Abstract
Candidiasis is one of the most important fungal diseases and generally refers to diseases of the skin or mucosal tissues caused by Candida species. Candida glabrata is an opportunistic human fungal pathogen. Infection with C. glabrata has significantly increased due to innate antifungal drug tolerance and the ability to adhere to mucocutaneous surfaces. Spt-Ada-Gcn5 acetyltransferase complex contains two different post-translational modifications, histone acetylation (HAT) module and deubiquitination (DUB) module, which are decisive in gene regulation and highly conserved in eukaryotes. Previous research in our laboratory found that the HAT module ADA2 could regulate C. glabrata oxidative stress tolerance, drug tolerance, cell wall integrity, and virulence. However, the roles of the DUB module that is comprised of UBP8, SGF11, SGF73, and SUS1 genes in those phenotypes are not yet understood. In this study, we found that DUB module genes UBP8, SGF11, and SUS1, but not SGF73 positively regulate histone H2B DUB. Furthermore, ubp8, sgf11, and sus1 mutants exhibited decreased biofilm formation and sensitivity to cell wall-perturbing agent sodium dodecyl sulfate and antifungal drug amphotericin B. In addition, the sgf73 mutant showed increased biofilm formation but was susceptible to oxidative stresses, antifungal drugs, and cell wall perturbing agents. The ubp8, sgf11, and sus1 mutants showed marginal hypovirulence, whereas the sgf73 mutant exhibited virulence similar to the wild type in a murine systemic infection model. In conclusion, the C. glabrata DUB module plays distinct roles in H2B ubiquitination, oxidative stress response, biofilm formation, cell wall integrity, and drug tolerance, but exhibits minor roles in virulence.
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Affiliation(s)
- Yue-Han Huang
- Department of Plant Pathology and Microbiology, National Taiwan University, 10617 Taipei, Taiwan
| | - Yi-Hang Lee
- Department of Plant Pathology and Microbiology, National Taiwan University, 10617 Taipei, Taiwan
| | - Chi-Jan Lin
- Institute of Molecular Biology, National Chung Hsing University, 40227 Taichung, Taiwan
| | - Li-Hang Hsu
- Department of Plant Pathology and Microbiology, National Taiwan University, 10617 Taipei, Taiwan
| | - Ying-Lien Chen
- Department of Plant Pathology and Microbiology, National Taiwan University, 10617 Taipei, Taiwan
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4
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Mahrik L, Stefanovie B, Maresova A, Princova J, Kolesar P, Lelkes E, Faux C, Helmlinger D, Prevorovsky M, Palecek JJ. The SAGA histone acetyltransferase module targets SMC5/6 to specific genes. Epigenetics Chromatin 2023; 16:6. [PMID: 36793083 PMCID: PMC9933293 DOI: 10.1186/s13072-023-00480-z] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/22/2022] [Accepted: 02/02/2023] [Indexed: 02/17/2023] Open
Abstract
BACKGROUND Structural Maintenance of Chromosomes (SMC) complexes are molecular machines driving chromatin organization at higher levels. In eukaryotes, three SMC complexes (cohesin, condensin and SMC5/6) play key roles in cohesion, condensation, replication, transcription and DNA repair. Their physical binding to DNA requires accessible chromatin. RESULTS We performed a genetic screen in fission yeast to identify novel factors required for SMC5/6 binding to DNA. We identified 79 genes of which histone acetyltransferases (HATs) were the most represented. Genetic and phenotypic analyses suggested a particularly strong functional relationship between the SMC5/6 and SAGA complexes. Furthermore, several SMC5/6 subunits physically interacted with SAGA HAT module components Gcn5 and Ada2. As Gcn5-dependent acetylation facilitates the accessibility of chromatin to DNA-repair proteins, we first analysed the formation of DNA-damage-induced SMC5/6 foci in the Δgcn5 mutant. The SMC5/6 foci formed normally in Δgcn5, suggesting SAGA-independent SMC5/6 localization to DNA-damaged sites. Next, we used Nse4-FLAG chromatin-immunoprecipitation (ChIP-seq) analysis in unchallenged cells to assess SMC5/6 distribution. A significant portion of SMC5/6 accumulated within gene regions in wild-type cells, which was reduced in Δgcn5 and Δada2 mutants. The drop in SMC5/6 levels was also observed in gcn5-E191Q acetyltransferase-dead mutant. CONCLUSION Our data show genetic and physical interactions between SMC5/6 and SAGA complexes. The ChIP-seq analysis suggests that SAGA HAT module targets SMC5/6 to specific gene regions and facilitates their accessibility for SMC5/6 loading.
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Affiliation(s)
- L Mahrik
- National Centre for Biomolecular Research, Faculty of Science, Masaryk University, Kotlarska 2, 61137, Brno, Czech Republic
- Mendel Centre for Plant Genomics and Proteomics, Central European Institute of Technology, Masaryk University, Kamenice 5, 62500, Brno, Czech Republic
| | - B Stefanovie
- National Centre for Biomolecular Research, Faculty of Science, Masaryk University, Kotlarska 2, 61137, Brno, Czech Republic
- Mendel Centre for Plant Genomics and Proteomics, Central European Institute of Technology, Masaryk University, Kamenice 5, 62500, Brno, Czech Republic
| | - A Maresova
- Department of Cell Biology, Faculty of Science, Charles University, Vinicna 7, 12800, Prague, Czech Republic
| | - J Princova
- Department of Cell Biology, Faculty of Science, Charles University, Vinicna 7, 12800, Prague, Czech Republic
| | - P Kolesar
- National Centre for Biomolecular Research, Faculty of Science, Masaryk University, Kotlarska 2, 61137, Brno, Czech Republic
| | - E Lelkes
- National Centre for Biomolecular Research, Faculty of Science, Masaryk University, Kotlarska 2, 61137, Brno, Czech Republic
| | - C Faux
- Centre de Recherche en Biologie Cellulaire de Montpellier, University of Montpellier, CNRS, 1919 Route de Mende, 34293, Montpellier Cedex 05, France
| | - D Helmlinger
- Centre de Recherche en Biologie Cellulaire de Montpellier, University of Montpellier, CNRS, 1919 Route de Mende, 34293, Montpellier Cedex 05, France
| | - M Prevorovsky
- Department of Cell Biology, Faculty of Science, Charles University, Vinicna 7, 12800, Prague, Czech Republic.
| | - J J Palecek
- National Centre for Biomolecular Research, Faculty of Science, Masaryk University, Kotlarska 2, 61137, Brno, Czech Republic.
- Mendel Centre for Plant Genomics and Proteomics, Central European Institute of Technology, Masaryk University, Kamenice 5, 62500, Brno, Czech Republic.
- National Centre for Biomolecular Research, Faculty of Science, Masaryk University, Kamenice 5, 62500, Brno, Czech Republic.
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5
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Lin CJ, Yang SY, Hsu LH, Yu SJ, Chen YL. The Gcn5-Ada2-Ada3 histone acetyltransferase module has divergent roles in pathogenesis of Candida glabrata. Med Mycol 2023; 61:myad004. [PMID: 36715154 DOI: 10.1093/mmy/myad004] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/05/2022] [Revised: 01/04/2023] [Accepted: 01/27/2023] [Indexed: 01/31/2023] Open
Abstract
Candida glabrata is an opportunistic fungal pathogen and the second most prevalent species isolated from candidiasis patients. C. glabrata has intrinsic tolerance to antifungal drugs and oxidative stresses and the ability to adhere to mucocutaneous surfaces. However, knowledge about the regulation of its virulence traits is limited. The Spt-Ada-Gcn5 acetyltransferase (SAGA) complex modulates gene transcription by histone acetylation through the histone acetyltransferase (HAT) module comprised of Gcn5-Ada2-Ada3. Previously, we showed that the ada2 mutant was hypervirulent but displayed decreased tolerance to antifungal drugs and cell wall perturbing agents. In this study, we further characterized the functions of Ada3 and Gcn5 in C. glabrata. We found that single, double, or triple deletions of the HAT module, as expected, resulted in a decreased level of acetylation on histone H3 lysine 9 (H3K9) and defective growth. These mutants were more susceptible to antifungal drugs, oxidative stresses, and cell wall perturbing agents compared with the wild-type. In addition, HAT module mutants exhibited enhanced agar invasion and upregulation of adhesin and proteases encoding genes, whereas the biofilm formation of those mutants was impaired. Interestingly, HAT module mutants exhibited enhanced induction of catalases (CTA1) expression upon treatment with H2O2 compared with the wild-type. Lastly, although ada3 and gcn5 exhibited marginal hypervirulence, the HAT double and triple mutants were hypervirulent in a murine model of candidiasis. In conclusion, the HAT module of the SAGA complex plays unique roles in H3K9 acetylation, drug tolerance, oxidative stress response, adherence, and virulence in C. glabrata.
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Affiliation(s)
- Chi-Jan Lin
- Institute of Molecular Biology, National Chung Hsing University, 40227 Taichung, Taiwan
- Department of Plant Pathology and Microbiology, National Taiwan University, 10617 Taipei, Taiwan
| | - Sheng-Yung Yang
- Department of Plant Pathology and Microbiology, National Taiwan University, 10617 Taipei, Taiwan
| | - Li-Hang Hsu
- Department of Plant Pathology and Microbiology, National Taiwan University, 10617 Taipei, Taiwan
| | - Shang-Jie Yu
- Department of Plant Pathology and Microbiology, National Taiwan University, 10617 Taipei, Taiwan
| | - Ying-Lien Chen
- Department of Plant Pathology and Microbiology, National Taiwan University, 10617 Taipei, Taiwan
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Princová J, Salat-Canela C, Daněk P, Marešová A, de Cubas L, Bähler J, Ayté J, Hidalgo E, Převorovský M. Perturbed fatty-acid metabolism is linked to localized chromatin hyperacetylation, increased stress-response gene expression and resistance to oxidative stress. PLoS Genet 2023; 19:e1010582. [PMID: 36626368 PMCID: PMC9870116 DOI: 10.1371/journal.pgen.1010582] [Citation(s) in RCA: 5] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/18/2022] [Revised: 01/23/2023] [Accepted: 12/19/2022] [Indexed: 01/11/2023] Open
Abstract
Oxidative stress is associated with cardiovascular and neurodegenerative diseases, diabetes, cancer, psychiatric disorders and aging. In order to counteract, eliminate and/or adapt to the sources of stress, cells possess elaborate stress-response mechanisms, which also operate at the level of regulating transcription. Interestingly, it is becoming apparent that the metabolic state of the cell and certain metabolites can directly control the epigenetic information and gene expression. In the fission yeast Schizosaccharomyces pombe, the conserved Sty1 stress-activated protein kinase cascade is the main pathway responding to most types of stresses, and regulates the transcription of hundreds of genes via the Atf1 transcription factor. Here we report that fission yeast cells defective in fatty acid synthesis (cbf11, mga2 and ACC/cut6 mutants; FAS inhibition) show increased expression of a subset of stress-response genes. This altered gene expression depends on Sty1-Atf1, the Pap1 transcription factor, and the Gcn5 and Mst1 histone acetyltransferases, is associated with increased acetylation of histone H3 at lysine 9 in the corresponding gene promoters, and results in increased cellular resistance to oxidative stress. We propose that changes in lipid metabolism can regulate the chromatin and transcription of specific stress-response genes, which in turn might help cells to maintain redox homeostasis.
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Affiliation(s)
- Jarmila Princová
- Laboratory of Microbial Genomics, Department of Cell Biology, Faculty of Science, Charles University, Prague, Czech Republic
| | - Clàudia Salat-Canela
- Oxidative Stress and Cell Cycle Group, Universitat Pompeu Fabra, C/Dr. Aiguader, Barcelona, Spain
| | - Petr Daněk
- Laboratory of Microbial Genomics, Department of Cell Biology, Faculty of Science, Charles University, Prague, Czech Republic
| | - Anna Marešová
- Laboratory of Microbial Genomics, Department of Cell Biology, Faculty of Science, Charles University, Prague, Czech Republic
| | - Laura de Cubas
- Oxidative Stress and Cell Cycle Group, Universitat Pompeu Fabra, C/Dr. Aiguader, Barcelona, Spain
| | - Jürg Bähler
- Institute of Healthy Ageing and Department of Genetics, Evolution & Environment, University College London, London, United Kingdom
| | - José Ayté
- Oxidative Stress and Cell Cycle Group, Universitat Pompeu Fabra, C/Dr. Aiguader, Barcelona, Spain
| | - Elena Hidalgo
- Oxidative Stress and Cell Cycle Group, Universitat Pompeu Fabra, C/Dr. Aiguader, Barcelona, Spain
| | - Martin Převorovský
- Laboratory of Microbial Genomics, Department of Cell Biology, Faculty of Science, Charles University, Prague, Czech Republic
- * E-mail:
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7
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Bari KA, Berg MD, Genereaux J, Brandl CJ, Lajoie P. Tra1 controls the transcriptional landscape of the aging cell. G3 (BETHESDA, MD.) 2022; 13:6782959. [PMID: 36315064 PMCID: PMC9836359 DOI: 10.1093/g3journal/jkac287] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 05/13/2022] [Accepted: 10/25/2022] [Indexed: 11/07/2022]
Abstract
Gene expression undergoes considerable changes during the aging process. The mechanisms regulating the transcriptional response to cellular aging remain poorly understood. Here, we employ the budding yeast Saccharomyces cerevisiae to better understand how organisms adapt their transcriptome to promote longevity. Chronological lifespan assays in yeast measure the survival of nondividing cells at stationary phase over time, providing insights into the aging process of postmitotic cells. Tra1 is an essential component of both the yeast Spt-Ada-Gcn5 acetyltransferase/Spt-Ada-Gcn5 acetyltransferase-like and nucleosome acetyltransferase of H4 complexes, where it recruits these complexes to acetylate histones at targeted promoters. Importantly, Tra1 regulates the transcriptional response to multiple stresses. To evaluate the role of Tra1 in chronological aging, we took advantage of a previously characterized mutant allele that carries mutations in the TRA1 PI3K domain (tra1Q3). We found that loss of functions associated with tra1Q3 sensitizes cells to growth media acidification and shortens lifespan. Transcriptional profiling reveals that genes differentially regulated by Tra1 during the aging process are enriched for components of the response to stress. Notably, expression of catalases (CTA1, CTT1) involved in hydrogen peroxide detoxification decreases in chronologically aged tra1Q3 cells. Consequently, they display increased sensitivity to oxidative stress. tra1Q3 cells are unable to grow on glycerol indicating a defect in mitochondria function. Aged tra1Q3 cells also display reduced expression of peroxisomal genes, exhibit decreased numbers of peroxisomes, and cannot grow on media containing oleate. Thus, Tra1 emerges as an important regulator of longevity in yeast via multiple mechanisms.
