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1
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Adams-Brown SE, Reid KZ. The Central FacilitaTOR: Coordinating Transcription and Translation in Eukaryotes. Int J Mol Sci 2025; 26:2845. [PMID: 40243440 PMCID: PMC11989106 DOI: 10.3390/ijms26072845] [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: 01/31/2025] [Revised: 03/11/2025] [Accepted: 03/17/2025] [Indexed: 04/18/2025] Open
Abstract
One of the biggest challenges to eukaryotic gene expression is coordinating transcription in the nucleus and protein synthesis in the cytoplasm. However, little is known about how these major steps in gene expression are connected. The Target of Rapamycin (TOR) signaling pathway is crucial in connecting these critical phases of gene expression. Highly conserved among eukaryotic cells, TOR regulates growth, metabolism, and cellular equilibrium in response to changes in nutrients, energy levels, and stress conditions. This review examines the extensive role of TOR in gene expression regulation. We highlight how TOR is involved in phosphorylation, remodeling chromatin structure, and managing the factors that facilitate transcription and translation. Furthermore, the critical functions of TOR extend to processing RNA, assembling RNA-protein complexes, and managing their export from the nucleus, demonstrating its wide-reaching impact throughout the cell. Our discussion emphasizes the integral roles of TOR in bridging the processes of transcription and translation and explores how it orchestrates these complex cellular processes.
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Affiliation(s)
| | - Ke Zhang Reid
- Department of Biology, Wake Forest University, Winston-Salem, NC 27109, USA
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2
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Liu M, Li Y, Yuan X, Rong S, Du J. Novel insights into RNA polymerase II transcription regulation: transcription factors, phase separation, and their roles in cardiovascular diseases. Biochem Cell Biol 2025; 103:1-21. [PMID: 39540550 DOI: 10.1139/bcb-2024-0094] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2024] Open
Abstract
Transcription factors (TFs) are specialized proteins that bind DNA in a sequence-specific manner and modulate RNA polymerase II (Pol II) in multiple steps of the transcription process. Phase separation is a spontaneous or driven process that can form membrane-less organelles called condensates. By creating different liquid phases at active transcription sites, the formation of transcription condensates can reduce the water content of the condensate and lower the dielectric constant in biological systems, which in turn alters the structure and function of proteins and nucleic acids in the condensate. In RNA Pol II transcription, phase separation formation shortens the time at which TFs bind to target DNA sites and promotes transcriptional bursting. RNA Pol II transcription is engaged in developing several diseases, such as cardiovascular disease, by regulating different TFs and mediating the occurrence of phase separation. This review aims to summarize the advances in the molecular mechanisms of RNA Pol II transcriptional regulation, in particular the effect of TFs and phase separation. The role of RNA Pol II transcriptional regulation in cardiovascular disease will be elucidated, providing potential therapeutic targets for the management and treatment of cardiovascular disease.
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Affiliation(s)
- Mengmeng Liu
- Department of Cardiology, The Second Affiliated Hospital of Chongqing Medical University, Chongqing 400010, China
| | - Yingrui Li
- Department of Cardiology, The Second Affiliated Hospital of Chongqing Medical University, Chongqing 400010, China
| | - Xin Yuan
- Department of Nephrology, The Second Affiliated Hospital of Chongqing Medical University, Chongqing 4000l0, China
| | - Shunkang Rong
- Department of Cardiology, The Second Affiliated Hospital of Chongqing Medical University, Chongqing 400010, China
| | - Jianlin Du
- Department of Cardiology, The Second Affiliated Hospital of Chongqing Medical University, Chongqing 400010, China
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3
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Sun S, Yu D, Guo M, Tang M, Yan Z, Sun W, Wu A. The transcription factor FgSfp1 orchestrates mycotoxin deoxynivalenol biosynthesis in Fusarium graminearum. Commun Biol 2024; 7:1584. [PMID: 39604708 PMCID: PMC11603076 DOI: 10.1038/s42003-024-07265-4] [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: 03/16/2024] [Accepted: 11/14/2024] [Indexed: 11/29/2024] Open
Abstract
Fusarium graminearum (F. graminearum) and its derivative mycotoxin deoxynivalenol (DON) are highly concerned with food safety and sustainability worldwide. Although several transcription factors (TFs) had been elucidated, molecular mechanism participates in DON biosynthesis regulation remains largely unrevealed. Here, we first characterized a zinc finger-contained TF in F. graminearum, FgSfp1, which is indispensable for DON production since its depletion resulting in a 95.4% DON yielding reduction. Interestingly, contrast to previous knowledge, all TRI-cluster genes were abnormally upregulated in ΔFgSfp1 while Tri proteins abundance rationally decreased simultaneously. Further evidence show FgSfp1 could coordinate genetic translation pace by manipulating ribosomal biogenesis process. Specifically, FgSfp1-depletion leads to ribosome biogenesis assembly factor (RiBi) expression attenuation along with DON precursor acetyl-CoA synthase reduction since FgSfp1 actively interacts with RNA 2'-O-methylation enzyme FgNop1 revealed by Bi-FC. It subsequently influences mRNA translation pace. In conclusion, we elucidated that the FgSfp1 orchestrates DON biosynthesis via participating RNA posttranscriptional modification for ribosomal RNA maturation, offering insights into the DON biosynthesis regulation. Ultimately, this TF might be a key regulator for DON contamination control in the whole food chain.
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Affiliation(s)
- Shuting Sun
- CAS Key Laboratory of Nutrition, Metabolism and Food Safety, Shanghai Institute of Nutrition and Health, University of Chinese Academy of Sciences, Chinese Academy of Sciences, Shanghai, China
| | - Dianzhen Yu
- CAS Key Laboratory of Nutrition, Metabolism and Food Safety, Shanghai Institute of Nutrition and Health, University of Chinese Academy of Sciences, Chinese Academy of Sciences, Shanghai, China
| | - Mingzhu Guo
- CAS Key Laboratory of Nutrition, Metabolism and Food Safety, Shanghai Institute of Nutrition and Health, University of Chinese Academy of Sciences, Chinese Academy of Sciences, Shanghai, China
| | - Muhai Tang
- CAS Key Laboratory of Nutrition, Metabolism and Food Safety, Shanghai Institute of Nutrition and Health, University of Chinese Academy of Sciences, Chinese Academy of Sciences, Shanghai, China
| | - Zheng Yan
- CAS Key Laboratory of Nutrition, Metabolism and Food Safety, Shanghai Institute of Nutrition and Health, University of Chinese Academy of Sciences, Chinese Academy of Sciences, Shanghai, China
| | - Wei Sun
- CAS Key Laboratory of Nutrition, Metabolism and Food Safety, Shanghai Institute of Nutrition and Health, University of Chinese Academy of Sciences, Chinese Academy of Sciences, Shanghai, China
| | - Aibo Wu
- CAS Key Laboratory of Nutrition, Metabolism and Food Safety, Shanghai Institute of Nutrition and Health, University of Chinese Academy of Sciences, Chinese Academy of Sciences, Shanghai, China.
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4
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Bi X. Hmo1: A versatile member of the high mobility group box family of chromosomal architecture proteins. World J Biol Chem 2024; 15:97938. [PMID: 39156122 PMCID: PMC11325855 DOI: 10.4331/wjbc.v15.i1.97938] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/13/2024] [Revised: 07/21/2024] [Accepted: 08/01/2024] [Indexed: 08/08/2024] Open
Abstract
Eukaryotic chromatin consisting of nucleosomes connected by linker DNA is organized into higher order structures, which is facilitated by linker histone H1. Formation of chromatin compacts and protects the genome, but also hinders DNA transactions. Cells have evolved mechanisms to modify/remodel chromatin resulting in chromatin states suitable for genome functions. The high mobility group box (HMGB) proteins are non-histone chromatin architectural factors characterized by one or more HMGB motifs that bind DNA in a sequence nonspecific fashion. They play a major role in chromatin dynamics. The Saccharomyces cerevisiae (yeast hereafter) HMGB protein Hmo1 contains two HMGB motifs. However, unlike a canonical HMGB protein that has an acidic C-terminus, Hmo1 ends with a lysine rich, basic, C-terminus, resembling linker histone H1. Hmo1 exhibits characteristics of both HMGB proteins and linker histones in its multiple functions. For instance, Hmo1 promotes transcription by RNA polymerases I and II like canonical HMGB proteins but makes chromatin more compact/stable like linker histones. Recent studies have demonstrated that Hmo1 destabilizes/disrupts nucleosome similarly as other HMGB proteins in vitro and acts to maintain a common topological architecture of genes in yeast genome. This minireview reviews the functions of Hmo1 and the underlying mechanisms, highlighting recent discoveries.
