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Xi BX, Liu SY, Xu YT, Zhang DD, Hu Q, Liu AG. Genetic Analysis of Two Novel GPI Variants Disrupting H Bonds and Localization Characteristics of 55 Gene Variants Associated with Glucose-6-phosphate Isomerase Deficiency. Curr Med Sci 2024; 44:426-434. [PMID: 38561594 DOI: 10.1007/s11596-024-2857-3] [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: 12/08/2023] [Accepted: 02/04/2024] [Indexed: 04/04/2024]
Abstract
OBJECTIVE Glucose-6-phosphate isomerase (GPI) deficiency is a rare hereditary nonspherocytic hemolytic anemia caused by GPI gene variants. This disorder exhibits wide heterogeneity in its clinical manifestations and molecular characteristics, often posing challenges for precise diagnoses using conventional methods. To this end, this study aimed to identify the novel variants responsible for GPI deficiency in a Chinese family. METHODS The clinical manifestations of the patient were summarized and analyzed for GPI deficiency phenotype diagnosis. Novel compound heterozygous variants of the GPI gene, c.174C>A (p.Asn58Lys) and c.1538G>T (p.Trp513Leu), were identified using whole-exome and Sanger sequencing. The AlphaFold program and Chimera software were used to analyze the effects of compound heterozygous variants on GPI structure. RESULTS By characterizing 53 GPI missense/nonsense variants from previous literature and two novel missense variants identified in this study, we found that most variants were located in exons 3, 4, 12, and 18, with a few localized in exons 8, 9, and 14. This study identified novel compound heterozygous variants associated with GPI deficiency. These pathogenic variants disrupt hydrogen bonds formed by highly conserved GPI amino acids. CONCLUSION Early family-based sequencing analyses, especially for patients with congenital anemia, can help increase diagnostic accuracy for GPI deficiency, improve child healthcare, and enable genetic counseling.
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Affiliation(s)
- Bi-Xin Xi
- Department of Pediatrics, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430030, China
| | - Si-Ying Liu
- Department of Pediatrics, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430030, China
| | - Yu-Ting Xu
- Department of Pediatrics, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430030, China
| | - De-Dong Zhang
- Department of Pediatrics, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430030, China
| | - Qun Hu
- Department of Pediatrics, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430030, China
| | - Ai-Guo Liu
- Department of Pediatrics, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430030, China.
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Zhang Y, Wang X, Chen G, Lu Y, Chen Q. Autocrine motility factor receptor promotes the malignancy of glioblastoma by regulating cell migration and invasion. Neurol Res 2024; 46:89-97. [PMID: 37703903 DOI: 10.1080/01616412.2023.2257463] [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: 05/04/2023] [Accepted: 07/30/2023] [Indexed: 09/15/2023]
Abstract
OBJECTIVE One of the important causes of death in cancer patients is malignant metastasis, invasion, and metastasis of tumor cells. Metastasis is also the most basic physiological characteristics and pathogenesis of various tumors. Previously published studies have suggested that autocrine motor factor receptor (AMFR) is the key regulator of tumor cell migration and invasion. Meanwhile, AMFR is highly expressed in esophageal tumors, gastrointestinal tumors, and bladder cancer, and it is also involved in its pathogenesis. However, the role of AMFR in glioblastoma has not been reported. METHODS In order to study the role of AMFR in the cell migration and invasion of glioblastoma, AMFR was silenced using siRNA and overexpressed using cDNA. Immunoblotting analysis and real-time quantitative polymerase chain reaction (PCR) were employed to assess the expression of AMFR. We conducted wound healing assay, cell migration assay, and tumorsphere formation assay to detect the invasion and metastatic ability of glioblastoma. RESULTS This study found that the level of AMFR expression was significantly correlated with the malignant degree of glioma tissue in clinic samples. AMFR silencing decreased cell migration and invasion of LN229. Overexpression of AMFR significantly increased cell migration and invasion of U251. CONCLUSION This study suggests that AMFR could be used as a therapeutic strategy for the clinical treatment of glioblastoma.
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Affiliation(s)
- Yao Zhang
- Department of Endocrinology, Liyuan Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
| | - Xiuping Wang
- Department of Pharmacy, Liyuan Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
| | - Guanghui Chen
- Department of Pharmacy, Renmin Hospital, Wuhan University, Wuhan, China
| | - Yajing Lu
- Institute of geriatric medicine, Liyuan Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
| | - Qiang Chen
- Department of Pharmacy, Liyuan Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
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3
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Zhang Y, Liu Z, Li X, Liu L, Wang L, Han X, Li Z. Comprehensive Molecular Analyses of a Six-Gene Signature for Predicting Late Recurrence of Hepatocellular Carcinoma. Front Oncol 2021; 11:732447. [PMID: 34568069 PMCID: PMC8459683 DOI: 10.3389/fonc.2021.732447] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/29/2021] [Accepted: 08/16/2021] [Indexed: 01/12/2023] Open
Abstract
A larger number of patients with stages I-III hepatocellular carcinoma (HCC) experience late recurrence (LR) after surgery. We sought to develop a novel tool to stratify patients with different LR risk for tailoring decision-making for postoperative recurrence surveillance and therapy modalities. We retrospectively enrolled two independent public cohorts and 103 HCC tissues. Using LASSO logical analysis, a six-gene model was developed in the The Cancer Genome Atlas liver hepatocellular carcinoma (TCGA-LIHC) and independently validated in GSE76427. Further experimental validation using qRT-PCR assays was performed to ensure the robustness and clinical feasible of this signature. We developed a novel LR-related signature consisting of six genes. This signature was validated to be significantly associated with dismal recurrence-free survival in three cohorts TCGA-LIHC, GSE76427, and qPCR assays [HR: 2.007 (1.200-3.357), p = 0.008; HR: 2.171 (1.068, 4.412), p-value = 0.032; HR: 3.383 (2.100, 5.450), p-value <0.001]. More importantly, this signature displayed robust discrimination in predicting the LR risk, with AUCs being 0.73 (TCGA-LIHC), 0.93 (GSE76427), and 0.85 (in-house cohort). Furthermore, we deciphered the specific landscape of molecular alterations among patients in nonrecurrence (NR) and LR group to analyze the mechanism contributing to LR. For high-risk group, we also identified several potential drugs with specific sensitivity to high- and low-risk groups, which is vital to improve prognosis of LR-HCC after surgery. We discovered and experimentally validated a novel gene signature with powerful performance for identifying patients at high LR risk in stages I-III HCC.