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Affiliation(s)
- Khaleda Afrin Bari
- Department of Anatomy and Cell Biology, The University of Western Ontario, London, ON N6A 5C1, Canada
| | - Matthew D Berg
- Present address for Matthew D Berg: Department of Genome Sciences, University of Washington, Seattle, WA, USA
| | - Julie Genereaux
- Department of Anatomy and Cell Biology, The University of Western Ontario, London, ON N6A 5C1, Canada,Department of Biochemistry, The University of Western Ontario, London, ON N6A 5C1, Canada
| | - Christopher J Brandl
- Department of Biochemistry, The University of Western Ontario, London, ON N6A 5C1, Canada
| | - Patrick Lajoie
- Corresponding author: Department of Anatomy and Cell Biology, The University of Western Ontario, London, ON N6A 5C1, Canada.
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8
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Detilleux D, Raynaud P, Pradet-Balade B, Helmlinger D. The TRRAP transcription cofactor represses interferon-stimulated genes in colorectal cancer cells. eLife 2022; 11:69705. [PMID: 35244540 PMCID: PMC8926402 DOI: 10.7554/elife.69705] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/23/2021] [Accepted: 03/03/2022] [Indexed: 11/30/2022] Open
Abstract
Transcription is essential for cells to respond to signaling cues and involves factors with multiple distinct activities. One such factor, TRRAP, functions as part of two large complexes, SAGA and TIP60, which have crucial roles during transcription activation. Structurally, TRRAP belongs to the phosphoinositide 3 kinase-related kinases (PIKK) family but is the only member classified as a pseudokinase. Recent studies established that a dedicated HSP90 co-chaperone, the triple T (TTT) complex, is essential for PIKK stabilization and activity. Here, using endogenous auxin-inducible degron alleles, we show that the TTT subunit TELO2 promotes TRRAP assembly into SAGA and TIP60 in human colorectal cancer cells (CRCs). Transcriptomic analysis revealed that TELO2 contributes to TRRAP regulatory roles in CRC cells, most notably of MYC target genes. Surprisingly, TELO2 and TRRAP depletion also induced the expression of type I interferon genes. Using a combination of nascent RNA, antibody-targeted chromatin profiling (CUT&RUN), ChIP, and kinetic analyses, we propose a model by which TRRAP directly represses the transcription of IRF9, which encodes a master regulator of interferon-stimulated genes. We have therefore uncovered an unexpected transcriptional repressor role for TRRAP, which we propose contributes to its tumorigenic activity.
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Affiliation(s)
| | - Peggy Raynaud
- CRBM, University of Montpellier, CNRS, Montpellier, France
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9
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Cohen A, Pataki E, Kupiec M, Weisman R. TOR complex 2 contributes to regulation of gene expression via inhibiting Gcn5 recruitment to subtelomeric and DNA replication stress genes. PLoS Genet 2022; 18:e1010061. [PMID: 35157728 PMCID: PMC8880919 DOI: 10.1371/journal.pgen.1010061] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/02/2021] [Revised: 02/25/2022] [Accepted: 01/28/2022] [Indexed: 11/18/2022] Open
Abstract
The fission yeast TOR complex 2 (TORC2) is required for gene silencing at subtelomeric regions and for the induction of gene transcription in response to DNA replication stress. Thus, TORC2 affects transcription regulation both negatively and positively. Whether these two TORC2-dependent functions share a common molecular mechanism is currently unknown. Here, we show that Gad8 physically interacts with proteins that regulate transcription, including subunits of the Spt-Ada-Gcn5-acetyltransferase (SAGA) complex and the BET bromodomain protein Bdf2. We demonstrate that in the absence of TORC2, Gcn5, the histone acetyltransferase subunit of SAGA, accumulates at subtelomeric genes and at non-induced promoters of DNA replication genes. Remarkably, the loss of Gcn5 in TORC2 mutant cells restores gene silencing as well as transcriptional induction in response to DNA replication stress. Loss of Bdf2 alleviates excess of Gcn5 binding in TORC2 mutant cells and also rescues the aberrant regulation of transcription in these cells. Furthermore, the loss of either SAGA or Bdf2 suppresses the sensitivity of TORC2 mutant cells to a variety of stresses, including DNA replication, DNA damage, temperature and nutrient stresses. We suggest a role of TORC2 in transcriptional regulation that is critical for gene silencing and gene induction in response to stress and involves the binding of Gcn5 to the chromatin.
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Affiliation(s)
- Adiel Cohen
- Department of Natural and Life Sciences, The Open University of Israel, Ra’anana, Israel
| | - Emese Pataki
- Department of Natural and Life Sciences, The Open University of Israel, Ra’anana, Israel
| | - Martin Kupiec
- The Shmunis School of Biomedicine & Cancer Research, Tel Aviv University, Tel Aviv, Israel
| | - Ronit Weisman
- Department of Natural and Life Sciences, The Open University of Israel, Ra’anana, Israel
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10
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Nikolenko JV, Vdovina YA, Fefelova EI, Glukhova AA, Nabirochkina EN, Kopytova DV. The SAGA Deubiquitinilation (DUB) Module Participates in Pol III-Dependent Transcription. Mol Biol 2021. [DOI: 10.1134/s0026893321020278] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/23/2022]
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11
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Yin BK, Wang ZQ. Beyond HAT Adaptor: TRRAP Liaisons with Sp1-Mediated Transcription. Int J Mol Sci 2021; 22:12445. [PMID: 34830324 PMCID: PMC8625110 DOI: 10.3390/ijms222212445] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/27/2021] [Revised: 11/09/2021] [Accepted: 11/15/2021] [Indexed: 12/19/2022] Open
Abstract
The members of the phosphatidylinositol 3-kinase-related kinase (PIKK) family play vital roles in multiple biological processes, including DNA damage response, metabolism, cell growth, mRNA decay, and transcription. TRRAP, as the only member lacking the enzymatic activity in this family, is an adaptor protein for several histone acetyltransferase (HAT) complexes and a scaffold protein for multiple transcription factors. TRRAP has been demonstrated to regulate various cellular functions in cell cycle progression, cell stemness maintenance and differentiation, as well as neural homeostasis. TRRAP is known to be an important orchestrator of many molecular machineries in gene transcription by modulating the activity of some key transcription factors, including E2F1, c-Myc, p53, and recently, Sp1. This review summarizes the biological and biochemical studies on the action mode of TRRAP together with the transcription factors, focusing on how TRRAP-HAT mediates the transactivation of Sp1-governing biological processes, including neurodegeneration.
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Affiliation(s)
- Bo-Kun Yin
- Leibniz Institute on Aging-Fritz Lipmann Institute (FLI), 07745 Jena, Germany;
| | - Zhao-Qi Wang
- Leibniz Institute on Aging-Fritz Lipmann Institute (FLI), 07745 Jena, Germany;
- Faculty of Biological Sciences, Friedrich Schiller University Jena, 07743 Jena, Germany
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12
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Toullec D, Elías-Villalobos A, Faux C, Noly A, Lledo G, Séveno M, Helmlinger D. The Hsp90 cochaperone TTT promotes cotranslational maturation of PIKKs prior to complex assembly. Cell Rep 2021; 37:109867. [PMID: 34686329 DOI: 10.1016/j.celrep.2021.109867] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/16/2020] [Revised: 07/30/2021] [Accepted: 09/30/2021] [Indexed: 01/28/2023] Open
Abstract
Phosphatidylinositol 3-kinase-related kinases (PIKKs) are a family of kinases that control fundamental processes, including cell growth, DNA damage repair, and gene expression. Although their regulation and activities are well characterized, little is known about how PIKKs fold and assemble into active complexes. Previous work has identified a heat shock protein 90 (Hsp90) cochaperone, the TTT complex, that specifically stabilizes PIKKs. Here, we describe a mechanism by which TTT promotes their de novo maturation in fission yeast. We show that TTT recognizes newly synthesized PIKKs during translation. Although PIKKs form multimeric complexes, we find that they do not engage in cotranslational assembly with their partners. Rather, our findings suggest a model by which TTT protects nascent PIKK polypeptides from misfolding and degradation because PIKKs acquire their native state after translation is terminated. Thus, PIKK maturation and assembly are temporally segregated, suggesting that the biogenesis of large complexes requires both dedicated chaperones and cotranslational interactions between subunits.
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Affiliation(s)
- Damien Toullec
- CRBM, University of Montpellier, CNRS, Montpellier, France
| | | | - Céline Faux
- CRBM, University of Montpellier, CNRS, Montpellier, France
| | - Ambre Noly
- CRBM, University of Montpellier, CNRS, Montpellier, France
| | | | - Martial Séveno
- BioCampus Montpellier, University of Montpellier, CNRS, INSERM, Montpellier, France
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13
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Razzaq I, Berg MD, Jiang Y, Genereaux J, Uthayakumar D, Kim GH, Agyare-Tabbi M, Halder V, Brandl CJ, Lajoie P, Shapiro RS. The SAGA and NuA4 component Tra1 regulates Candida albicans drug resistance and pathogenesis. Genetics 2021; 219:iyab131. [PMID: 34849885 PMCID: PMC8633099 DOI: 10.1093/genetics/iyab131] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/29/2021] [Accepted: 08/02/2021] [Indexed: 11/14/2022] Open
Abstract
Candida albicans is the most common cause of death from fungal infections. The emergence of resistant strains reducing the efficacy of first-line therapy with echinocandins, such as caspofungin calls for the identification of alternative therapeutic strategies. Tra1 is an essential component of the SAGA and NuA4 transcriptional co-activator complexes. As a PIKK family member, Tra1 is characterized by a C-terminal phosphoinositide 3-kinase domain. In Saccharomyces cerevisiae, the assembly and function of SAGA and NuA4 are compromised by a Tra1 variant (Tra1Q3) with three arginine residues in the putative ATP-binding cleft changed to glutamine. Whole transcriptome analysis of the S. cerevisiae tra1Q3 strain highlights Tra1's role in global transcription, stress response, and cell wall integrity. As a result, tra1Q3 increases susceptibility to multiple stressors, including caspofungin. Moreover, the same tra1Q3 allele in the pathogenic yeast C. albicans causes similar phenotypes, suggesting that Tra1 broadly mediates the antifungal response across yeast species. Transcriptional profiling in C. albicans identified 68 genes that were differentially expressed when the tra1Q3 strain was treated with caspofungin, as compared to gene expression changes induced by either tra1Q3 or caspofungin alone. Included in this set were genes involved in cell wall maintenance, adhesion, and filamentous growth. Indeed, the tra1Q3 allele reduces filamentation and other pathogenesis traits in C. albicans. Thus, Tra1 emerges as a promising therapeutic target for fungal infections.