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Affiliation(s)
- Xin Bi
- Department of Biology, University of Rochester, Rochester, NY 14627, United States
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5
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Patil S, Cremosnik G, Dötsch L, Flegel J, Schulte B, Maier KC, Žumer K, Cramer P, Janning P, Sievers S, Ziegler S, Waldmann H. The Pseudo-Natural Product Tafbromin Selectively Targets the TAF1 Bromodomain 2. Angew Chem Int Ed Engl 2024; 63:e202404645. [PMID: 38801173 DOI: 10.1002/anie.202404645] [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: 03/07/2024] [Revised: 05/17/2024] [Accepted: 05/23/2024] [Indexed: 05/29/2024]
Abstract
Phenotypic assays detect small-molecule bioactivity at functionally relevant cellular sites, and inherently cover a variety of targets and mechanisms of action. They can uncover new small molecule-target pairs and may give rise to novel biological insights. By means of an osteoblast differentiation assay which employs a Hedgehog (Hh) signaling agonist as stimulus and which monitors an endogenous marker for osteoblasts, we identified a pyrrolo[3,4-g]quinoline (PQ) pseudo-natural product (PNP) class of osteogenesis inhibitors. The most potent PQ, termed Tafbromin, impairs canonical Hh signaling and modulates osteoblast differentiation through binding to the bromodomain 2 of the TATA-box binding protein-associated factor 1 (TAF1). Tafbromin is the most selective TAF1 bromodomain 2 ligand and promises to be an invaluable tool for the study of biological processes mediated by TAF1(2) bromodomains.
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Affiliation(s)
- Sohan Patil
- Max-Planck Institute of Molecular Physiology, Department of Chemical Biology, Otto-Hahn-Strasse 11, Dortmund, 44227, Germany
| | - Gregor Cremosnik
- Max-Planck Institute of Molecular Physiology, Department of Chemical Biology, Otto-Hahn-Strasse 11, Dortmund, 44227, Germany
| | - Lara Dötsch
- Max-Planck Institute of Molecular Physiology, Department of Chemical Biology, Otto-Hahn-Strasse 11, Dortmund, 44227, Germany
- Technical University Dortmund, Faculty of Chemistry and Chemical Biology, Otto-Hahn-Strasse 6, Dortmund, 44227, Germany
| | - Jana Flegel
- Max-Planck Institute of Molecular Physiology, Department of Chemical Biology, Otto-Hahn-Strasse 11, Dortmund, 44227, Germany
| | - Britta Schulte
- Max-Planck Institute of Molecular Physiology, Department of Chemical Biology, Otto-Hahn-Strasse 11, Dortmund, 44227, Germany
| | - Kerstin C Maier
- Max Planck Institute for Multidisciplinary Sciences, Department of Molecular Biology, Am Fassberg 11, 37077, Göttingen, Germany
| | - Kristina Žumer
- Max Planck Institute for Multidisciplinary Sciences, Department of Molecular Biology, Am Fassberg 11, 37077, Göttingen, Germany
| | - Patrick Cramer
- Max Planck Institute for Multidisciplinary Sciences, Department of Molecular Biology, Am Fassberg 11, 37077, Göttingen, Germany
| | - Petra Janning
- Max-Planck Institute of Molecular Physiology, Department of Chemical Biology, Otto-Hahn-Strasse 11, Dortmund, 44227, Germany
| | - Sonja Sievers
- Max-Planck Institute of Molecular Physiology, Department of Chemical Biology, Otto-Hahn-Strasse 11, Dortmund, 44227, Germany
| | - Slava Ziegler
- Max-Planck Institute of Molecular Physiology, Department of Chemical Biology, Otto-Hahn-Strasse 11, Dortmund, 44227, Germany
| | - Herbert Waldmann
- Max-Planck Institute of Molecular Physiology, Department of Chemical Biology, Otto-Hahn-Strasse 11, Dortmund, 44227, Germany
- Technical University Dortmund, Faculty of Chemistry and Chemical Biology, Otto-Hahn-Strasse 6, Dortmund, 44227, Germany
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Wang M, Li J, Wang Y, Fu H, Qiu H, Li Y, Li M, Lu Y, Fu YV. Single-molecule study reveals Hmo1, not Hho1, promotes chromatin assembly in budding yeast. mBio 2023; 14:e0099323. [PMID: 37432033 PMCID: PMC10470511 DOI: 10.1128/mbio.00993-23] [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: 05/09/2023] [Accepted: 05/25/2023] [Indexed: 07/12/2023] Open
Abstract
Linker histone H1 plays a crucial role in various biological processes, including nucleosome stabilization, high-order chromatin structure organization, gene expression, and epigenetic regulation in eukaryotic cells. Unlike higher eukaryotes, little about the linker histone in Saccharomyces cerevisiae is known. Hho1 and Hmo1 are two long-standing controversial histone H1 candidates in budding yeast. In this study, we directly observed at the single-molecule level that Hmo1, but not Hho1, is involved in chromatin assembly in the yeast nucleoplasmic extracts (YNPE), which can replicate the physiological condition of the yeast nucleus. The presence of Hmo1 facilitates the assembly of nucleosomes on DNA in YNPE, as revealed by single-molecule force spectroscopy. Further single-molecule analysis showed that the lysine-rich C-terminal domain (CTD) of Hmo1 is essential for the function of chromatin compaction, while the second globular domain at the C-terminus of Hho1 impairs its ability. In addition, Hmo1, but not Hho1, forms condensates with double-stranded DNA via reversible phase separation. The phosphorylation fluctuation of Hmo1 coincides with metazoan H1 during the cell cycle. Our data suggest that Hmo1, but not Hho1, possesses some functionality similar to that of linker histone in Saccharomyces cerevisiae, even though some properties of Hmo1 differ from those of a canonical linker histone H1. Our study provides clues for the linker histone H1 in budding yeast and provides insights into the evolution and diversity of histone H1 across eukaryotes. IMPORTANCE There has been a long-standing debate regarding the identity of linker histone H1 in budding yeast. To address this issue, we utilized YNPE, which accurately replicate the physiological conditions in yeast nuclei, in combination with total internal reflection fluorescence microscopy and magnetic tweezers. Our findings demonstrated that Hmo1, rather than Hho1, is responsible for chromatin assembly in budding yeast. Additionally, we found that Hmo1 shares certain characteristics with histone H1, including phase separation and phosphorylation fluctuations throughout the cell cycle. Furthermore, we discovered that the lysine-rich domain of Hho1 is buried by its second globular domain at the C-terminus, resulting in the loss of function that is similar to histone H1. Our study provides compelling evidence to suggest that Hmo1 shares linker histone H1 function in budding yeast and contributes to our understanding of the evolution of linker histone H1 across eukaryotes.