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Affiliation(s)
- Iqra Razzaq
- Department of Molecular and Cellular Biology, University of Guelph, Guelph, ON N1G2W1, Canada
| | - Matthew D Berg
- Department of Biochemistry, The University of Western Ontario, London, ON N6A 5C1, Canada
| | - Yuwei Jiang
- Department of Anatomy and Cell Biology, The University of Western Ontario, London, ON N6A 5C1, Canada
| | - Julie Genereaux
- Department of Biochemistry, The University of Western Ontario, London, ON N6A 5C1, Canada
- Department of Anatomy and Cell Biology, The University of Western Ontario, London, ON N6A 5C1, Canada
| | - Deeva Uthayakumar
- Department of Molecular and Cellular Biology, University of Guelph, Guelph, ON N1G2W1, Canada
| | - Grace H Kim
- Department of Molecular and Cellular Biology, University of Guelph, Guelph, ON N1G2W1, Canada
| | - Michelle Agyare-Tabbi
- Department of Molecular and Cellular Biology, University of Guelph, Guelph, ON N1G2W1, Canada
| | - Viola Halder
- Department of Molecular and Cellular Biology, University of Guelph, Guelph, ON N1G2W1, Canada
| | - Christopher J Brandl
- Department of Biochemistry, The University of Western Ontario, London, ON N6A 5C1, Canada
| | - Patrick Lajoie
- Department of Anatomy and Cell Biology, The University of Western Ontario, London, ON N6A 5C1, Canada
| | - Rebecca S Shapiro
- Department of Molecular and Cellular Biology, University of Guelph, Guelph, ON N1G2W1, Canada
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14
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Fischer V, Plassard D, Ye T, Reina-San-Martin B, Stierle M, Tora L, Devys D. The related coactivator complexes SAGA and ATAC control embryonic stem cell self-renewal through acetyltransferase-independent mechanisms. Cell Rep 2021; 36:109598. [PMID: 34433046 PMCID: PMC8430043 DOI: 10.1016/j.celrep.2021.109598] [Citation(s) in RCA: 13] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/27/2021] [Revised: 06/17/2021] [Accepted: 08/03/2021] [Indexed: 01/10/2023] Open
Abstract
SAGA (Spt-Ada-Gcn5 acetyltransferase) and ATAC (Ada-two-A-containing) are two related coactivator complexes, sharing the same histone acetyltransferase (HAT) subunit. The HAT activities of SAGA and ATAC are required for metazoan development, but the role of these complexes in RNA polymerase II transcription is less understood. To determine whether SAGA and ATAC have redundant or specific functions, we compare the effects of HAT inactivation in each complex with that of inactivation of either SAGA or ATAC core subunits in mouse embryonic stem cells (ESCs). We show that core subunits of SAGA or ATAC are required for complex assembly and mouse ESC growth and self-renewal. Surprisingly, depletion of HAT module subunits causes a global decrease in histone H3K9 acetylation, but does not result in significant phenotypic or transcriptional defects. Thus, our results indicate that SAGA and ATAC are differentially required for self-renewal of mouse ESCs by regulating transcription through different pathways in a HAT-independent manner.
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Affiliation(s)
- Veronique Fischer
- Institut de Génétique et de Biologie Moléculaire et Cellulaire, 67404 Illkirch Cedex, France; Centre National de la Recherche Scientifique, UMR7104, 67404 Illkirch Cedex, France; Institut National de la Santé et de la Recherche Médicale, U964, 67404 Illkirch Cedex, France; Université de Strasbourg, 67000 Strasbourg, France
| | - Damien Plassard
- Institut de Génétique et de Biologie Moléculaire et Cellulaire, 67404 Illkirch Cedex, France; Centre National de la Recherche Scientifique, UMR7104, 67404 Illkirch Cedex, France; Institut National de la Santé et de la Recherche Médicale, U964, 67404 Illkirch Cedex, France; Université de Strasbourg, 67000 Strasbourg, France; Plateforme GenomEast, infrastructure France Génomique, Illkirch, France
| | - Tao Ye
- Institut de Génétique et de Biologie Moléculaire et Cellulaire, 67404 Illkirch Cedex, France; Centre National de la Recherche Scientifique, UMR7104, 67404 Illkirch Cedex, France; Institut National de la Santé et de la Recherche Médicale, U964, 67404 Illkirch Cedex, France; Université de Strasbourg, 67000 Strasbourg, France; Plateforme GenomEast, infrastructure France Génomique, Illkirch, France
| | - Bernardo Reina-San-Martin
- Institut de Génétique et de Biologie Moléculaire et Cellulaire, 67404 Illkirch Cedex, France; Centre National de la Recherche Scientifique, UMR7104, 67404 Illkirch Cedex, France; Institut National de la Santé et de la Recherche Médicale, U964, 67404 Illkirch Cedex, France; Université de Strasbourg, 67000 Strasbourg, France
| | - Matthieu Stierle
- Institut de Génétique et de Biologie Moléculaire et Cellulaire, 67404 Illkirch Cedex, France; Centre National de la Recherche Scientifique, UMR7104, 67404 Illkirch Cedex, France; Institut National de la Santé et de la Recherche Médicale, U964, 67404 Illkirch Cedex, France; Université de Strasbourg, 67000 Strasbourg, France
| | - Laszlo Tora
- Institut de Génétique et de Biologie Moléculaire et Cellulaire, 67404 Illkirch Cedex, France; Centre National de la Recherche Scientifique, UMR7104, 67404 Illkirch Cedex, France; Institut National de la Santé et de la Recherche Médicale, U964, 67404 Illkirch Cedex, France; Université de Strasbourg, 67000 Strasbourg, France
| | - Didier Devys
- Institut de Génétique et de Biologie Moléculaire et Cellulaire, 67404 Illkirch Cedex, France; Centre National de la Recherche Scientifique, UMR7104, 67404 Illkirch Cedex, France; Institut National de la Santé et de la Recherche Médicale, U964, 67404 Illkirch Cedex, France; Université de Strasbourg, 67000 Strasbourg, France.
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15
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Chen YJC, Dent SYR. Conservation and diversity of the eukaryotic SAGA coactivator complex across kingdoms. Epigenetics Chromatin 2021; 14:26. [PMID: 34112237 PMCID: PMC8194025 DOI: 10.1186/s13072-021-00402-x] [Citation(s) in RCA: 17] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/22/2021] [Accepted: 06/03/2021] [Indexed: 12/27/2022] Open
Abstract
The SAGA complex is an evolutionarily conserved transcriptional coactivator that regulates gene expression through its histone acetyltransferase and deubiquitylase activities, recognition of specific histone modifications, and interactions with transcription factors. Multiple lines of evidence indicate the existence of distinct variants of SAGA among organisms as well as within a species, permitting diverse functions to dynamically regulate cellular pathways. Our co-expression analysis of genes encoding human SAGA components showed enrichment in reproductive organs, brain tissues and the skeletal muscle, which corresponds to their established roles in developmental programs, emerging roles in neurodegenerative diseases, and understudied functions in specific cell types. SAGA subunits modulate growth, development and response to various stresses from yeast to plants and metazoans. In metazoans, SAGA further participates in the regulation of differentiation and maturation of both innate and adaptive immune cells, and is associated with initiation and progression of diseases including a broad range of cancers. The evolutionary conservation of SAGA highlights its indispensable role in eukaryotic life, thus deciphering the mechanisms of action of SAGA is key to understanding fundamental biological processes throughout evolution. To illuminate the diversity and conservation of this essential complex, here we discuss variations in composition, essentiality and co-expression of component genes, and its prominent functions across Fungi, Plantae and Animalia kingdoms.
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Affiliation(s)
- Ying-Jiun C Chen
- Department of Epigenetics and Molecular Carcinogenesis, The University of Texas MD Anderson Cancer Center, Smithville, TX, USA
- The Center for Cancer Epigenetics, The University of Texas MD Anderson Cancer Center, Houston, TX, USA
| | - Sharon Y R Dent
- Department of Epigenetics and Molecular Carcinogenesis, The University of Texas MD Anderson Cancer Center, Smithville, TX, USA.
- The Center for Cancer Epigenetics, The University of Texas MD Anderson Cancer Center, Houston, TX, USA.
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16
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Badmos H, Cobbe N, Campbell A, Jackson R, Bennett D. Drosophila USP22/nonstop polarizes the actin cytoskeleton during collective border cell migration. J Cell Biol 2021; 220:212101. [PMID: 33988679 PMCID: PMC8129793 DOI: 10.1083/jcb.202007005] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/02/2020] [Revised: 02/06/2021] [Accepted: 04/23/2021] [Indexed: 01/04/2023] Open
Abstract
Polarization of the actin cytoskeleton is vital for the collective migration of cells in vivo. During invasive border cell migration in Drosophila, actin polarization is directly controlled by the Hippo signaling complex, which resides at contacts between border cells in the cluster. Here, we identify, in a genetic screen for deubiquitinating enzymes involved in border cell migration, an essential role for nonstop/USP22 in the expression of Hippo pathway components expanded and merlin. Loss of nonstop function consequently leads to a redistribution of F-actin and the polarity determinant Crumbs, loss of polarized actin protrusions, and tumbling of the border cell cluster. Nonstop is a component of the Spt-Ada-Gcn5-acetyltransferase (SAGA) transcriptional coactivator complex, but SAGA’s histone acetyltransferase module, which does not bind to expanded or merlin, is dispensable for migration. Taken together, our results uncover novel roles for SAGA-independent nonstop/USP22 in collective cell migration, which may help guide studies in other systems where USP22 is necessary for cell motility and invasion.
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Affiliation(s)
- Hammed Badmos
- Department of Biochemistry, Institute of Integrative Biology, University of Liverpool, Liverpool, UK.,Department of Molecular Physiology and Cell Signalling, Institute of Systems, Molecular and Integrative Biology, University of Liverpool, Liverpool, UK
| | - Neville Cobbe
- Department of Biochemistry, Institute of Integrative Biology, University of Liverpool, Liverpool, UK
| | - Amy Campbell
- Department of Biochemistry, Institute of Integrative Biology, University of Liverpool, Liverpool, UK
| | - Richard Jackson
- Liverpool Clinical Trials Centre, University of Liverpool, Liverpool, UK
| | - Daimark Bennett
- Department of Biochemistry, Institute of Integrative Biology, University of Liverpool, Liverpool, UK.,Department of Molecular Physiology and Cell Signalling, Institute of Systems, Molecular and Integrative Biology, University of Liverpool, Liverpool, UK
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17
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Soffers JHM, Workman JL. The SAGA chromatin-modifying complex: the sum of its parts is greater than the whole. Genes Dev 2021; 34:1287-1303. [PMID: 33004486 PMCID: PMC7528701 DOI: 10.1101/gad.341156.120] [Citation(s) in RCA: 33] [Impact Index Per Article: 8.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
Abstract
In this review, Soffers and Workman discuss the initial discovery of the canonical SAGA complex, the subsequent studies that have shaped our view on the internal organization of its subunits into modules, and the latest structural work that visualizes the modules and provides insights into their function. There are many large protein complexes involved in transcription in a chromatin context. However, recent studies on the SAGA coactivator complex are generating new paradigms for how the components of these complexes function, both independently and in concert. This review highlights the initial discovery of the canonical SAGA complex 23 years ago, our evolving understanding of its modular structure and the relevance of its modular nature for its coactivator function in gene regulation.
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Affiliation(s)
- Jelly H M Soffers
- Stowers Institute for Medical Research, Kansas City, Missouri 64110, USA
| | - Jerry L Workman
- Stowers Institute for Medical Research, Kansas City, Missouri 64110, USA
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18
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High-Throughput Flow Cytometry Combined with Genetic Analysis Brings New Insights into the Understanding of Chromatin Regulation of Cellular Quiescence. Int J Mol Sci 2020; 21:ijms21239022. [PMID: 33260998 PMCID: PMC7729564 DOI: 10.3390/ijms21239022] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/26/2020] [Revised: 11/19/2020] [Accepted: 11/23/2020] [Indexed: 11/21/2022] Open
Abstract
Cellular quiescence is a reversible differentiation state when cells are changing the gene expression program to reduce metabolic functions and adapt to a new cellular environment. When fission yeast cells are deprived of nitrogen in the absence of any mating partner, cells can reversibly arrest in a differentiated G0-like cellular state, called quiescence. This change is accompanied by a marked alteration of nuclear organization and a global reduction of transcription. Using high-throughput flow cytometry combined with genetic analysis, we describe the results of a comprehensive screen for genes encoding chromatin components and regulators that are required for the entry and the maintenance of cellular quiescence. We show that the histone acetylase and deacetylase complexes, SAGA and Rpd3, have key roles both for G0 entry and survival during quiescence. We reveal a novel function for the Ino80 nucleosome remodeling complex in cellular quiescence. Finally, we demonstrate that components of the MRN complex, Rad3, the nonhomologous end-joining, and nucleotide excision DNA repair pathways are essential for viability in G0.
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19
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What do the structures of GCN5-containing complexes teach us about their function? BIOCHIMICA ET BIOPHYSICA ACTA-GENE REGULATORY MECHANISMS 2020; 1864:194614. [PMID: 32739556 DOI: 10.1016/j.bbagrm.2020.194614] [Citation(s) in RCA: 25] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [Subscribe] [Scholar Register] [Received: 05/15/2020] [Revised: 07/28/2020] [Accepted: 07/29/2020] [Indexed: 12/20/2022]
Abstract
Transcription initiation is a major regulatory step in eukaryotic gene expression. It involves the assembly of general transcription factors and RNA polymerase II into a functional pre-initiation complex at core promoters. The degree of chromatin compaction controls the accessibility of the transcription machinery to template DNA. Co-activators have critical roles in this process by actively regulating chromatin accessibility. Many transcriptional coactivators are multisubunit complexes, organized into distinct structural and functional modules and carrying multiple regulatory activities. The first nuclear histone acetyltransferase (HAT) characterized was General Control Non-derepressible 5 (Gcn5). Gcn5 was subsequently identified as a subunit of the HAT module of the Spt-Ada-Gcn5-acetyltransferase (SAGA) complex, which is an experimental paradigm for multifunctional co-activators. We know today that Gcn5 is the catalytic subunit of multiple distinct co-activator complexes with specific functions. In this review, we summarize recent advances in the structure of Gcn5-containing co-activator complexes, most notably SAGA, and discuss how these new structural insights contribute to better understand their functions.
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20
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Torres-Zelada EF, Weake VM. The Gcn5 complexes in Drosophila as a model for metazoa. BIOCHIMICA ET BIOPHYSICA ACTA-GENE REGULATORY MECHANISMS 2020; 1864:194610. [PMID: 32735945 DOI: 10.1016/j.bbagrm.2020.194610] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Subscribe] [Scholar Register] [Received: 04/30/2020] [Revised: 07/21/2020] [Accepted: 07/22/2020] [Indexed: 01/14/2023]
Abstract
The histone acetyltransferase Gcn5 is conserved throughout eukaryotes where it functions as part of large multi-subunit transcriptional coactivator complexes that stimulate gene expression. Here, we describe how studies in the model insect Drosophila melanogaster have provided insight into the essential roles played by Gcn5 in the development of multicellular organisms. We outline the composition and activity of the four different Gcn5 complexes in Drosophila: the Spt-Ada-Gcn5 Acetyltransferase (SAGA), Ada2a-containing (ATAC), Ada2/Gcn5/Ada3 transcription activator (ADA), and Chiffon Histone Acetyltransferase (CHAT) complexes. Whereas the SAGA and ADA complexes are also present in the yeast Saccharomyces cerevisiae, ATAC has only been identified in other metazoa such as humans, and the CHAT complex appears to be unique to insects. Each of these Gcn5 complexes is nucleated by unique Ada2 homologs or splice isoforms that share conserved N-terminal domains, and differ only in their C-terminal domains. We describe the common and specialized developmental functions of each Gcn5 complex based on phenotypic analysis of mutant flies. In addition, we outline how gene expression studies in mutant flies have shed light on the different biological roles of each complex. Together, these studies highlight the key role that Drosophila has played in understanding the expanded biological function of Gcn5 in multicellular eukaryotes.
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Affiliation(s)
| | - Vikki M Weake
- Department of Biochemistry, Purdue University, West Lafayette, IN 47907, USA; Purdue University Center for Cancer Research, Purdue University, West Lafayette, IN 47907, USA.
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21
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Nuño-Cabanes C, Rodríguez-Navarro S. The promiscuity of the SAGA complex subunits: Multifunctional or moonlighting proteins? BIOCHIMICA ET BIOPHYSICA ACTA-GENE REGULATORY MECHANISMS 2020; 1864:194607. [PMID: 32712338 DOI: 10.1016/j.bbagrm.2020.194607] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Subscribe] [Scholar Register] [Received: 04/27/2020] [Revised: 07/09/2020] [Accepted: 07/13/2020] [Indexed: 12/15/2022]
Abstract
Gene expression, the decoding of DNA information into accessible instructions for protein synthesis, is a complex process in which multiple steps, including transcription, mRNA processing and mRNA export, are regulated by different factors. One of the first steps in this process involves chemical and structural changes in chromatin to allow transcription. For such changes to occur, histone tail and DNA epigenetic modifications foster the binding of transcription factors to promoter regions. The SAGA coactivator complex plays a crucial role in this process by mediating histone acetylation through Gcn5, and histone deubiquitination through Ubp8 enzymes. However, most SAGA subunits interact physically with other proteins beyond the SAGA complex. These interactions could represent SAGA-independent functions or a mechanism to widen SAGA multifunctionality. Among the different mechanisms to perform more than one function, protein moonlighting defines unrelated molecular activities for the same polypeptide sequence. Unlike pleiotropy, where a single gene can affect different phenotypes, moonlighting necessarily involves separate functions of a protein at the molecular level. In this review we describe in detail some of the alternative physical interactions of several SAGA subunits. In some cases, the alternative role constitutes a clear moonlighting function, whereas in most of them the lack of molecular evidence means that we can only define these interactions as promiscuous that require further work to verify if these are moonlighting functions.
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Affiliation(s)
- Carme Nuño-Cabanes
- Gene Expression and RNA Metabolism Laboratory, Instituto de Biomedicina de Valencia (CSIC), Jaume Roig, 11, E-46010 Valencia, Spain
| | - Susana Rodríguez-Navarro
- Gene Expression and RNA Metabolism Laboratory, Instituto de Biomedicina de Valencia (CSIC), Jaume Roig, 11, E-46010 Valencia, Spain.
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22
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Cheon Y, Kim H, Park K, Kim M, Lee D. Dynamic modules of the coactivator SAGA in eukaryotic transcription. Exp Mol Med 2020; 52:991-1003. [PMID: 32616828 PMCID: PMC8080568 DOI: 10.1038/s12276-020-0463-4] [Citation(s) in RCA: 29] [Impact Index Per Article: 5.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/20/2020] [Revised: 05/15/2020] [Accepted: 05/19/2020] [Indexed: 02/08/2023] Open
Abstract
SAGA (Spt-Ada-Gcn5 acetyltransferase) is a highly conserved transcriptional coactivator that consists of four functionally independent modules. Its two distinct enzymatic activities, histone acetylation and deubiquitylation, establish specific epigenetic patterns on chromatin and thereby regulate gene expression. Whereas earlier studies emphasized the importance of SAGA in regulating global transcription, more recent reports have indicated that SAGA is involved in other aspects of gene expression and thus plays a more comprehensive role in regulating the overall process. Here, we discuss recent structural and functional studies of each SAGA module and compare the subunit compositions of SAGA with related complexes in yeast and metazoans. We discuss the regulatory role of the SAGA deubiquitylating module (DUBm) in mRNA surveillance and export, and in transcription initiation and elongation. The findings suggest that SAGA plays numerous roles in multiple stages of transcription. Further, we describe how SAGA is related to human disease. Overall, in this report, we illustrate the newly revealed understanding of SAGA in transcription regulation and disease implications for fine-tuning gene expression. A protein that helps add epigenetic information to genome, SAGA, controls many aspects of gene activation, potentially making it a target for cancer therapies. To fit inside the tiny cell nucleus, the genome is tightly packaged, and genes must be unpacked before they can be activated. Known to be important in genome opening, SAGA has now been shown to also play many roles in gene activation. Daeyoup Lee at the KAIST, Daejeon, South Korea, and co-workers have reviewed recent discoveries about SAGA’s structure, function, and roles in disease. They report that SAGA’s complex (19 subunits organized into four modules) allows it to play so many roles, genome opening, initiating transcription, and efficiently exporting mRNAs. Its master role means that malfunction of SAGA may be linked to many diseases.
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Affiliation(s)
- Youngseo Cheon
- Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Daejeon, 34141, South Korea
| | - Harim Kim
- Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Daejeon, 34141, South Korea
| | - Kyubin Park
- Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Daejeon, 34141, South Korea
| | - Minhoo Kim
- Leonard Davis School of Gerontology, University of Southern California, Los Angeles, CA, 90089, USA
| | - Daeyoup Lee
- Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Daejeon, 34141, South Korea.
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23
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Donczew R, Warfield L, Pacheco D, Erijman A, Hahn S. Two roles for the yeast transcription coactivator SAGA and a set of genes redundantly regulated by TFIID and SAGA. eLife 2020; 9:e50109. [PMID: 31913117 PMCID: PMC6977968 DOI: 10.7554/elife.50109] [Citation(s) in RCA: 60] [Impact Index Per Article: 12.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/11/2019] [Accepted: 01/07/2020] [Indexed: 12/31/2022] Open
Abstract
Deletions within genes coding for subunits of the transcription coactivator SAGA caused strong genome-wide defects in transcription and SAGA-mediated chromatin modifications. In contrast, rapid SAGA depletion produced only modest transcription defects at 13% of protein-coding genes - genes that are generally more sensitive to rapid TFIID depletion. However, transcription of these 'coactivator-redundant' genes is strongly affected by rapid depletion of both factors, showing the overlapping functions of TFIID and SAGA at this gene set. We suggest that this overlapping function is linked to TBP-DNA recruitment. The remaining 87% of expressed genes that we term 'TFIID-dependent' are highly sensitive to rapid TFIID depletion and insensitive to rapid SAGA depletion. Genome-wide mapping of SAGA and TFIID found binding of both factors at many genes independent of gene class. Promoter analysis suggests that the distinction between the gene classes is due to multiple components rather than any single regulatory factor or promoter sequence motif.
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Affiliation(s)
- Rafal Donczew
- Fred Hutchinson Cancer Research CenterSeattleUnited States
| | - Linda Warfield
- Fred Hutchinson Cancer Research CenterSeattleUnited States
| | - Derek Pacheco
- Fred Hutchinson Cancer Research CenterSeattleUnited States
| | - Ariel Erijman
- Fred Hutchinson Cancer Research CenterSeattleUnited States
| | - Steven Hahn
- Fred Hutchinson Cancer Research CenterSeattleUnited States
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24
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Bu B, Chen L, Zheng L, He W, Zhang L. Nipped-A regulates the Drosophila circadian clock via histone deubiquitination. EMBO J 2020; 39:e101259. [PMID: 31538360 PMCID: PMC6939192 DOI: 10.15252/embj.2018101259] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/29/2018] [Revised: 08/25/2019] [Accepted: 08/28/2019] [Indexed: 12/19/2022] Open
Abstract
Psychiatric diseases are often accompanied by circadian disruptions, but the molecular underpinnings remain largely unclear. To address this, we screened genes that have been previously reported to be associated with psychiatric diseases and found that TRRAP, a gene associated with schizophrenia, is involved in circadian rhythm regulation. Knocking down Nipped-A, the Drosophila homolog of human TRRAP, leads to lengthened period of locomotor rhythms in flies. Molecular analysis demonstrates that NIPPED-A sets the pace of the clock by increasing the mRNA and protein levels of core clock genes timeless (tim) and Par domain protein 1ε (Pdp1ε). Furthermore, we found that NIPPED-A promotes the transcription of tim and Pdp1ε possibly by facilitating deubiquitination of histone H2B via the deubiquitination module of the transcription co-activator Spt-Ada-Gcn5 acetyltransferase complex. Taken together, these findings reveal a novel role for NIPPED-A in epigenetic regulation of the clock.
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Affiliation(s)
- Bei Bu
- Key Laboratory of Molecular Biophysics of Ministry of EducationCollege of Life Science and TechnologyHuazhong University of Science and TechnologyWuhanHubeiChina
- Henan Key Laboratory of Reproduction and GeneticsCenter for Reproductive MedicineThe First Affiliated Hospital of Zhengzhou UniversityZhengzhouHenanChina
| | - Lixia Chen
- Key Laboratory of Molecular Biophysics of Ministry of EducationCollege of Life Science and TechnologyHuazhong University of Science and TechnologyWuhanHubeiChina
| | - Liubin Zheng
- Key Laboratory of Molecular Biophysics of Ministry of EducationCollege of Life Science and TechnologyHuazhong University of Science and TechnologyWuhanHubeiChina
| | - Weiwei He
- Key Laboratory of Molecular Biophysics of Ministry of EducationCollege of Life Science and TechnologyHuazhong University of Science and TechnologyWuhanHubeiChina
| | - Luoying Zhang
- Key Laboratory of Molecular Biophysics of Ministry of EducationCollege of Life Science and TechnologyHuazhong University of Science and TechnologyWuhanHubeiChina
- Institute of Brain ResearchHuazhong University of Science and TechnologyWuhanHubeiChina
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25
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Espinosa-Cores L, Bouza-Morcillo L, Barrero-Gil J, Jiménez-Suárez V, Lázaro A, Piqueras R, Jarillo JA, Piñeiro M. Insights Into the Function of the NuA4 Complex in Plants. FRONTIERS IN PLANT SCIENCE 2020; 11:125. [PMID: 32153620 PMCID: PMC7047200 DOI: 10.3389/fpls.2020.00125] [Citation(s) in RCA: 21] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 11/22/2019] [Accepted: 01/28/2020] [Indexed: 05/14/2023]
Abstract
Chromatin remodeling plays a key role in the establishment and maintenance of gene expression patterns essential for plant development and responses to environmental factors. Post-translational modification of histones, including acetylation, is one of the most relevant chromatin remodeling mechanisms that operate in eukaryotic cells. Histone acetylation is an evolutionarily conserved chromatin signature commonly associated with transcriptional activation. Histone acetylation levels are tightly regulated through the antagonistic activity of histone acetyltransferases (HATs) and histone deacetylases (HDACs). In plants, different families of HATs are present, including the MYST family, which comprises homologs of the catalytic subunit of the Nucleosome Acetyltransferase of H4 (NuA4) complex in yeast. This complex mediates acetylation of histones H4, H2A, and H2A.Z, and is involved in transcriptional regulation, heterochromatin silencing, cell cycle progression, and DNA repair in yeast. In Arabidopsis and, other plant species, homologs for most of the yeast NuA4 subunits are present and although the existence of this complex has not been demonstrated yet, compelling evidence supports the notion that this type of HAT complex functions from mosses to angiosperms. Recent proteomic studies show that several Arabidopsis homologs of NuA4 components, including the assembly platform proteins and the catalytic subunit, are associated in vivo with additional members of this complex suggesting that a NuA4-like HAT complex is present in plants. Furthermore, the functional characterization of some Arabidopsis NuA4 subunits has uncovered the involvement of these proteins in the regulation of different plant biological processes. Interestingly, for most of the mutant plants deficient in subunits of this complex characterized so far, conspicuous defects in flowering time are observed, suggesting a role for NuA4 in the control of this plant developmental program. Moreover, the participation of Arabidopsis NuA4 homologs in other developmental processes, such as gametophyte development, as well as in cell proliferation and stress and hormone responses, has also been reported. In this review, we summarize the current state of knowledge on plant putative NuA4 subunits and discuss the latest progress concerning the function of this chromatin modifying complex.