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Affiliation(s)
- Mengxue Wang
- State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China
- College of Life Sciences, University of Chinese Academy of Sciences, Beijing, China
| | - Jinghua Li
- Beijing National Laboratory for Condensed Matter Physics, Institute of Physics, Chinese Academy of Sciences, Beijing, China
| | - Yong Wang
- State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China
- College of Life Sciences, University of Chinese Academy of Sciences, Beijing, China
| | - Hang Fu
- Wenzhou Institute, University of Chinese Academy of Sciences, Wenzhou, Zhejiang, China
- School of Physics, University of Chinese Academy of Sciences, Beijing, China
| | - Haoning Qiu
- State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China
- College of Life Sciences, University of Chinese Academy of Sciences, Beijing, China
| | - Yanying Li
- State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China
- College of Life Sciences, University of Chinese Academy of Sciences, Beijing, China
| | - Ming Li
- Beijing National Laboratory for Condensed Matter Physics, Institute of Physics, Chinese Academy of Sciences, Beijing, China
- School of Physics, University of Chinese Academy of Sciences, Beijing, China
- Songshan Lake Materials Laboratory, Dongguan, Guangdong, China
| | - Ying Lu
- Beijing National Laboratory for Condensed Matter Physics, Institute of Physics, Chinese Academy of Sciences, Beijing, China
- School of Physics, University of Chinese Academy of Sciences, Beijing, China
- Songshan Lake Materials Laboratory, Dongguan, Guangdong, China
| | - Yu Vincent Fu
- State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China
- Savaid Medical School, University of Chinese Academy of Sciences, Beijing, China
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7
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Franco-Losilla M, Nordzieke S, Feldmann I, Limón MC, Avalos J. HmbC, a Protein of the HMG Family, Participates in the Regulation of Carotenoid Biosynthesis in Fusarium fujikuroi. Genes (Basel) 2023; 14:1661. [PMID: 37628712 PMCID: PMC10454146 DOI: 10.3390/genes14081661] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/15/2023] [Revised: 08/12/2023] [Accepted: 08/16/2023] [Indexed: 08/27/2023] Open
Abstract
In the fungus Fusarium fujikuroi, carotenoid production is up-regulated by light and down-regulated by the CarS RING finger protein, which modulates the mRNA levels of carotenoid pathway genes (car genes). To identify new potential regulators of car genes, we used a biotin-mediated pull-down procedure to detect proteins capable of binding to their promoters. We focused our attention on one of the proteins found in the screening, belonging to the High-Mobility Group (HMG) family that was named HmbC. The deletion of the hmbC gene resulted in increased carotenoid production due to higher mRNA levels of car biosynthetic genes. In addition, the deletion resulted in reduced carS mRNA levels, which could also explain the partial deregulation of the carotenoid pathway. The mutants exhibited other phenotypic traits, such as alterations in development under certain stress conditions, or reduced sensitivity to cell wall degrading enzymes, revealed by less efficient protoplast formation, indicating that HmbC is also involved in other cellular processes. In conclusion, we identified a protein of the HMG family that participates in the regulation of carotenoid biosynthesis. This is probably achieved through an epigenetic mechanism related to chromatin structure, as is frequent in this class of proteins.
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Affiliation(s)
- Marta Franco-Losilla
- Departamento de Genética, Facultad de Biología, Universidad de Sevilla, 41012 Sevilla, Spain; (M.F.-L.); (J.A.)
| | - Steffen Nordzieke
- Departamento de Genética, Facultad de Biología, Universidad de Sevilla, 41012 Sevilla, Spain; (M.F.-L.); (J.A.)
| | - Ingo Feldmann
- Leibniz-Institut für Analytische Wissenschaften—ISAS—e.V., 44227 Dortmund, Germany;
| | - M. Carmen Limón
- Departamento de Genética, Facultad de Biología, Universidad de Sevilla, 41012 Sevilla, Spain; (M.F.-L.); (J.A.)
| | - Javier Avalos
- Departamento de Genética, Facultad de Biología, Universidad de Sevilla, 41012 Sevilla, Spain; (M.F.-L.); (J.A.)
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Kumar S, Mashkoor M, Grove A. Yeast Crf1p: An activator in need is an activator indeed. Comput Struct Biotechnol J 2022; 20:107-116. [PMID: 34976315 PMCID: PMC8688861 DOI: 10.1016/j.csbj.2021.12.003] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/28/2021] [Revised: 11/15/2021] [Accepted: 12/03/2021] [Indexed: 11/10/2022] Open
Abstract
Ribosome biogenesis is an energetically costly process, and tight regulation is required for stoichiometric balance between components. This requires coordination of RNA polymerases I, II, and III. Lack of nutrients or the presence of stress leads to downregulation of ribosome biogenesis, a process for which mechanistic target of rapamycin complex I (mTORC1) is key. mTORC1 activity is communicated by means of specific transcription factors, and in yeast, which is a primary model system in which transcriptional coordination has been delineated, transcription factors involved in regulation of ribosomal protein genes include Fhl1p and its cofactors, Ifh1p and Crf1p. Ifh1p is an activator, whereas Crf1p has been implicated in maintaining the repressed state upon mTORC1 inhibition. Computational analyses of evolutionary relationships have indicated that Ifh1p and Crf1p descend from a common ancestor. Here, we discuss recent evidence, which suggests that Crf1p also functions as an activator. We propose a model that consolidates available experimental evidence, which posits that Crf1p functions as an alternate activator to prevent the stronger activator Ifh1p from re-binding gene promoters upon mTORC1 inhibition. The correlation between retention of Crf1p in related yeast strains and duplication of ribosomal protein genes suggests that this backup activation may be important to ensure gene expression when Ifh1p is limiting. With ribosome biogenesis as a hallmark of cell growth, failure to control assembly of ribosomal components leads to several human pathologies. A comprehensive understanding of mechanisms underlying this process is therefore of the essence.
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Key Words
- CK2, casein kinase 2
- Crf1, corepressor with forkhead like
- Crf1p
- FHA, forkhead-associated
- FHB, forkhead-binding
- FKBP, FK506 binding protein
- Fhl1, forkhead like
- Fpr1, FK506-sensitive proline rotamase
- Gene regulation
- Hmo1, high mobility group
- Ifh1, interacts with forkhead like
- Ifh1p
- RASTR, ribosome assembly stress response
- RP, ribosomal protein
- Rap1, repressor/activator protein
- RiBi, ribosome biogenesis
- Ribosomal protein
- Ribosome biogenesis
- Sfp1, split finger protein
- WGD, whole genome duplication
- mTORC1
- mTORC1, mechanistic target of rapamycin complex 1
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Affiliation(s)
- Sanjay Kumar
- Department of Biological Sciences, Louisiana State University, Baton Rouge, LA 70803, USA
| | - Muneera Mashkoor
- Department of Biological Sciences, Louisiana State University, Baton Rouge, LA 70803, USA
| | - Anne Grove
- Department of Biological Sciences, Louisiana State University, Baton Rouge, LA 70803, USA
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9
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Amigo R, Farkas C, Gidi C, Hepp MI, Cartes N, Tarifeño E, Workman JL, Gutiérrez JL. The linker histone Hho1 modulates the activity of ATP-dependent chromatin remodeling complexes. BIOCHIMICA ET BIOPHYSICA ACTA. GENE REGULATORY MECHANISMS 2022; 1865:194781. [PMID: 34963628 DOI: 10.1016/j.bbagrm.2021.194781] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 06/05/2021] [Revised: 11/30/2021] [Accepted: 12/12/2021] [Indexed: 02/07/2023]
Abstract
Diverse factors play roles in chromatin dynamics, including linker proteins. Among them are high mobility group (HMG) box family proteins and linker histones. In the yeast Saccharomyces cerevisiae, Hmo1 has been identified as an HMG-box protein. This protein displays properties that are in agreement with this allocation. However, a number of studies have postulated that Hmo1 functions as a linker histone in yeast. On the other hand, when discovered, the Hho1 protein was identified as a linker histone. While multiple studies support this classification, some findings point to characteristics of Hho1 that are dissimilar to those commonly assigned to linker histones. In order to better understand the roles played by Hmo1 and Hho1 in chromatin dynamics and transcriptional regulation, we performed several analyses directly comparing these two proteins. Our analyses of genome-wide binding profiles support the belonging of Hmo1 to the HMGB family and Hho1 to the linker histones family. Interestingly, by performing protein-protein interaction analyses we found that both Hmo1 and Hho1 display physical interaction with the ATP-dependent chromatin remodeling complexes RSC, ISW1a and SWI/SNF. Moreover, by carrying out nucleosome remodeling assays, we found that both proteins stimulate the activity of the ISW1a complex. However, in the case of RSC, Hmo1 and Hho1 displayed differential properties, with Hho1 mainly showing an inhibitory effect. Our results are in agreement with the opposite roles played by RSC and ISW1a in chromatin dynamics and transcriptional regulation, and expand the view for the roles played by Hho1 and linker histones.