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26
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González-Medina A, Hidalgo E, Ayté J. Gcn5-mediated acetylation at MBF-regulated promoters induces the G1/S transcriptional wave. Nucleic Acids Res 2019; 47:8439-8451. [PMID: 31260531 PMCID: PMC6895280 DOI: 10.1093/nar/gkz561] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/24/2018] [Revised: 06/17/2019] [Accepted: 06/21/2019] [Indexed: 11/26/2022] Open
Abstract
In fission yeast, MBF-dependent transcription is inactivated at the end of S phase through a negative feedback loop that involves the co-repressors, Yox1 and Nrm1. Although this repression system is well known, the molecular mechanisms involved in MBF activation remain largely unknown. Compacted chromatin constitutes a barrier to activators accessing promoters. Here, we show that chromatin regulation plays a key role in activating MBF-dependent transcription. Gcn5, a part of the SAGA complex, binds to MBF-regulated promoters through the MBF co-activator Rep2 in a cell cycle-dependent manner and in a reverse correlation to the binding of the MBF co-repressors, Nrm1 or Yox1. We propose that the co-repressors function as physical barriers to SAGA recruitment onto MBF promoters. We also show that Gcn5 acetylates specific lysine residues on histone H3 in a cell cycle-regulated manner. Furthermore, either in a gcn5 mutant or in a strain in which histone H3 is kept in an unacetylated form, MBF-dependent transcription is downregulated. In summary, Gcn5 is required for the full activation and correct timing of MBF-regulated gene transcription.
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Affiliation(s)
| | - Elena Hidalgo
- Oxidative Stress and Cell Cycle Group, Universitat Pompeu Fabra, Barcelona 08003, Spain
| | - José Ayté
- Oxidative Stress and Cell Cycle Group, Universitat Pompeu Fabra, Barcelona 08003, Spain
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27
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New insights into the evolutionary conservation of the sole PIKK pseudokinase Tra1/TRRAP. Biochem Soc Trans 2019; 47:1597-1608. [DOI: 10.1042/bst20180496] [Citation(s) in RCA: 20] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/23/2019] [Revised: 07/25/2019] [Accepted: 11/05/2019] [Indexed: 02/07/2023]
Abstract
Phosphorylation by protein kinases is a fundamental mechanism of signal transduction. Many kinase families contain one or several members that, although evolutionarily conserved, lack the residues required for catalytic activity. Studies combining structural, biochemical, and functional approaches revealed that these pseudokinases have crucial roles in vivo and may even represent attractive targets for pharmacological intervention. Pseudokinases mediate signal transduction by a diversity of mechanisms, including allosteric regulation of their active counterparts, assembly of signaling hubs, or modulation of protein localization. One such pseudokinase, named Tra1 in yeast and transformation/transcription domain-associated protein (TRRAP) in mammals, is the only member lacking all catalytic residues within the phosphatidylinositol 3-kinase related kinase (PIKK) family of kinases. PIKKs are related to the PI3K family of lipid kinases, but function as Serine/Threonine protein kinases and have pivotal roles in diverse processes such as DNA damage sensing and repair, metabolic control of cell growth, nonsense-mediated decay, or transcription initiation. Tra1/TRRAP is the largest subunit of two distinct transcriptional co-activator complexes, SAGA and NuA4/TIP60, which it recruits to promoters upon transcription factor binding. Here, we review our current knowledge on the Tra1/TRRAP pseudokinase, focusing on its role as a scaffold for SAGA and NuA4/TIP60 complex assembly and recruitment to chromatin. We further discuss its evolutionary history within the PIKK family and highlight recent findings that reveal the importance of molecular chaperones in pseudokinase folding, function, and conservation.
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28
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Elías-Villalobos A, Toullec D, Faux C, Séveno M, Helmlinger D. Chaperone-mediated ordered assembly of the SAGA and NuA4 transcription co-activator complexes in yeast. Nat Commun 2019; 10:5237. [PMID: 31748520 PMCID: PMC6868236 DOI: 10.1038/s41467-019-13243-w] [Citation(s) in RCA: 27] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/08/2019] [Accepted: 10/25/2019] [Indexed: 12/19/2022] Open
Abstract
Transcription initiation involves the coordinated activities of large multimeric complexes, but little is known about their biogenesis. Here we report several principles underlying the assembly and topological organization of the highly conserved SAGA and NuA4 co-activator complexes, which share the Tra1 subunit. We show that Tra1 contributes to the overall integrity of NuA4, whereas, within SAGA, it specifically controls the incorporation of the de-ubiquitination module (DUB), as part of an ordered assembly pathway. Biochemical and functional analyses reveal the mechanism by which Tra1 specifically interacts with either SAGA or NuA4. Finally, we demonstrate that Hsp90 and its cochaperone TTT promote Tra1 de novo incorporation into both complexes, indicating that Tra1, the sole pseudokinase of the PIKK family, shares a dedicated chaperone machinery with its cognate kinases. Overall, our work brings mechanistic insights into the assembly of transcriptional complexes and reveals the contribution of dedicated chaperones to this process. Transcription initiation involves the coordinated assembly and activity of large multimeric complexes. Here the authors report on the chaperone-mediated ordered assembly of the SAGA and NuA4 transcription co-activator complexes in fission yeast, providing insight into the de novo assembly of transcriptional complexes and the contribution of dedicated chaperones to this process.
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Affiliation(s)
| | - Damien Toullec
- CRBM, CNRS, University of Montpellier, Montpellier, France
| | - Céline Faux
- CRBM, CNRS, University of Montpellier, Montpellier, France
| | - Martial Séveno
- BioCampus Montpellier, CNRS, INSERM, University of Montpellier, Montpellier, France
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29
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Bao K, Jia S. Noncatalytic Function of a JmjC Domain Protein Disrupts Heterochromatin. Epigenet Insights 2019; 12:2516865719862249. [PMID: 31321383 PMCID: PMC6624913 DOI: 10.1177/2516865719862249] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/03/2019] [Accepted: 06/13/2019] [Indexed: 11/29/2022] Open
Abstract
Chromatin-modifying enzymes are frequently overexpressed in cancer cells, and
their enzymatic activities play important roles in changing the epigenetic
landscape responsible for tumorigenesis. However, many of these proteins also
execute noncatalytic functions, which are poorly understood. In fission yeast,
overexpression of Epe1, a histone demethylase homolog, causes heterochromatin
defects. Interestingly, in our recent work, we discovered that overexpressed
Epe1 recruits SAGA, a histone acetyltransferase complex important for
transcriptional regulation, to disrupt heterochromatin, independent of its
demethylase activity. Our findings suggest that overexpressed
chromatin-modifying enzymes can alter the epigenetic landscape through changing
their proteomic environments, an area that needs to be further explored in
dissecting disease etiology associated with overexpression of chromatin
regulators.
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Affiliation(s)
- Kehan Bao
- Department of Biological Sciences, Columbia University, New York, NY, USA
| | - Songtao Jia
- Department of Biological Sciences, Columbia University, New York, NY, USA
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30
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Cheung ACM, Díaz-Santín LM. Share and share alike: the role of Tra1 from the SAGA and NuA4 coactivator complexes. Transcription 2019; 10:37-43. [PMID: 30375921 PMCID: PMC6351133 DOI: 10.1080/21541264.2018.1530936] [Citation(s) in RCA: 13] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/25/2018] [Revised: 09/21/2018] [Accepted: 09/24/2018] [Indexed: 01/12/2023] Open
Abstract
SAGA and NuA4 are coactivator complexes required for transcription on chromatin. Although they contain different enzymatic and biochemical activities, both contain the large Tra1 subunit. Recent electron microscopy studies have resolved the complete structure of Tra1 and its integration in SAGA/NuA4, providing important insight into Tra1 function.
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Affiliation(s)
- Alan C. M. Cheung
- Department of Structural and Molecular Biology, University College London, Institute of Structural and Molecular Biology, London, UK
- Institute of Structural and Molecular Biology, Biological Sciences, Birkbeck College, London, UK
| | - Luis Miguel Díaz-Santín
- Department of Structural and Molecular Biology, University College London, Institute of Structural and Molecular Biology, London, UK
- Institute of Structural and Molecular Biology, Biological Sciences, Birkbeck College, London, UK
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31
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Bao K, Shan CM, Moresco J, Yates J, Jia S. Anti-silencing factor Epe1 associates with SAGA to regulate transcription within heterochromatin. Genes Dev 2018; 33:116-126. [PMID: 30573453 PMCID: PMC6317313 DOI: 10.1101/gad.318030.118] [Citation(s) in RCA: 26] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/19/2018] [Accepted: 10/19/2018] [Indexed: 12/11/2022]
Abstract
In this study, Bao et al. investigated how transcription is regulated within heterochromatin in fission yeast. They show that overexpressed Epe1 associates with SAGA and recruits SAGA to heterochromatin regions (which leads to an increase in histone acetylation, transcription of repeats, and the disruption of heterochromatin) and that Epe1 recruits SAGA to regulate transcription within heterochromatin when expressed at normal levels. Heterochromatin is a highly condensed form of chromatin that silences gene transcription. Although high levels of transcriptional activities disrupt heterochromatin, transcription of repetitive DNA elements and subsequent processing of the transcripts by the RNAi machinery are required for heterochromatin assembly. In fission yeast, a JmjC domain protein, Epe1, promotes transcription of DNA repeats to facilitate heterochromatin formation, but overexpression of Epe1 leads to heterochromatin defects. However, the molecular function of Epe1 is not well understood. By screening the fission yeast deletion library, we found that heterochromatin defects associated with Epe1 overexpression are alleviated by mutations of the SAGA histone acetyltransferase complex. Overexpressed Epe1 associates with SAGA and recruits SAGA to heterochromatin regions, which leads to increased histone acetylation, transcription of repeats, and the disruption of heterochromatin. At its normal expression levels, Epe1 also associates with SAGA, albeit weakly. Such interaction regulates histone acetylation levels at heterochromatin and promotes transcription of repeats for heterochromatin assembly. Our results also suggest that increases of certain chromatin protein levels, which frequently occur in cancer cells, might strengthen relatively weak interactions to affect the epigenetic landscape.
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Affiliation(s)
- Kehan Bao
- Department of Biological Sciences, Columbia University, New York, New York 10027, USA
| | - Chun-Min Shan
- Department of Biological Sciences, Columbia University, New York, New York 10027, USA
| | - James Moresco
- Department of Molecular Medicine and Neurobiology, The Scripps Research Institute, La Jolla, California 92037, USA
| | - John Yates
- Department of Molecular Medicine and Neurobiology, The Scripps Research Institute, La Jolla, California 92037, USA
| | - Songtao Jia
- Department of Biological Sciences, Columbia University, New York, New York 10027, USA
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32
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Fischer V, Schumacher K, Tora L, Devys D. Global role for coactivator complexes in RNA polymerase II transcription. Transcription 2018; 10:29-36. [PMID: 30299209 PMCID: PMC6351120 DOI: 10.1080/21541264.2018.1521214] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/19/2022] Open
Abstract
SAGA and TFIID are related transcription complexes, which were proposed to alternatively deliver TBP at different promoter classes. Recent genome-wide studies in yeast revealed that both complexes are required for the transcription of a vast majority of genes by RNA polymerase II raising new questions about the role of coactivators.