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Affiliation(s)
- Roberto Amigo
- Departamento de Bioquímica y Biología Molecular, Facultad de Ciencias Biológicas, Universidad de Concepción, Barrio Universitario s/n, Concepción 4070043, Chile
| | - Carlos Farkas
- Departamento de Bioquímica y Biología Molecular, Facultad de Ciencias Biológicas, Universidad de Concepción, Barrio Universitario s/n, Concepción 4070043, Chile
| | - Cristian Gidi
- Departamento de Bioquímica y Biología Molecular, Facultad de Ciencias Biológicas, Universidad de Concepción, Barrio Universitario s/n, Concepción 4070043, Chile
| | - Matias I Hepp
- Departamento de Bioquímica y Biología Molecular, Facultad de Ciencias Biológicas, Universidad de Concepción, Barrio Universitario s/n, Concepción 4070043, Chile
| | - Natalia Cartes
- Departamento de Bioquímica y Biología Molecular, Facultad de Ciencias Biológicas, Universidad de Concepción, Barrio Universitario s/n, Concepción 4070043, Chile
| | - Estefanía Tarifeño
- Departamento de Bioquímica y Biología Molecular, Facultad de Ciencias Biológicas, Universidad de Concepción, Barrio Universitario s/n, Concepción 4070043, Chile
| | - Jerry L Workman
- Stowers Institute for Medical Research, 1000 E 50(th) Street, Kansas City, MO 64110, USA
| | - José L Gutiérrez
- Departamento de Bioquímica y Biología Molecular, Facultad de Ciencias Biológicas, Universidad de Concepción, Barrio Universitario s/n, Concepción 4070043, Chile.
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10
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Transcriptional control of ribosome biogenesis in yeast: links to growth and stress signals. Biochem Soc Trans 2021; 49:1589-1599. [PMID: 34240738 PMCID: PMC8421047 DOI: 10.1042/bst20201136] [Citation(s) in RCA: 29] [Impact Index Per Article: 7.3] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/10/2021] [Revised: 06/14/2021] [Accepted: 06/18/2021] [Indexed: 12/15/2022]
Abstract
Ribosome biogenesis requires prodigious transcriptional output in rapidly growing yeast cells and is highly regulated in response to both growth and stress signals. This minireview focuses on recent developments in our understanding of this regulatory process, with an emphasis on the 138 ribosomal protein genes (RPGs) themselves and a group of >200 ribosome biogenesis (RiBi) genes whose products contribute to assembly but are not part of the ribosome. Expression of most RPGs depends upon Rap1, a pioneer transcription factor (TF) required for the binding of a pair of RPG-specific TFs called Fhl1 and Ifh1. RPG expression is correlated with Ifh1 promoter binding, whereas Rap1 and Fhl1 remain promoter-associated upon stress-induced down regulation. A TF called Sfp1 has also been implicated in RPG regulation, though recent work reveals that its primary function is in activation of RiBi and other growth-related genes. Sfp1 plays an important regulatory role at a small number of RPGs where Rap1–Fhl1–Ifh1 action is subsidiary or non-existent. In addition, nearly half of all RPGs are bound by Hmo1, which either stabilizes or re-configures Fhl1–Ifh1 binding. Recent studies identified the proline rotamase Fpr1, known primarily for its role in rapamycin-mediated inhibition of the TORC1 kinase, as an additional TF at RPG promoters. Fpr1 also affects Fhl1–Ifh1 binding, either independently or in cooperation with Hmo1. Finally, a major recent development was the discovery of a protein homeostasis mechanism driven by unassembled ribosomal proteins, referred to as the Ribosome Assembly Stress Response (RASTR), that controls RPG transcription through the reversible condensation of Ifh1.
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11
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Kasahara K, Nakayama R, Shiwa Y, Kanesaki Y, Ishige T, Yoshikawa H, Kokubo T. Fpr1, a primary target of rapamycin, functions as a transcription factor for ribosomal protein genes cooperatively with Hmo1 in Saccharomyces cerevisiae. PLoS Genet 2020; 16:e1008865. [PMID: 32603360 PMCID: PMC7357790 DOI: 10.1371/journal.pgen.1008865] [Citation(s) in RCA: 12] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/27/2019] [Revised: 07/13/2020] [Accepted: 05/15/2020] [Indexed: 11/18/2022] Open
Abstract
Fpr1 (FK506-sensitive proline rotamase 1), a protein of the FKBP12 (FK506-binding protein 12 kDa) family in Saccharomyces cerevisiae, is a primary target for the immunosuppressive agents FK506 and rapamycin. Fpr1 inhibits calcineurin and TORC1 (target of rapamycin complex 1) when bound to FK506 and rapamycin, respectively. Although Fpr1 is recognised to play a crucial role in the efficacy of these drugs, its physiological functions remain unclear. In a hmo1Δ (high mobility group family 1-deleted) yeast strain, deletion of FPR1 induced severe growth defects, which could be alleviated by increasing the copy number of RPL25 (ribosome protein of the large subunit 25), suggesting that RPL25 expression was affected in hmo1Δfpr1Δ cells. In the current study, extensive chromatin immunoprecipitation (ChIP) and ChIP-sequencing analyses revealed that Fpr1 associates specifically with the upstream activating sequences of nearly all RPG (ribosomal protein gene) promoters, presumably in a manner dependent on Rap1 (repressor/activator site binding protein 1). Intriguingly, Fpr1 promotes the binding of Fhl1/Ifh1 (forkhead-like 1/interacts with forkhead 1), two key regulators of RPG transcription, to certain RPG promoters independently of and/or cooperatively with Hmo1. Furthermore, mutation analyses of Fpr1 indicated that for transcriptional function on RPG promoters, Fpr1 requires its N-terminal domain and the binding surface for rapamycin, but not peptidyl-prolyl isomerase activity. Notably, Fpr1 orthologues from other species also inhibit TORC1 when bound to rapamycin, but do not regulate transcription in yeast, which suggests that these two functions of Fpr1 are independent of each other.
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Affiliation(s)
- Koji Kasahara
- Department of Molecular Microbiology, Tokyo University of Agriculture, Tokyo, Japan
- * E-mail:
| | - Risa Nakayama
- Department of Bioscience, Tokyo University of Agriculture, Tokyo, Japan
| | - Yuh Shiwa
- Department of Molecular Microbiology, Tokyo University of Agriculture, Tokyo, Japan
| | - Yu Kanesaki
- Research Institute of Green Science and Technology, Shizuoka University, Shizuoka, Japan
| | - Taichiro Ishige
- NODAI Genome Research Center, Tokyo University of Agriculture, Tokyo, Japan
| | | | - Tetsuro Kokubo
- Graduate School of Medical Life Science, Yokohama City University, Yokohama, Japan
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12
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African trypanosomes expressing multiple VSGs are rapidly eliminated by the host immune system. Proc Natl Acad Sci U S A 2019; 116:20725-20735. [PMID: 31554700 PMCID: PMC6789922 DOI: 10.1073/pnas.1905120116] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/18/2022] Open
Abstract
Many parasites escape the host immune system by undergoing antigenic variation, a process in which surface antigens are regularly shed and replaced by new ones. Trypanosoma brucei employs multiple sophisticated molecular mechanisms to ensure the expression of a homogeneous VSG coat. We generated a mutant parasite that expresses multiple distinct VSGs and studied the consequences of having a multi-VSG coat during an infection. We showed that expression of multiple VSGs makes the parasites more vulnerable to the immune response, which can now control the trypanosomes from the onset of the infection, allowing most mice to survive. In the future, trypanosome infections may be treated using drugs that generate parasites with multi-VSG coats. Trypanosoma brucei parasites successfully evade the host immune system by periodically switching the dense coat of variant surface glycoprotein (VSG) at the cell surface. Each parasite expresses VSGs in a monoallelic fashion that is tightly regulated. The consequences of exposing multiple VSGs during an infection, in terms of antibody response and disease severity, remain unknown. In this study, we overexpressed a high-mobility group box protein, TDP1, which was sufficient to open the chromatin of silent VSG expression sites, to disrupt VSG monoallelic expression, and to generate viable and healthy parasites with a mixed VSG coat. Mice infected with these parasites mounted a multi-VSG antibody response, which rapidly reduced parasitemia. Consequently, we observed prolonged survival in which nearly 90% of the mice survived a 30-d period of infection with undetectable parasitemia. Immunodeficient RAG2 knock-out mice were unable to control infection with TDP1-overexpressing parasites, showing that the adaptive immune response is critical to reducing disease severity. This study shows that simultaneous exposure of multiple VSGs is highly detrimental to the parasite, even at the very early stages of infection, suggesting that drugs that disrupt VSG monoallelic expression could be used to treat trypanosomiasis.
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13
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Bokor E, Ámon J, Keisham K, Karácsony Z, Vágvölgyi C, Hamari Z. HMGB proteins are required for sexual development in Aspergillus nidulans. PLoS One 2019; 14:e0216094. [PMID: 31022275 PMCID: PMC6483251 DOI: 10.1371/journal.pone.0216094] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/14/2019] [Accepted: 04/12/2019] [Indexed: 11/18/2022] Open
Abstract
Aspergillus nidulans has three high mobility group box (HMGB) proteins, HmbA, HmbB and HmbC that are chromatin-associated architectural proteins involved in DNA-related functions. By creating and studying deletion strains in both veA+ and veA1 background, we have characterized the role of HmbA, HmbB and HmbC in sexual development. Expression of the mating-type MAT1-1 and MAT1-2 coding genes were found to be extremely down-regulated in all three mutants on day 4 of sexual development, which results in deficient ascospore production and/or ascospore viability in the mutants. In addition, we found that HmbA and HmbB play also a role in sensing of and response to environmental signals, while HmbC functionally interacts with VeA, a key regulator of the coordination of asexual and sexual development, as well as of secondary metabolism.
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Affiliation(s)
- Eszter Bokor
- University of Szeged, Faculty of Science and Informatics, Department of Microbiology, Szeged, Hungary
| | - Judit Ámon
- University of Szeged, Faculty of Science and Informatics, Department of Microbiology, Szeged, Hungary
| | - Kabichandra Keisham
- University of Szeged, Faculty of Science and Informatics, Department of Microbiology, Szeged, Hungary
| | - Zoltán Karácsony
- University of Szeged, Faculty of Science and Informatics, Department of Microbiology, Szeged, Hungary
| | - Csaba Vágvölgyi
- University of Szeged, Faculty of Science and Informatics, Department of Microbiology, Szeged, Hungary
| | - Zsuzsanna Hamari
- University of Szeged, Faculty of Science and Informatics, Department of Microbiology, Szeged, Hungary
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14
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Kozlova AL, Valieva ME, Maluchenko NV, Studitsky VM. HMGB Proteins as DNA Chaperones That Modulate Chromatin Activity. Mol Biol 2018. [DOI: 10.1134/s0026893318050096] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/28/2022]
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15
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HMGB proteins involved in TOR signaling as general regulators of cell growth by controlling ribosome biogenesis. Curr Genet 2018; 64:1205-1213. [PMID: 29713761 DOI: 10.1007/s00294-018-0842-8] [Citation(s) in RCA: 18] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/13/2018] [Revised: 04/21/2018] [Accepted: 04/24/2018] [Indexed: 12/14/2022]
Abstract
The number of ribosomes and their activity need to be highly regulated because their function is crucial for the cell. Ribosome biogenesis is necessary for cell growth and proliferation in accordance with nutrient availability and other external and intracellular signals. High-mobility group B (HMGB) proteins are conserved from yeasts to human and are decisive in cellular fate. These proteins play critical functions, from the maintenance of chromatin structure, DNA repair, or transcriptional regulation, to facilitation of ribosome biogenesis. They are also involved in cancer and other pathologies. In this review, we summarize evidence of how HMGB proteins contribute to ribosome-biogenesis control, with special emphasis on a common nexus to the target of rapamycin (TOR) pathway, a signaling cascade essential for cell growth and proliferation from yeast to human. Perspectives in this field are also discussed.
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16
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Johnson AN, Weil PA. Identification of a transcriptional activation domain in yeast repressor activator protein 1 (Rap1) using an altered DNA-binding specificity variant. J Biol Chem 2017; 292:5705-5723. [PMID: 28196871 PMCID: PMC5392566 DOI: 10.1074/jbc.m117.779181] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/30/2017] [Revised: 02/13/2017] [Indexed: 01/06/2023] Open
Abstract
Repressor activator protein 1 (Rap1) performs multiple vital cellular functions in the budding yeast Saccharomyces cerevisiae These include regulation of telomere length, transcriptional repression of both telomere-proximal genes and the silent mating type loci, and transcriptional activation of hundreds of mRNA-encoding genes, including the highly transcribed ribosomal protein- and glycolytic enzyme-encoding genes. Studies of the contributions of Rap1 to telomere length regulation and transcriptional repression have yielded significant mechanistic insights. However, the mechanism of Rap1 transcriptional activation remains poorly understood because Rap1 is encoded by a single copy essential gene and is involved in many disparate and essential cellular functions, preventing easy interpretation of attempts to directly dissect Rap1 structure-function relationships. Moreover, conflicting reports on the ability of Rap1-heterologous DNA-binding domain fusion proteins to serve as chimeric transcriptional activators challenge use of this approach to study Rap1. Described here is the development of an altered DNA-binding specificity variant of Rap1 (Rap1AS). We used Rap1AS to map and characterize a 41-amino acid activation domain (AD) within the Rap1 C terminus. We found that this AD is required for transcription of both chimeric reporter genes and authentic chromosomal Rap1 enhancer-containing target genes. Finally, as predicted for a bona fide AD, mutation of this newly identified AD reduced the efficiency of Rap1 binding to a known transcriptional coactivator TFIID-binding target, Taf5. In summary, we show here that Rap1 contains an AD required for Rap1-dependent gene transcription. The Rap1AS variant will likely also be useful for studies of the functions of Rap1 in other biological pathways.
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Affiliation(s)
- Amanda N Johnson
- From the Department of Molecular Physiology and Biophysics, Vanderbilt University School of Medicine, Nashville, Tennessee 37232
| | - P Anthony Weil
- From the Department of Molecular Physiology and Biophysics, Vanderbilt University School of Medicine, Nashville, Tennessee 37232
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17
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Abstract
Eukaryotic genomes are packaged in chromatin. The higher-order organization of nucleosome core particles is controlled by the association of the intervening linker DNA with either the linker histone H1 or high mobility group box (HMGB) proteins. While H1 is thought to stabilize the nucleosome by preventing DNA unwrapping, the DNA bending imposed by HMGB may propagate to the nucleosome to destabilize chromatin. For metazoan H1, chromatin compaction requires its lysine-rich C-terminal domain, a domain that is buried between globular domains in the previously characterized yeast Saccharomyces cerevisiae linker histone Hho1p. Here, we discuss the functions of S. cerevisiae HMO1, an HMGB family protein unique in containing a terminal lysine-rich domain and in stabilizing genomic DNA. On ribosomal DNA (rDNA) and genes encoding ribosomal proteins, HMO1 appears to exert its role primarily by stabilizing nucleosome-free regions or "fragile" nucleosomes. During replication, HMO1 likewise appears to ensure low nucleosome density at DNA junctions associated with the DNA damage response or the need for topoisomerases to resolve catenanes. Notably, HMO1 shares with the mammalian linker histone H1 the ability to stabilize chromatin, as evidenced by the absence of HMO1 creating a more dynamic chromatin environment that is more sensitive to nuclease digestion and in which chromatin-remodeling events associated with DNA double-strand break repair occur faster; such chromatin stabilization requires the lysine-rich extension of HMO1. Thus, HMO1 appears to have evolved a unique linker histone-like function involving the ability to stabilize both conventional nucleosome arrays as well as DNA regions characterized by low nucleosome density or the presence of noncanonical nucleosomes.