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Affiliation(s)
- Veronique Fischer
- a Institut de Génétique et de Biologie Moléculaire et Cellulaire , Illkirch , France.,b Centre National de la Recherche Scientifique , UMR7104 , Illkirch , France.,c Institut National de la Santé et de la Recherche Médicale , Illkirch , France.,d Université de Strasbourg , Illkirch , France
| | - Kenny Schumacher
- a Institut de Génétique et de Biologie Moléculaire et Cellulaire , Illkirch , France.,b Centre National de la Recherche Scientifique , UMR7104 , Illkirch , France.,c Institut National de la Santé et de la Recherche Médicale , Illkirch , France.,d Université de Strasbourg , Illkirch , France
| | - Laszlo Tora
- a Institut de Génétique et de Biologie Moléculaire et Cellulaire , Illkirch , France.,b Centre National de la Recherche Scientifique , UMR7104 , Illkirch , France.,c Institut National de la Santé et de la Recherche Médicale , Illkirch , France.,d Université de Strasbourg , Illkirch , France
| | - Didier Devys
- a Institut de Génétique et de Biologie Moléculaire et Cellulaire , Illkirch , France.,b Centre National de la Recherche Scientifique , UMR7104 , Illkirch , France.,c Institut National de la Santé et de la Recherche Médicale , Illkirch , France.,d Université de Strasbourg , Illkirch , France
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33
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Bruzzone MJ, Grünberg S, Kubik S, Zentner GE, Shore D. Distinct patterns of histone acetyltransferase and Mediator deployment at yeast protein-coding genes. Genes Dev 2018; 32:1252-1265. [PMID: 30108132 PMCID: PMC6120713 DOI: 10.1101/gad.312173.118] [Citation(s) in RCA: 34] [Impact Index Per Article: 4.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/23/2018] [Accepted: 06/19/2018] [Indexed: 02/07/2023]
Abstract
Here, Bruzzone et al. explore the relative contributions of the transcriptional coactivators Mediator and two histone acetyltransferase (HAT) complexes, NuA4 and SAGA, to RNA polymerase II association at specific genes and gene classes by rapid nuclear depletion of key complex subunits. They show that the NuA4 HAT Esa1 differentially affects certain groups of genes, whereas the SAGA HAT Gcn5 has a weaker but more uniform effect, and their findings suggest that at least three distinct combinations of coactivator deployment are used to generate moderate or high transcription levels. The transcriptional coactivators Mediator and two histone acetyltransferase (HAT) complexes, NuA4 and SAGA, play global roles in transcriptional activation. Here we explore the relative contributions of these factors to RNA polymerase II association at specific genes and gene classes by rapid nuclear depletion of key complex subunits. We show that the NuA4 HAT Esa1 differentially affects certain groups of genes, whereas the SAGA HAT Gcn5 has a weaker but more uniform effect. Relative dependence on Esa1 and Tra1, a shared component of NuA4 and SAGA, distinguishes two large groups of coregulated growth-promoting genes. In contrast, we show that the activity of Mediator is particularly important at a separate, small set of highly transcribed TATA-box-containing genes. Our analysis indicates that at least three distinct combinations of coactivator deployment are used to generate moderate or high transcription levels and suggests that each may be associated with distinct forms of regulation.
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Affiliation(s)
- Maria Jessica Bruzzone
- Department of Molecular Biology, Institute of Genetics and Genomics in Geneva, 1211 Geneva 4, Switzerland
| | - Sebastian Grünberg
- Basic Sciences Division, Fred Hutchinson Cancer Research Center, Seattle, Washington 98109, USA
| | - Slawomir Kubik
- Department of Molecular Biology, Institute of Genetics and Genomics in Geneva, 1211 Geneva 4, Switzerland
| | - Gabriel E Zentner
- Department of Biology, Indiana University, Bloomington, Indiana 47405, USA
| | - David Shore
- Department of Molecular Biology, Institute of Genetics and Genomics in Geneva, 1211 Geneva 4, Switzerland
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34
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Berg MD, Genereaux J, Karagiannis J, Brandl CJ. The Pseudokinase Domain of Saccharomyces cerevisiae Tra1 Is Required for Nuclear Localization and Incorporation into the SAGA and NuA4 Complexes. G3 (BETHESDA, MD.) 2018; 8:1943-1957. [PMID: 29626083 PMCID: PMC5982823 DOI: 10.1534/g3.118.200288] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 01/03/2018] [Accepted: 04/04/2018] [Indexed: 12/29/2022]
Abstract
Tra1 is an essential component of the SAGA/SLIK and NuA4 complexes in S. cerevisiae, recruiting these co-activator complexes to specific promoters. As a PIKK family member, Tra1 is characterized by a C-terminal phosphoinositide 3-kinase (PI3K) domain. Unlike other PIKK family members (e.g., Tor1, Tor2, Mec1, Tel1), Tra1 has no demonstrable kinase activity. We identified three conserved arginine residues in Tra1 that reside proximal or within the cleft between the N- and C-terminal subdomains of the PI3K domain. To establish a function for Tra1's PI3K domain and specifically the cleft region, we characterized a tra1 allele where these three arginine residues are mutated to glutamine. The half-life of the Tra1[Formula: see text] protein is reduced but its steady state level is maintained at near wild-type levels by a transcriptional feedback mechanism. The tra1[Formula: see text] allele results in slow growth under stress and alters the expression of genes also regulated by other components of the SAGA complex. Tra1[Formula: see text] is less efficiently transported to the nucleus than the wild-type protein. Likely related to this, Tra1[Formula: see text] associates poorly with SAGA/SLIK and NuA4. The ratio of Spt7SLIK to Spt7SAGA increases in the tra1[Formula: see text] strain and truncated forms of Spt20 become apparent upon isolation of SAGA/SLIK. Intragenic suppressor mutations of tra1[Formula: see text] map to the cleft region further emphasizing its importance. We propose that the PI3K domain of Tra1 is directly or indirectly important for incorporating Tra1 into SAGA and NuA4 and thus the biosynthesis and/or stability of the intact complexes.
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Affiliation(s)
- Matthew D Berg
- Department of Biochemistry, Schulich School of Medicine & Dentistry, Western University, London, Ontario, Canada N6A5C1
| | - Julie Genereaux
- Department of Biochemistry, Schulich School of Medicine & Dentistry, Western University, London, Ontario, Canada N6A5C1
| | - Jim Karagiannis
- Department of Biology, Western University, London, Ontario, Canada N6A5B7
| | - Christopher J Brandl
- Department of Biochemistry, Schulich School of Medicine & Dentistry, Western University, London, Ontario, Canada N6A5C1
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35
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Helmlinger D, Tora L. Sharing the SAGA. Trends Biochem Sci 2017; 42:850-861. [PMID: 28964624 PMCID: PMC5660625 DOI: 10.1016/j.tibs.2017.09.001] [Citation(s) in RCA: 122] [Impact Index Per Article: 15.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/13/2017] [Revised: 08/30/2017] [Accepted: 09/05/2017] [Indexed: 12/14/2022]
Abstract
Transcription initiation is a major regulatory step in eukaryotic gene expression. Co-activators establish transcriptionally competent promoter architectures and chromatin signatures to allow the formation of the pre-initiation complex (PIC), comprising RNA polymerase II (Pol II) and general transcription factors (GTFs). Many GTFs and co-activators are multisubunit complexes, in which individual components are organized into functional modules carrying specific activities. Recent advances in affinity purification and mass spectrometry analyses have revealed that these complexes often share functional modules, rather than containing unique components. This observation appears remarkably prevalent for chromatin-modifying and remodeling complexes. Here, we use the modular organization of the evolutionary conserved Spt-Ada-Gcn5 acetyltransferase (SAGA) complex as a paradigm to illustrate how co-activators share and combine a relatively limited set of functional tools.
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Affiliation(s)
| | - László Tora
- Institut de Génétique et de Biologie Moléculaire et Cellulaire, 67404 Illkirch, France; Centre National de la Recherche Scientifique, UMR7104, 67404 Illkirch, France; Institut National de la Santé et de la Recherche Médicale, U964, 67404 Illkirch, France; Université de Strasbourg, 67404 Illkirch, France.
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36
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Laboucarié T, Detilleux D, Rodriguez-Mias RA, Faux C, Romeo Y, Franz-Wachtel M, Krug K, Maček B, Villén J, Petersen J, Helmlinger D. TORC1 and TORC2 converge to regulate the SAGA co-activator in response to nutrient availability. EMBO Rep 2017; 18:2197-2218. [PMID: 29079657 DOI: 10.15252/embr.201744942] [Citation(s) in RCA: 34] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/31/2017] [Revised: 08/31/2017] [Accepted: 09/07/2017] [Indexed: 12/19/2022] Open
Abstract
Gene expression regulation is essential for cells to adapt to changes in their environment. Co-activator complexes have well-established roles in transcriptional regulation, but less is known about how they sense and respond to signaling cues. We have previously shown that, in fission yeast, one such co-activator, the SAGA complex, controls gene expression and the switch from proliferation to differentiation in response to nutrient availability. Here, using a combination of genetic, biochemical, and proteomic approaches, we show that SAGA responds to nutrients through the differential phosphorylation of its Taf12 component, downstream of both the TORC1 and TORC2 pathways. Taf12 phosphorylation increases early upon starvation and is controlled by the opposing activities of the PP2A phosphatase, which is activated by TORC1, and the TORC2-activated Gad8AKT kinase. Mutational analyses suggest that Taf12 phosphorylation prevents cells from committing to differentiation until starvation reaches a critical level. Overall, our work reveals that SAGA is a direct target of nutrient-sensing pathways and has uncovered a mechanism by which TORC1 and TORC2 converge to control gene expression and cell fate decisions.
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Affiliation(s)
| | | | | | - Céline Faux
- CRBM, CNRS, University of Montpellier, Montpellier, France
| | - Yves Romeo
- CRBM, CNRS, University of Montpellier, Montpellier, France
| | | | | | - Boris Maček
- Proteome Center Tübingen, Tuebingen, Germany
| | - Judit Villén
- Department of Genome Sciences, University of Washington, Seattle, WA, USA
| | - Janni Petersen
- Flinders Centre for Innovation in Cancer, School of Medicine, Faculty of Health Science, Flinders University, Adelaide, SA, Australia
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37
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Li X, Seidel CW, Szerszen LT, Lange JJ, Workman JL, Abmayr SM. Enzymatic modules of the SAGA chromatin-modifying complex play distinct roles in Drosophila gene expression and development. Genes Dev 2017; 31:1588-1600. [PMID: 28887412 PMCID: PMC5630023 DOI: 10.1101/gad.300988.117] [Citation(s) in RCA: 39] [Impact Index Per Article: 4.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/25/2017] [Accepted: 08/11/2017] [Indexed: 01/03/2023]
Abstract
In this study, Li et al. demonstrate that the two enzymatic modules of the Drosophila Spt–Ada–Gcn5–acetyltransferase (SAGA) chromatin-modifying complex are differently required in oogenesis. Their findings demonstrate that loss of the histone acetyltransferase (HAT) activity blocks oogenesis, while loss of H2B deubiquitinase (DUB) activity does not, suggesting that the DUB module has functions within SAGA as well as independent functions. The Spt–Ada–Gcn5–acetyltransferase (SAGA) chromatin-modifying complex is a transcriptional coactivator that contains four different modules of subunits. The intact SAGA complex has been well characterized for its function in transcription regulation and development. However, little is known about the roles of individual modules within SAGA and whether they have any SAGA-independent functions. Here we demonstrate that the two enzymatic modules of Drosophila SAGA are differently required in oogenesis. Loss of the histone acetyltransferase (HAT) activity blocks oogenesis, while loss of the H2B deubiquitinase (DUB) activity does not. However, the DUB module regulates a subset of genes in early embryogenesis, and loss of the DUB subunits causes defects in embryogenesis. ChIP-seq (chromatin immunoprecipitation [ChIP] combined with high-throughput sequencing) analysis revealed that both the DUB and HAT modules bind most SAGA target genes even though many of these targets do not require the DUB module for expression. Furthermore, we found that the DUB module can bind to chromatin and regulate transcription independently of the HAT module. Our results suggest that the DUB module has functions within SAGA and independent functions.