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18
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Kasahara K, Higashino A, Unzai S, Yoshikawa H, Kokubo T. Oligomerization of Hmo1 mediated by box A is essential for DNA binding in vitro and in vivo. Genes Cells 2016; 21:1333-1352. [PMID: 27860073 DOI: 10.1111/gtc.12449] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/11/2016] [Accepted: 10/05/2016] [Indexed: 11/30/2022]
Abstract
Hmo1, a member of HMGB family proteins in Saccharomyces cerevisiae, binds to and regulates the transcription of genes encoding ribosomal RNA and ribosomal proteins. The functional motifs of Hmo1 include two HMG-like motifs, box A and box B, and a C-terminal tail. To elucidate the molecular roles of the HMG-like boxes in DNA binding in vivo, we analyzed the DNA-binding activity of various Hmo1 mutants using ChIP or reporter assays that enabled us to conveniently detect Hmo1 binding to the promoter of RPS5, a major target gene of Hmo1. Our mutational analyses showed that box B is a bona fide DNA-binding motif and that it also plays other important roles in cell growth. However, box A, especially its first α-helix, contributes to DNA binding of Hmo1 by inducing self-assembly of Hmo1. Intriguingly, box A mediated formation of oligomers of more than two proteins on DNA in vivo. Furthermore, duplication of the box B partially alleviates the requirement for box A. These findings suggest that the principal role of box A is to assemble multiple box B in the appropriate orientation, thereby stabilizing the binding of Hmo1 to DNA and nucleating specific chromosomal architecture on its target genes.
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Affiliation(s)
- Koji Kasahara
- Isotope Center, Tokyo University of Agriculture, Setagaya, Tokyo, Japan
| | - Ayako Higashino
- Department of Bioscience, Tokyo University of Agriculture, Tokyo, Japan
| | - Satoru Unzai
- Department of Frontier Bioscience, Hosei University, Koganei, Tokyo, Japan
| | | | - Tetsuro Kokubo
- Graduate School of Medical Life Science, Yokohama City University, Yokohama, Kanagawa, Japan
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19
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Zhang Y, Najmi SM, Schneider DA. Transcription factors that influence RNA polymerases I and II: To what extent is mechanism of action conserved? BIOCHIMICA ET BIOPHYSICA ACTA-GENE REGULATORY MECHANISMS 2016; 1860:246-255. [PMID: 27989933 DOI: 10.1016/j.bbagrm.2016.10.010] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 08/05/2016] [Revised: 10/07/2016] [Accepted: 10/25/2016] [Indexed: 01/05/2023]
Abstract
In eukaryotic cells, nuclear RNA synthesis is accomplished by at least three unique, multisubunit RNA polymerases. The roles of these enzymes are generally partitioned into the synthesis of the three major classes of RNA: rRNA, mRNA, and tRNA for RNA polymerases I, II, and III respectively. Consistent with their unique cellular roles, each enzyme has a complement of specialized transcription factors and enzymatic properties. However, not all transcription factors have evolved to affect only one eukaryotic RNA polymerase. In fact, many factors have been shown to influence the activities of multiple nuclear RNA polymerases. This review focuses on a subset of these factors, specifically addressing the mechanisms by which these proteins influence RNA polymerases I and II.
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Affiliation(s)
- Yinfeng Zhang
- Department of Biochemistry and Molecular Genetics, University of Alabama at Birmingham, Birmingham, AL 35294
| | - Saman M Najmi
- Department of Biochemistry and Molecular Genetics, University of Alabama at Birmingham, Birmingham, AL 35294
| | - David A Schneider
- Department of Biochemistry and Molecular Genetics, University of Alabama at Birmingham, Birmingham, AL 35294
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20
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Abstract
The 137 ribosomal protein genes (RPG) of Saccharomyces provide a model for gene coregulation. Reja et al. examine the positional and functional organization of their regulators (Rap1, Fhl1, Ifh1, Sfp1, and Hmo1), the transcription machinery (TFIIB, TFIID, and RNA polymerase II), and chromatin at near-base-pair resolution using ChIP-exo. The 137 ribosomal protein genes (RPGs) of Saccharomyces provide a model for gene coregulation. We examined the positional and functional organization of their regulators (Rap1 [repressor activator protein 1], Fhl1, Ifh1, Sfp1, and Hmo1), the transcription machinery (TFIIB, TFIID, and RNA polymerase II), and chromatin at near-base-pair resolution using ChIP-exo, as RPGs are coordinately reprogrammed. Where Hmo1 is enriched, Fhl1, Ifh1, Sfp1, and Hmo1 cross-linked broadly to promoter DNA in an RPG-specific manner and demarcated by general minor groove widening. Importantly, Hmo1 extended 20–50 base pairs (bp) downstream from Fhl1. Upon RPG repression, Fhl1 remained in place. Hmo1 dissociated, which was coupled to an upstream shift of the +1 nucleosome, as reflected by the Hmo1 extension and core promoter region. Fhl1 and Hmo1 may create two regulatable and positionally distinct barriers, against which chromatin remodelers position the +1 nucleosome into either an activating or a repressive state. Consistent with in vitro studies, we found that specific TFIID subunits, in addition to cross-linking at the core promoter, made precise cross-links at Rap1 sites, which we interpret to reflect native Rap1–TFIID interactions. Our findings suggest how sequence-specific DNA binding regulates nucleosome positioning and transcription complex assembly >300 bp away and how coregulation coevolved with coding sequences.
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Affiliation(s)
- Rohit Reja
- Center for Eukaryotic Gene Regulation, Pennsylvania State University, University Park, Pennsylvania 16802, USA
| | - Vinesh Vinayachandran
- Center for Eukaryotic Gene Regulation, Pennsylvania State University, University Park, Pennsylvania 16802, USA
| | - Sujana Ghosh
- Center for Eukaryotic Gene Regulation, Pennsylvania State University, University Park, Pennsylvania 16802, USA
| | - B Franklin Pugh
- Center for Eukaryotic Gene Regulation, Pennsylvania State University, University Park, Pennsylvania 16802, USA
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21
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Watanabe K, Yabe M, Kasahara K, Kokubo T. A Random Screen Using a Novel Reporter Assay System Reveals a Set of Sequences That Are Preferred as the TATA or TATA-Like Elements in the CYC1 Promoter of Saccharomyces cerevisiae. PLoS One 2015; 10:e0129357. [PMID: 26046838 PMCID: PMC4457894 DOI: 10.1371/journal.pone.0129357] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/08/2015] [Accepted: 05/08/2015] [Indexed: 12/11/2022] Open
Abstract
In Saccharomyces cerevisiae, the core promoters of class II genes contain either TATA or TATA-like elements to direct accurate transcriptional initiation. Genome-wide analyses show that the consensus sequence of the TATA element is TATAWAWR (8 bp), whereas TATA-like elements carry one or two mismatches to this consensus. The fact that several functionally distinct TATA sequences have been identified indicates that these elements may function, at least to some extent, in a gene-specific manner. The purpose of the present study was to identify functional TATA sequences enriched in one particular core promoter and compare them with the TATA or TATA-like elements that serve as the pre-initiation complex (PIC) assembly sites on the yeast genome. For this purpose, we conducted a randomized screen of the TATA element in the CYC1 promoter by using a novel reporter assay system and identified several hundreds of unique sequences that were tentatively classified into nine groups. The results indicated that the 7 bp TATA element (i.e., TATAWAD) and several sets of TATA-like sequences are preferred specifically by this promoter. Furthermore, we find that the most frequently isolated TATA-like sequence, i.e., TATTTAAA, is actually utilized as a functional core promoter element for the endogenous genes, e.g., ADE5,7 and ADE6. Collectively, these results indicate that the sequence requirements for the functional TATA or TATA-like elements in one particular core promoter are not as stringent. However, the variation of these sequences differs significantly from that of the PIC assembly sites on the genome, presumably depending on promoter structures and reflecting the gene-specific function of these sequences.
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Affiliation(s)
- Kiyoshi Watanabe
- Molecular and Cellular Biology Laboratory, Graduate School of Medical Life Science, Yokohama City University, Yokohama, Kanagawa, Japan
| | - Makoto Yabe
- Molecular and Cellular Biology Laboratory, Graduate School of Medical Life Science, Yokohama City University, Yokohama, Kanagawa, Japan
| | - Koji Kasahara
- Molecular and Cellular Biology Laboratory, Graduate School of Medical Life Science, Yokohama City University, Yokohama, Kanagawa, Japan
| | - Tetsuro Kokubo
- Molecular and Cellular Biology Laboratory, Graduate School of Medical Life Science, Yokohama City University, Yokohama, Kanagawa, Japan
- * E-mail:
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22
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Higashino A, Shiwa Y, Yoshikawa H, Kokubo T, Kasahara K. Both HMG boxes in Hmo1 are essential for DNA binding in vitro and in vivo. Biosci Biotechnol Biochem 2014; 79:384-93. [PMID: 25410521 DOI: 10.1080/09168451.2014.978258] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/24/2022]
Abstract
Hmo1, a member of the high mobility group B family proteins in Saccharomyces cerevisiae, associates with the promoters of ribosomal protein genes (RPGs) to direct accurate transcriptional initiation. Here, to identify factors involved in the binding of Hmo1 to its targets and the mechanism of Hmo1-dependent transcriptional initiation, we developed a novel reporter system using the promoter of the RPG RPS5. A genetic screen did not identify any factors that influence Hmo1 binding, but did identify a number of mutations in Hmo1 that impair its DNA binding activity in vivo and in vitro. These results suggest that Hmo1 binds to its target promoters autonomously without any aid of additional factors. Furthermore, characterization of Hmo1 mutants showed that the box A domain plays a pivotal role in DNA binding and may be required for the recognition of structural properties of target promoters that occur in native chromatin.
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Affiliation(s)
- Ayako Higashino
- a Department of Bioscience , Tokyo University of Agriculture , Tokyo , Japan
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23
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Albert B, Colleran C, Léger-Silvestre I, Berger AB, Dez C, Normand C, Perez-Fernandez J, McStay B, Gadal O. Structure-function analysis of Hmo1 unveils an ancestral organization of HMG-Box factors involved in ribosomal DNA transcription from yeast to human. Nucleic Acids Res 2013; 41:10135-49. [PMID: 24021628 PMCID: PMC3905846 DOI: 10.1093/nar/gkt770] [Citation(s) in RCA: 40] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/12/2022] Open
Abstract
Ribosome biogenesis is a major metabolic effort for growing cells. In Saccharomyces cerevisiae, Hmo1, an abundant high-mobility group box protein (HMGB) binds to the coding region of the RNA polymerase I transcribed ribosomal RNAs genes and the promoters of ∼70% of ribosomal protein genes. In this study, we have demonstrated the functional conservation of eukaryotic HMGB proteins involved in ribosomal DNA (rDNA) transcription. We have shown that when expressed in budding yeast, human UBF1 and a newly identified Sp-Hmo1 (Schizosaccharomyces pombe) localize to the nucleolus and suppress growth defect of the RNA polymerase I mutant rpa49-Δ. Owing to the multiple functions of both proteins, Hmo1 and UBF1 are not fully interchangeable. By deletion and domains swapping in Hmo1, we identified essential domains that stimulate rDNA transcription but are not fully required for stimulation of ribosomal protein genes expression. Hmo1 is organized in four functional domains: a dimerization module, a canonical HMGB motif followed by a conserved domain and a C-terminal nucleolar localization signal. We propose that Hmo1 has acquired species-specific functions and shares with UBF1 and Sp-Hmo1 an ancestral function to stimulate rDNA transcription.
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Affiliation(s)
- Benjamin Albert
- LBME du CNRS, Université de Toulouse, 118 route de Narbonne, F-31000 Toulouse, France, Laboratoire de Biologie Moléculaire Eucaryote, Université de Toulouse, 118 route de Narbonne, F-31000 Toulouse, France and Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, University Road, Galway, Ireland
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Eguchi K, Yoshioka Y, Yoshida H, Morishita K, Miyata S, Hiai H, Yamaguchi M. The Drosophila DOCK family protein sponge is involved in differentiation of R7 photoreceptor cells. Exp Cell Res 2013; 319:2179-95. [DOI: 10.1016/j.yexcr.2013.05.024] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/24/2013] [Revised: 05/16/2013] [Accepted: 05/18/2013] [Indexed: 01/17/2023]
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The fate of the messenger is pre-determined: a new model for regulation of gene expression. BIOCHIMICA ET BIOPHYSICA ACTA-GENE REGULATORY MECHANISMS 2013; 1829:643-53. [PMID: 23337853 DOI: 10.1016/j.bbagrm.2013.01.004] [Citation(s) in RCA: 85] [Impact Index Per Article: 7.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 10/22/2012] [Revised: 01/07/2013] [Accepted: 01/08/2013] [Indexed: 02/08/2023]
Abstract
Recent years have seen a rise in publications demonstrating coupling between transcription and mRNA decay. This coupling most often accompanies cellular processes that involve transitions in gene expression patterns, for example during mitotic division and cellular differentiation and in response to cellular stress. Transcription can affect the mRNA fate by multiple mechanisms. The most novel finding is the process of co-transcriptional imprinting of mRNAs with proteins, which in turn regulate cytoplasmic mRNA stability. Transcription therefore is not only a catalyst of mRNA synthesis but also provides a platform that enables imprinting, which coordinates between transcription and mRNA decay. Here we present an overview of the literature, which provides the evidence of coupling between transcription and decay, review the mechanisms and regulators by which the two processes are coupled, discuss why such coupling is beneficial and present a new model for regulation of gene expression. This article is part of a Special Issue entitled: RNA Decay mechanisms.
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Xiao L, Kamau E, Donze D, Grove A. Expression of yeast high mobility group protein HMO1 is regulated by TOR signaling. Gene 2011; 489:55-62. [DOI: 10.1016/j.gene.2011.08.017] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/26/2011] [Revised: 07/19/2011] [Accepted: 08/26/2011] [Indexed: 10/17/2022]
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27
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Xiao L, Grove A. Coordination of Ribosomal Protein and Ribosomal RNA Gene Expression in Response to TOR Signaling. Curr Genomics 2011; 10:198-205. [PMID: 19881913 PMCID: PMC2705853 DOI: 10.2174/138920209788185261] [Citation(s) in RCA: 55] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/02/2009] [Revised: 03/04/2009] [Accepted: 03/06/2009] [Indexed: 01/22/2023] Open
Abstract
Cells grow in response to nutrients or growth factors, whose presence is detected and communicated by elaborate signaling pathways. Protein kinases play crucial roles in processes such as cell cycle progression and gene expression, and misregulation of such pathways has been correlated with various diseased states. Signals intended to promote cell growth converge on ribosome biogenesis, as the ability to produce cellular proteins is intimately tied to cell growth. Part of the response to growth signals is therefore the coordinate expression of genes encoding ribosomal RNA (rRNA) and ribosomal proteins (RP). A key player in regulating cell growth is the Target of Rapamycin (TOR) kinase, one of the gatekeepers that prevent cell cycle progression from G1 to S under conditions of nutritional stress. TOR is structurally and functionally conserved in all eukaryotes. Under favorable growth conditions, TOR is active and cells maintain a robust rate of ribosome biogenesis, translation initiation and nutrient import. Under stress conditions, TOR signaling is suppressed, leading to cell cycle arrest, while the failure of TOR to respond appropriately to environmental or nutritional signals leads to uncontrolled cell growth. Emerging evidence from Saccharomyces cerevisiae indicates that High Mobility Group (HMGB) proteins, non-sequence-specific chromosomal proteins, participate in mediating responses to growth signals. As HMGB proteins are distinguished by their ability to alter DNA topology, they frequently function in the assembly of higher-order nucleoprotein complexes. We review here recent evidence, which suggests that HMGB proteins may function to coordinate TOR-dependent regulation of rRNA and RP gene expression.