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Affiliation(s)
- Xuanying Li
- Stowers Institute for Medical Research, Kansas City, Missouri 64110, USA.,Department of Anatomy and Cell Biology, University of Kansas Medical Center, Kansas City, Kansas 66160, USA
| | | | - Leanne T Szerszen
- Stowers Institute for Medical Research, Kansas City, Missouri 64110, USA
| | - Jeffrey J Lange
- Stowers Institute for Medical Research, Kansas City, Missouri 64110, USA
| | - Jerry L Workman
- Stowers Institute for Medical Research, Kansas City, Missouri 64110, USA
| | - Susan M Abmayr
- Stowers Institute for Medical Research, Kansas City, Missouri 64110, USA.,Department of Anatomy and Cell Biology, University of Kansas Medical Center, Kansas City, Kansas 66160, USA
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38
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Díaz-Santín LM, Lukoyanova N, Aciyan E, Cheung AC. Cryo-EM structure of the SAGA and NuA4 coactivator subunit Tra1 at 3.7 angstrom resolution. eLife 2017; 6:28384. [PMID: 28767037 PMCID: PMC5576489 DOI: 10.7554/elife.28384] [Citation(s) in RCA: 33] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/05/2017] [Accepted: 07/31/2017] [Indexed: 01/30/2023] Open
Abstract
Coactivator complexes SAGA and NuA4 stimulate transcription by post-translationally modifying chromatin. Both complexes contain the Tra1 subunit, a highly conserved 3744-residue protein from the Phosphoinositide 3-Kinase-related kinase (PIKK) family and a direct target for multiple sequence-specific activators. We present the Cryo-EM structure of Saccharomyces cerevsisae Tra1 to 3.7 Å resolution, revealing an extensive network of alpha-helical solenoids organized into a diamond ring conformation and is strikingly reminiscent of DNA-PKcs, suggesting a direct role for Tra1 in DNA repair. The structure was fitted into an existing SAGA EM reconstruction and reveals limited contact surfaces to Tra1, hence it does not act as a molecular scaffold within SAGA. Mutations that affect activator targeting are distributed across the Tra1 structure, but also cluster within the N-terminal Finger region, indicating the presence of an activator interaction site. The structure of Tra1 is a key milestone in deciphering the mechanism of multiple coactivator complexes. Inside our cells, histone proteins package and condense DNA so that it can fit into the cell nucleus. However, this also switches off the genes, since the machines that read and interpret them can no longer access the underlying DNA. Turning genes on requires specific enzymes that chemically modify the histone proteins to regain access to the DNA. This must be carefully controlled, otherwise the ‘wrong’ genes can be activated, causing undesired effects and endangering the cell. Histone modifying enzymes often reside in large protein complexes. Two well-known examples are the SAGA and NuA4 complexes. Both have different roles during gene activation, but share a protein called Tra1. This protein enables SAGA and NuA4 to act on specific genes by binding to ‘activator proteins’ that are found on the DNA. Tra1 is one of the biggest proteins in the cell, but its size makes it difficult to study and until now, its structure was unknown. To determine the structure of Tra1, Díaz-Santín et al. extracted the protein from baker’s yeast, and examined it using electron microscopy. The structure of Tra1 resembled a diamond ring with multiple protein domains that correspond to a band, setting and a centre stone. The structure was detailed enough so that Díaz-Santín et al. could locate various mutations that affect the role of Tra1. These locations are likely to be direct interfaces to the ‘activator proteins’. Moreover, the study showed that Tra1 was similar to another protein that repairs damaged DNA. These results suggest that Tra1 not only works as an activator target, but may also have a role in repairing damaged DNA, and might even connect these two processes. Yeast Tra1 is also very similar to its human counterpart, which has been shown to stimulate cells to become cancerous. Further studies based on these results may help us understand how cancer begins.
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Affiliation(s)
- Luis Miguel Díaz-Santín
- Department of Structural and Molecular Biology, Institute of Structural and Molecular Biology, University College London, London, United Kingdom
| | - Natasha Lukoyanova
- Institute of Structural and Molecular Biology, Biological Sciences, Birkbeck College, London, United Kingdom
| | - Emir Aciyan
- Institute of Structural and Molecular Biology, Biological Sciences, Birkbeck College, London, United Kingdom
| | - Alan Cm Cheung
- Department of Structural and Molecular Biology, Institute of Structural and Molecular Biology, University College London, London, United Kingdom.,Institute of Structural and Molecular Biology, Biological Sciences, Birkbeck College, London, United Kingdom
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Vosnakis N, Koch M, Scheer E, Kessler P, Mély Y, Didier P, Tora L. Coactivators and general transcription factors have two distinct dynamic populations dependent on transcription. EMBO J 2017; 36:2710-2725. [PMID: 28724529 PMCID: PMC5599802 DOI: 10.15252/embj.201696035] [Citation(s) in RCA: 17] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/07/2016] [Revised: 06/08/2017] [Accepted: 06/15/2017] [Indexed: 12/29/2022] Open
Abstract
SAGA and ATAC are two distinct chromatin modifying co‐activator complexes with distinct enzymatic activities involved in RNA polymerase II (Pol II) transcription regulation. To investigate the mobility of co‐activator complexes and general transcription factors in live‐cell nuclei, we performed imaging experiments based on photobleaching. SAGA and ATAC, but also two general transcription factors (TFIID and TFIIB), were highly dynamic, exhibiting mainly transient associations with chromatin, contrary to Pol II, which formed more stable chromatin interactions. Fluorescence correlation spectroscopy analyses revealed that the mobile pool of the two co‐activators, as well as that of TFIID and TFIIB, can be subdivided into “fast” (free) and “slow” (chromatin‐interacting) populations. Inhibiting transcription elongation decreased H3K4 trimethylation and reduced the “slow” population of SAGA, ATAC, TFIIB and TFIID. In addition, inhibiting histone H3K4 trimethylation also reduced the “slow” populations of SAGA and ATAC. Thus, our results demonstrate that in the nuclei of live cells the equilibrium between fast and slow population of SAGA or ATAC complexes is regulated by active transcription via changes in the abundance of H3K4me3 on chromatin.
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Affiliation(s)
- Nikolaos Vosnakis
- Institut de Génétique et de Biologie Moléculaire et Cellulaire, Illkirch, France.,Centre National de la Recherche Scientifique, UMR7104, Illkirch, France.,Institut National de la Santé et de la Recherche Médicale, U964, Illkirch, France.,Université de Strasbourg, Illkirch, France
| | - Marc Koch
- Institut de Génétique et de Biologie Moléculaire et Cellulaire, Illkirch, France.,Centre National de la Recherche Scientifique, UMR7104, Illkirch, France.,Institut National de la Santé et de la Recherche Médicale, U964, Illkirch, France.,Université de Strasbourg, Illkirch, France
| | - Elisabeth Scheer
- Institut de Génétique et de Biologie Moléculaire et Cellulaire, Illkirch, France.,Centre National de la Recherche Scientifique, UMR7104, Illkirch, France.,Institut National de la Santé et de la Recherche Médicale, U964, Illkirch, France.,Université de Strasbourg, Illkirch, France
| | - Pascal Kessler
- Institut de Génétique et de Biologie Moléculaire et Cellulaire, Illkirch, France.,Centre National de la Recherche Scientifique, UMR7104, Illkirch, France.,Institut National de la Santé et de la Recherche Médicale, U964, Illkirch, France.,Université de Strasbourg, Illkirch, France
| | - Yves Mély
- Université de Strasbourg, Illkirch, France.,Laboratoire de Biophotonique et Pharmacologie, Illkirch, France
| | - Pascal Didier
- Université de Strasbourg, Illkirch, France.,Laboratoire de Biophotonique et Pharmacologie, Illkirch, France
| | - László Tora
- Institut de Génétique et de Biologie Moléculaire et Cellulaire, Illkirch, France .,Centre National de la Recherche Scientifique, UMR7104, Illkirch, France.,Institut National de la Santé et de la Recherche Médicale, U964, Illkirch, France.,Université de Strasbourg, Illkirch, France
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Global Fitness Profiling Identifies Arsenic and Cadmium Tolerance Mechanisms in Fission Yeast. G3-GENES GENOMES GENETICS 2016; 6:3317-3333. [PMID: 27558664 PMCID: PMC5068951 DOI: 10.1534/g3.116.033829] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Indexed: 01/08/2023]
Abstract
Heavy metals and metalloids such as cadmium [Cd(II)] and arsenic [As(III)] are widespread environmental toxicants responsible for multiple adverse health effects in humans. However, the molecular mechanisms underlying metal-induced cytotoxicity and carcinogenesis, as well as the detoxification and tolerance pathways, are incompletely understood. Here, we use global fitness profiling by barcode sequencing to quantitatively survey the Schizosaccharomyces pombe haploid deletome for genes that confer tolerance of cadmium or arsenic. We identified 106 genes required for cadmium resistance and 110 genes required for arsenic resistance, with a highly significant overlap of 36 genes. A subset of these 36 genes account for almost all proteins required for incorporating sulfur into the cysteine-rich glutathione and phytochelatin peptides that chelate cadmium and arsenic. A requirement for Mms19 is explained by its role in directing iron–sulfur cluster assembly into sulfite reductase as opposed to promoting DNA repair, as DNA damage response genes were not enriched among those required for cadmium or arsenic tolerance. Ubiquinone, siroheme, and pyridoxal 5′-phosphate biosynthesis were also identified as critical for Cd/As tolerance. Arsenic-specific pathways included prefoldin-mediated assembly of unfolded proteins and protein targeting to the peroxisome, whereas cadmium-specific pathways included plasma membrane and vacuolar transporters, as well as Spt–Ada–Gcn5-acetyltransferase (SAGA) transcriptional coactivator that controls expression of key genes required for cadmium tolerance. Notable differences are apparent with corresponding screens in the budding yeast Saccharomyces cerevisiae, underscoring the utility of analyzing toxic metal defense mechanisms in both organisms.
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Sgf73, a subunit of SAGA complex, is required for the assembly of RITS complex in fission yeast. Sci Rep 2015; 5:14707. [PMID: 26443059 PMCID: PMC4595766 DOI: 10.1038/srep14707] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/07/2015] [Accepted: 09/07/2015] [Indexed: 02/04/2023] Open
Abstract
RNA interference (RNAi) is a widespread gene-silencing mechanism and is required for heterochromatin assembly in a variety of organisms. The RNA-induced transcriptional silencing complex (RITS), composed of Ago1, Tas3 and Chp1, is a key component of RNAi machinery in fission yeast that connects short interference RNA (siRNA) and heterochromatin formation. However, the process by which RITS is assembled is not well understood. Here, we identified Sgf73, a subunit of the SAGA co-transcriptional complex, is required for pericentromeric heterochromatin silencing and the generation of siRNA. This novel role of Sgf73 is independent of enzymatic activities or structural integrity of SAGA. Instead, Sgf73 is physically associated with Ago1 and Chp1. The interactions among the subunits of the RITS, including those between Tas3 and Chp1, between Chp1 and Ago1, between Ago1 and Tas3, were all impaired by the deletion of sgf73+. Consistently, the recruitment of Ago1 and Chp1 to the pericentromeric region was abolished in sgf73Δ cells. Our study unveils a moonlighting function of a SAGA subunit. It suggests Sgf73 is a novel factor that promotes assembly of RITS and RNAi-mediated heterochromatin formation.
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Kurabe N, Murakami S, Tashiro F. SGF29 and Sry pathway in hepatocarcinogenesis. World J Biol Chem 2015; 6:139-147. [PMID: 26322172 PMCID: PMC4549758 DOI: 10.4331/wjbc.v6.i3.139] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/27/2015] [Revised: 05/31/2015] [Accepted: 07/02/2015] [Indexed: 02/05/2023] Open
Abstract
Deregulated c-Myc expression is a hallmark of many human cancers. We have recently identified a role of mammalian homolog of yeast SPT-ADA-GCN5-acetyltransferas (SAGA) complex component, SAGA-associated factor 29 (SGF29), in regulating the c-Myc overexpression. Here, we discuss the molecular nature of SFG29 in SPT3-TAF9-GCN5-acetyltransferase complex, a counterpart of yeast SAGA complex, and the mechanism through which the elevated SGF29 expression contribute to oncogenic potential of c-Myc in hepatocellularcarcinoma (HCC). We propose that the upstream regulation of SGF29 elicited by sex-determining region Y (Sry) is also augmented in HCC. We hypothesize that c-Myc elevation driven by the deregulated Sry and SGF29 pathway is implicated in the male specific acquisition of human HCCs.
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Bonnet J, Wang CY, Baptista T, Vincent SD, Hsiao WC, Stierle M, Kao CF, Tora L, Devys D. The SAGA coactivator complex acts on the whole transcribed genome and is required for RNA polymerase II transcription. Genes Dev 2014; 28:1999-2012. [PMID: 25228644 PMCID: PMC4173158 DOI: 10.1101/gad.250225.114] [Citation(s) in RCA: 160] [Impact Index Per Article: 14.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/24/2022]
Abstract
The SAGA coactivator complex contains distinct chromatin-modifying activities and is recruited by DNA-bound activators to regulate the expression of a subset of genes. Bonnet et al. discovered that SAGA acetylates the promoters and deubiquitinates the transcribed region of all expressed genes. SAGA also plays a critical role for RNA polymerase II recruitment at all expressed genes. This study uncovers a new function for SAGA as a bona fide cofactor for all RNA polymerase II transcription. The SAGA (Spt–Ada–Gcn5 acetyltransferase) coactivator complex contains distinct chromatin-modifying activities and is recruited by DNA-bound activators to regulate the expression of a subset of genes. Surprisingly, recent studies revealed little overlap between genome-wide SAGA-binding profiles and changes in gene expression upon depletion of subunits of the complex. As indicators of SAGA recruitment on chromatin, we monitored in yeast and human cells the genome-wide distribution of histone H3K9 acetylation and H2B ubiquitination, which are respectively deposited or removed by SAGA. Changes in these modifications after inactivation of the corresponding enzyme revealed that SAGA acetylates the promoters and deubiquitinates the transcribed region of all expressed genes. In agreement with this broad distribution, we show that SAGA plays a critical role for RNA polymerase II recruitment at all expressed genes. In addition, through quantification of newly synthesized RNA, we demonstrated that SAGA inactivation induced a strong decrease of mRNA synthesis at all tested genes. Analysis of the SAGA deubiquitination activity further revealed that SAGA acts on the whole transcribed genome in a very fast manner, indicating a highly dynamic association of the complex with chromatin. Thus, our study uncovers a new function for SAGA as a bone fide cofactor for all RNA polymerase II transcription.
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Affiliation(s)
- Jacques Bonnet
- Institut de Génétique et de Biologie Moléculaire et Cellulaire, 67404 Illkirch, France; UMR7104, Centre National de la Recherche Scientifique, 67404 Illkirch, France; U964, Institut National de la Santé et de la Recherche Médicale, 67404 Illkirch, France; Université de Strasbourg, 67404 Illkirch, Cedex, France
| | - Chen-Yi Wang
- Institut de Génétique et de Biologie Moléculaire et Cellulaire, 67404 Illkirch, France; Institute of Cellular and Organismic Biology, Academia Sinica, Taipei 11529, Taiwan
| | - Tiago Baptista
- Institut de Génétique et de Biologie Moléculaire et Cellulaire, 67404 Illkirch, France; UMR7104, Centre National de la Recherche Scientifique, 67404 Illkirch, France; U964, Institut National de la Santé et de la Recherche Médicale, 67404 Illkirch, France; Université de Strasbourg, 67404 Illkirch, Cedex, France
| | - Stéphane D Vincent
- Institut de Génétique et de Biologie Moléculaire et Cellulaire, 67404 Illkirch, France; UMR7104, Centre National de la Recherche Scientifique, 67404 Illkirch, France; U964, Institut National de la Santé et de la Recherche Médicale, 67404 Illkirch, France; Université de Strasbourg, 67404 Illkirch, Cedex, France
| | - Wei-Chun Hsiao
- Institute of Cellular and Organismic Biology, Academia Sinica, Taipei 11529, Taiwan
| | - Matthieu Stierle
- Institut de Génétique et de Biologie Moléculaire et Cellulaire, 67404 Illkirch, France; UMR7104, Centre National de la Recherche Scientifique, 67404 Illkirch, France; U964, Institut National de la Santé et de la Recherche Médicale, 67404 Illkirch, France; Université de Strasbourg, 67404 Illkirch, Cedex, France
| | - Cheng-Fu Kao
- Institute of Cellular and Organismic Biology, Academia Sinica, Taipei 11529, Taiwan
| | - László Tora
- Institut de Génétique et de Biologie Moléculaire et Cellulaire, 67404 Illkirch, France; UMR7104, Centre National de la Recherche Scientifique, 67404 Illkirch, France; U964, Institut National de la Santé et de la Recherche Médicale, 67404 Illkirch, France; Université de Strasbourg, 67404 Illkirch, Cedex, France
| | - Didier Devys
- Institut de Génétique et de Biologie Moléculaire et Cellulaire, 67404 Illkirch, France; UMR7104, Centre National de la Recherche Scientifique, 67404 Illkirch, France; U964, Institut National de la Santé et de la Recherche Médicale, 67404 Illkirch, France; Université de Strasbourg, 67404 Illkirch, Cedex, France;
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Histone H2B ubiquitination promotes the function of the anaphase-promoting complex/cyclosome in Schizosaccharomyces pombe. G3-GENES GENOMES GENETICS 2014; 4:1529-38. [PMID: 24948786 PMCID: PMC4132182 DOI: 10.1534/g3.114.012625] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Indexed: 11/29/2022]
Abstract
Ubiquitination and deubiquitination of proteins are reciprocal events involved in many cellular processes, including the cell cycle. During mitosis, the metaphase to anaphase transition is regulated by the ubiquitin ligase activity of the anaphase-promoting complex/cyclosome (APC/C). Although the E3 ubiquitin ligase function of the APC/C has been well characterized, it is not clear whether deubiquitinating enzymes (DUBs) play a role in reversing APC/C substrate ubiquitination. Here we performed a genetic screen to determine what DUB, if any, antagonizes the function of the APC/C in the fission yeast Schizosaccharomyces pombe. We found that deletion of ubp8, encoding the Spt-Ada-Gcn5-Acetyl transferase (SAGA) complex associated DUB, suppressed temperature-sensitive phenotypes of APC/C mutants cut9-665, lid1-6, cut4-533, and slp1-362. Our analysis revealed that Ubp8 antagonizes APC/C function in a mechanism independent of the spindle assembly checkpoint and proteasome activity. Notably, suppression of APC/C mutants was linked to loss of Ubp8 catalytic activity and required histone H2B ubiquitination. On the basis of these data, we conclude that Ubp8 antagonizes APC/C function indirectly by modulating H2B ubiquitination status.
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45
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Lei B, Zhou N, Guo Y, Zhao W, Tan YW, Yu Y, Lu H. Septin ring assembly is regulated by Spt20, a structural subunit of SAGA complex. J Cell Sci 2014; 127:4024-36. [DOI: 10.1242/jcs.151910] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022] Open
Abstract
Accurate cell division requires proper assembly of high-order septin structures. In fission yeast, Spn1-4 are assembled into a primary septin ring at the division site, and the subsequent recruitment of Mid2 to the structure results in a stable septin ring. However, not much is known about the regulation of this key process. Here, we found deletion of Spt20, a structural subunit of SAGA transcriptional activation complex, caused a severe cell separation defect. The defect is mainly due to impaired septin ring assembly, as 80% of spt20Δ cells lost septin rings at the division sites. Spt20 regulates septin ring assembly partially through the transcriptional activation of mid2+. Spt20 also interacts with Spn2 and Mid2 in vitro and is associated with other components of the ring in vivo. Spt20 is co-localized with the septin ring, but does not separate when the septin ring splits. Importantly, Spt20 regulates the stability of the septin ring and is required for the recruitment of Mid2. The transcription-dependent and -independent roles of Spt20 in the septin ring assembly highlight a multifaceted regulation of one process by a SAGA subunit.
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46
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Ansari SA, Morse RH. Mechanisms of Mediator complex action in transcriptional activation. Cell Mol Life Sci 2013; 70:2743-56. [PMID: 23361037 PMCID: PMC11113466 DOI: 10.1007/s00018-013-1265-9] [Citation(s) in RCA: 72] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/25/2012] [Revised: 01/07/2013] [Accepted: 01/09/2013] [Indexed: 12/14/2022]
Abstract
Mediator is a large multisubunit complex that plays a central role in the regulation of RNA Pol II transcribed genes. Conserved in overall structure and function among eukaryotes, Mediator comprises 25-30 protein subunits that reside in four distinct modules, termed head, middle, tail, and CDK8/kinase. Different subunits of Mediator contact other transcriptional regulators including activators, co-activators, general transcription factors, subunits of RNA Pol II, and specifically modified histones, leading to the regulated expression of target genes. This review is focused on the interactions of specific Mediator subunits with diverse transcription regulators and how those interactions contribute to Mediator function in transcriptional activation.
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Affiliation(s)
- Suraiya A. Ansari
- Laboratory of Molecular Genetics, Wadsworth Center, New York State Department of Health, Albany, NY 12201–0509 USA
| | - Randall H. Morse
- Laboratory of Molecular Genetics, Wadsworth Center, New York State Department of Health, Albany, NY 12201–0509 USA
- Department of Biomedical Science, University at Albany School of Public Health, Albany, NY USA
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Kamata K, Hatanaka A, Goswami G, Shinmyozu K, Nakayama JI, Urano T, Hatashita M, Uchida H, Oki M. C-terminus of the Sgf73 subunit of SAGA and SLIK is important for retention in the larger complex and for heterochromatin boundary function. Genes Cells 2013; 18:823-37. [DOI: 10.1111/gtc.12075] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/21/2013] [Accepted: 05/22/2013] [Indexed: 11/26/2022]
Affiliation(s)
- Kazuma Kamata
- Department of Applied Chemistry & Biotechnology; Graduate School of Engineering; University of Fukui; Fukui 910-8507; Japan
| | - Akira Hatanaka
- Department of Applied Chemistry & Biotechnology; Graduate School of Engineering; University of Fukui; Fukui 910-8507; Japan
| | - Gayatri Goswami
- Department of Applied Chemistry & Biotechnology; Graduate School of Engineering; University of Fukui; Fukui 910-8507; Japan
| | - Kaori Shinmyozu
- Center for Developmental Biology; Laboratory for Chromatin Dynamics; RIKEN; Kobe 650-0047; Japan
| | | | - Takeshi Urano
- Department of Biochemistry; Shimane University Faculty of Medicine; Izumo 693-8501; Japan
| | - Masanori Hatashita
- Research and Development Department; Wakasa Wan Energy Research Center; Tsuruga 914-0192; Japan
| | - Hiroyuki Uchida
- Department of Applied Chemistry & Biotechnology; Graduate School of Engineering; University of Fukui; Fukui 910-8507; Japan
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Takahashi H, Sun X, Hamamoto M, Yashiroda Y, Yoshida M. The SAGA histone acetyltransferase complex regulates leucine uptake through the Agp3 permease in fission yeast. J Biol Chem 2012; 287:38158-67. [PMID: 22992726 DOI: 10.1074/jbc.m112.411165] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
Metabolic responses of unicellular organisms are mostly acute, transient, and cell-autonomous. Regulation of nutrient uptake in yeast is one such rapid response. High quality nitrogen sources such as NH(4)(+) inhibit uptake of poor nitrogen sources, such as amino acids. Both transcriptional and posttranscriptional mechanisms operate in nutrient uptake regulation; however, many components of this system remain uncharacterized in the fission yeast, Schizosaccharomyces pombe. Here, we demonstrate that the Spt-Ada-Gcn acetyltransferase (SAGA) complex modulates leucine uptake. Initially, we noticed that a branched-chain amino acid auxotroph exhibits a peculiar adaptive growth phenotype on solid minimal media containing certain nitrogen sources. In fact, the growth of many auxotrophic strains is inhibited by excess NH(4)Cl, possibly through nitrogen-mediated uptake inhibition of the corresponding nutrients. Surprisingly, DNA microarray analysis revealed that the transcriptional reprogramming during the adaptation of the branched-chain amino acid auxotroph was highly correlated with reprogramming observed in deletions of the SAGA histone acetyltransferase module genes. Deletion of gcn5(+) increased leucine uptake in the prototrophic background and rendered the leucine auxotroph resistant to NH(4)Cl. Deletion of tra1(+) caused the opposite phenotypes. The increase in leucine uptake in the gcn5Δ mutant was dependent on an amino acid permease gene, SPCC965.11c(+). The closest budding yeast homolog of this permease is a relatively nonspecific amino acid permease AGP3, which functions in poor nutrient conditions. Our analysis identified the regulation of nutrient uptake as a physiological function for the SAGA complex, providing a potential link between cellular metabolism and chromatin regulation.
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Affiliation(s)
- Hidekazu Takahashi
- Chemical Genetics Laboratory/Chemical Genomics Research Group, RIKEN Advanced Science Institute, Wako, Saitama 351-0198, Japan
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Helmlinger D. New insights into the SAGA complex from studies of the Tra1 subunit in budding and fission yeast. Transcription 2012; 3:13-8. [PMID: 22456315 DOI: 10.4161/trns.3.1.19271] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/19/2022] Open
Abstract
The SAGA complex is a conserved, multifunctional co-activator that controls the transcription of many inducible genes in response to environmental changes. Recent studies have provided new insights into the functions of one of its subunits, Tra1/TRRAP, and suggest that it controls SAGA activity in response to external stimuli.
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Affiliation(s)
- Dominique Helmlinger
- Macromolecular Biochemistry Research Center, CNRS UMR 5237, University of Montpellier 1 & 2, Montpellier, France.
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50
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The Schizosaccharomyces pombe inv1+ regulatory region is unusually large and contains redundant cis-acting elements that function in a SAGA- and Swi/Snf-dependent fashion. EUKARYOTIC CELL 2012; 11:1067-74. [PMID: 22707486 DOI: 10.1128/ec.00141-12] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/01/2023]
Abstract
The Schizosaccharomyces pombe inv1(+) gene encodes invertase, the enzyme required for hydrolysis of sucrose and raffinose. Transcription of inv1(+) is regulated by glucose levels, with transcription tightly repressed in high glucose and strongly induced in low glucose. To understand this regulation, we have analyzed the inv1(+) cis-regulatory region and the requirement for the trans-acting coactivators SAGA and Swi/Snf. Surprisingly, deletion of the entire 1-kilobase intergenic region between the inv1(+) TATA element and the upstream open reading frame SPCC191.10 does not significantly alter regulation of inv1(+) transcription. However, a longer deletion that extends through SPCC191.10 abolishes inv1(+) induction in low glucose. Additional analysis demonstrates that there are multiple, redundant regulatory regions spread over 1.5 kb 5' of inv1(+), including within SPCC191.10, that can confer glucose-mediated transcriptional regulation to inv1(+). Furthermore, SPCC191.10 can regulate inv1(+) transcription in an orientation-independent fashion and from a distance as great as 3 kb. With respect to trans-acting factors, both SAGA and Swi/Snf are recruited to SPCC191.10 and to other locations in the large inv1(+) regulatory region in a glucose-dependent fashion, and both are required for inv1(+) derepression. Taken together, these results demonstrate that inv1(+) regulation in S. pombe occurs via the use of multiple regulatory elements and that activation can occur over a great distance, even from elements within other open reading frames.
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