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Affiliation(s)
- Lijuan Xiao
- Department of Biological Sciences, Louisiana State University, Baton Rouge, LA 70803, USA
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Kasahara K, Ohyama Y, Kokubo T. Hmo1 directs pre-initiation complex assembly to an appropriate site on its target gene promoters by masking a nucleosome-free region. Nucleic Acids Res 2011; 39:4136-50. [PMID: 21288884 PMCID: PMC3105432 DOI: 10.1093/nar/gkq1334] [Citation(s) in RCA: 23] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/14/2022] Open
Abstract
Saccharomyces cerevisiae Hmo1 binds to the promoters of ∼70% of ribosomal protein genes (RPGs) at high occupancy, but is observed at lower occupancy on the remaining RPG promoters. In Δhmo1 cells, the transcription start site (TSS) of the Hmo1-enriched RPS5 promoter shifted upstream, while the TSS of the Hmo1-limited RPL10 promoter did not shift. Analyses of chimeric RPS5/RPL10 promoters revealed a region between the RPS5 upstream activating sequence (UAS) and core promoter, termed the intervening region (IVR), responsible for strong Hmo1 binding and an upstream TSS shift in Δhmo1 cells. Chromatin immunoprecipitation analyses showed that the RPS5-IVR resides within a nucleosome-free region and that pre-initiation complex (PIC) assembly occurs at a site between the IVR and a nucleosome overlapping the TSS (+1 nucleosome). The PIC assembly site was shifted upstream in Δhmo1 cells on this promoter, indicating that Hmo1 normally masks the RPS5-IVR to prevent PIC assembly at inappropriate site(s). This novel mechanism ensures accurate transcriptional initiation by delineating the 5′- and 3′-boundaries of the PIC assembly zone.
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Affiliation(s)
- Koji Kasahara
- Division of Molecular and Cellular Biology, Graduate School of Nanobioscience, Yokohama City University, Yokohama 230-0045, Japan.
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Sugihara F, Kasahara K, Kokubo T. Highly redundant function of multiple AT-rich sequences as core promoter elements in the TATA-less RPS5 promoter of Saccharomyces cerevisiae. Nucleic Acids Res 2010; 39:59-75. [PMID: 20805245 PMCID: PMC3017598 DOI: 10.1093/nar/gkq741] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/26/2022] Open
Abstract
In eukaryotes, protein-coding genes are transcribed by RNA polymerase II (pol II) together with general transcription factors (GTFs). TFIID, the largest GTF composed of TATA element-binding protein (TBP) and 14 TBP-associated factors (TAFs), plays a critical role in transcription from TATA-less promoters. In metazoans, several core promoter elements other than the TATA element are thought to be recognition sites for TFIID. However, it is unclear whether functionally homologous elements also exist in TATA-less promoters in Saccharomyces cerevisiae. Here, we identify the cis-elements required to support normal levels of transcription and accurate initiation from sites within the TATA-less and TFIID-dependent RPS5 core promoter. Systematic mutational analyses show that multiple AT-rich sequences are required for these activities and appear to function as recognition sites for TFIID. A single copy of these sequences can support accurate initiation from the endogenous promoter, indicating that they carry highly redundant functions. These results show a novel architecture of yeast TATA-less promoters and support a model in which pol II scans DNA downstream from a recruited site, while searching for appropriate initiation site(s).
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Affiliation(s)
- Fuminori Sugihara
- Division of Molecular and Cellular Biology, Graduate School of Nanobioscience, Yokohama City University, Yokohama, Kanagawa, Japan
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Xiao L, Williams AM, Grove A. The C-terminal domain of yeast high mobility group protein HMO1 mediates lateral protein accretion and in-phase DNA bending. Biochemistry 2010; 49:4051-9. [PMID: 20402481 DOI: 10.1021/bi1003603] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2022]
Abstract
The Saccharomyces cerevisiae high mobility group protein HMO1 has two DNA binding domains, box A and box B, and a lysine-rich C-terminal extension. Among other functions, HMO1 has been implicated as a component of the RNA polymerase I transcription machinery. We report here that HMO1 promotes DNA apposition as evidenced by its stimulation of end-joining in the presence of T4 DNA ligase. Analysis of truncated HMO1 variants shows that enhanced DNA end-joining requires the C-terminal domain but that box A is dispensable. The efficiency of joining DNA ends with different nucleotide content parallels that of DNA ligase, and optimal ligation efficiency is attained when DNA is effectively saturated with protein, implying that HMO1 binds internal sites in preference to DNA ends. Removal of the C-terminal tail does not attenuate the self-association characteristic of HMO1 but alters the stoichiometry of binding and prevents intramolecular DNA cyclization. This suggests that the C-terminal domain mediates an accretion of HMO1 on DNA that causes in-phase DNA bending and that binding of HMO1 lacking the C-terminal domain results in out-of-phase bending. Taken together, our results show that HMO1 shares with mammalian HMGB proteins the ability to promote DNA association. Notably, the C-terminal domain mediates both DNA end-joining and an accretion of multiple HMO1 protomers on duplex DNA that produces in-phase DNA bending. This mode of binding is reminiscent of that proposed for the mammalian RNA polymerase I transcription factor UBF.
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Affiliation(s)
- Lijuan Xiao
- Department of Biological Sciences, Louisiana State University, Baton Rouge, Louisiana 70803, USA
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Ohtsuki K, Kasahara K, Shirahige K, Kokubo T. Genome-wide localization analysis of a complete set of Tafs reveals a specific effect of the taf1 mutation on Taf2 occupancy and provides indirect evidence for different TFIID conformations at different promoters. Nucleic Acids Res 2009; 38:1805-20. [PMID: 20026583 PMCID: PMC2847235 DOI: 10.1093/nar/gkp1172] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/14/2022] Open
Abstract
In Saccharomyces cerevisiae, TFIID and SAGA principally mediate transcription of constitutive housekeeping genes and stress-inducible genes, respectively, by delivering TBP to the core promoter. Both are multi-protein complexes composed of 15 and 20 subunits, respectively, five of which are common and which may constitute a core sub-module in each complex. Although genome-wide gene expression studies have been conducted extensively in several TFIID and/or SAGA mutants, there are only a limited number of studies investigating genome-wide localization of the components of these two complexes. Specifically, there are no previous reports on localization of a complete set of Tafs and the effects of taf mutations on localization. Here, we examine the localization profiles of a complete set of Tafs, Gcn5, Bur6/Ncb2, Sua7, Tfa2, Tfg1, Tfb3 and Rpb1, on chromosomes III, IV and V by chromatin immunoprecipitation (ChIP)-chip analysis in wild-type and taf1-T657K mutant strains. In addition, we conducted conventional and sequential ChIP analysis of several ribosomal protein genes (RPGs) and non-RPGs. Intriguingly, the results revealed a novel relationship between TFIIB and NC2, simultaneous co-localization of SAGA and TFIID on RPG promoters, specific effects of taf1 mutation on Taf2 occupancy, and an indirect evidence for the existence of different TFIID conformations.
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Affiliation(s)
- Kazushige Ohtsuki
- Division of Molecular and Cellular Biology, Graduate School of Nanobioscience, Yokohama City University, Yokohama 230-0045, Japan
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Current awareness on yeast. Yeast 2008. [DOI: 10.1002/yea.1558] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/05/2022] Open
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Yeast Rap1 contributes to genomic integrity by activating DNA damage repair genes. EMBO J 2008; 27:1575-84. [PMID: 18480842 DOI: 10.1038/emboj.2008.93] [Citation(s) in RCA: 27] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/10/2007] [Accepted: 04/14/2008] [Indexed: 11/08/2022] Open
Abstract
Rap1 (repressor-activator protein 1) is a multifunctional protein that controls telomere function, silencing and the activation of glycolytic and ribosomal protein genes. We have identified a novel function for Rap1, regulating the ribonucleotide reductase (RNR) genes that are required for DNA repair and telomere expansion. Both the C terminus and DNA-binding domain of Rap1 are required for the activation of the RNR genes, and the phenotypes of different Rap1 mutants suggest that it utilizes both regions to carry out distinct steps in the activation process. Recruitment of Rap1 to the RNR3 gene is dependent on activation of the DNA damage checkpoint and chromatin remodelling by SWI/SNF. The dependence on SWI/SNF for binding suggests that Rap1 acts after remodelling to prevent the repositioning of nucleosomes back to the repressed state. Furthermore, the recruitment of Rap1 requires TAF(II)s, suggesting a role for TFIID in stabilizing activator binding in vivo. We propose that Rap1 acts as a rheostat controlling nucleotide pools in response to shortened telomeres and DNA damage, providing a mechanism for fine-tuning the RNR genes during checkpoint activation.
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