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Affiliation(s)
- Yuyuan Zhang
- Department of Interventional Radiology, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, China.,Interventional Institute of Zhengzhou University, Zhengzhou, China.,Interventional Treatment and Clinical Research Center of Henan Province, Zhengzhou, China
| | - Zaoqu Liu
- Department of Interventional Radiology, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, China.,Interventional Institute of Zhengzhou University, Zhengzhou, China.,Interventional Treatment and Clinical Research Center of Henan Province, Zhengzhou, China
| | - Xin Li
- Department of Interventional Radiology, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, China.,Interventional Institute of Zhengzhou University, Zhengzhou, China.,Interventional Treatment and Clinical Research Center of Henan Province, Zhengzhou, China
| | - Long Liu
- Department of Hepatobiliary and Pancreatic Surgery, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, China
| | - Libo Wang
- Department of Hepatobiliary and Pancreatic Surgery, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, China
| | - Xinwei Han
- Department of Interventional Radiology, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, China.,Interventional Institute of Zhengzhou University, Zhengzhou, China.,Interventional Treatment and Clinical Research Center of Henan Province, Zhengzhou, China
| | - Zhen Li
- Department of Interventional Radiology, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, China.,Interventional Institute of Zhengzhou University, Zhengzhou, China.,Interventional Treatment and Clinical Research Center of Henan Province, Zhengzhou, China
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The chemokine CCL1 triggers an AMFR-SPRY1 pathway that promotes differentiation of lung fibroblasts into myofibroblasts and drives pulmonary fibrosis. Immunity 2021; 54:2042-2056.e8. [PMID: 34407391 DOI: 10.1016/j.immuni.2021.06.008] [Citation(s) in RCA: 54] [Impact Index Per Article: 13.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/28/2020] [Revised: 02/20/2021] [Accepted: 06/11/2021] [Indexed: 12/28/2022]
Abstract
Recruitment of immune cells to the site of inflammation by the chemokine CCL1 is important in the pathology of inflammatory diseases. Here, we examined the role of CCL1 in pulmonary fibrosis (PF). Bronchoalveolar lavage fluid from PF mouse models contained high amounts of CCL1, as did lung biopsies from PF patients. Immunofluorescence analyses revealed that alveolar macrophages and CD4+ T cells were major producers of CCL1 and targeted deletion of Ccl1 in these cells blunted pathology. Deletion of the CCL1 receptor Ccr8 in fibroblasts limited migration, but not activation, in response to CCL1. Mass spectrometry analyses of CCL1 complexes identified AMFR as a CCL1 receptor, and deletion of Amfr impaired fibroblast activation. Mechanistically, CCL1 binding triggered ubiquitination of the ERK inhibitor Spry1 by AMFR, thus activating Ras-mediated profibrotic protein synthesis. Antibody blockade of CCL1 ameliorated PF pathology, supporting the therapeutic potential of targeting this pathway for treating fibroproliferative lung diseases.
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5
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Autocrine motility factor and its receptor expression in musculoskeletal tumors. J Bone Oncol 2020; 24:100318. [PMID: 33101887 PMCID: PMC7574284 DOI: 10.1016/j.jbo.2020.100318] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/19/2020] [Revised: 09/03/2020] [Accepted: 09/07/2020] [Indexed: 11/21/2022] Open
Abstract
Discovery of Autocrine Motility factor (AMF) and its receptor (AMFR), both triggering tumor invasion and metastasis, may alter the therapeutic concept. Here, in this review, we show a novel outlook suggesting a cross-talking between musculoskeletal tumors and the skeletal milieu regulated by AMF-AMFR signaling. This review will highlight the pharmacological need for AMF and AMFR inhibitors for patients with malignant musculoskeletal tumors. Management of aggressive malignant musculoskeletal tumors is clinically challenging and awaits the identification of regulator(s) that can be therapeutically used to improve patient outcome. Autocrine motility factor (AMF), a secreted cytokine, is known to alter the bone microenvironment by linking to its receptor AMFR (AMF Receptor), leading to tumor progression. It was noted that both the ligand and its receptor belong to the moonlighting family of proteins, as they contribute to intracellular metabolic function such as glycolysis and gluconeogenesis by expressing glucose-6-phosphate isomerase AMF/GPI and higher protein degradation by expressing AMFR/gp78 functioning as ubiquitin ligase activity. Thus, AMF/GPI and AMFR/gp78 contribute to higher metabolic turnover of protein and glucose. Recently, a large-scale cohort study including 23 different histological types of musculoskeletal tumors revealed that patients with osteosarcoma, multiple myeloma, rhabdomyosarcoma, and angiosarcoma tend to express higher levels of AMF, whereas multiple myeloma patients expressed high levels of AMFR. Consistently, the cellular data showed that a variety of musculoskeletal tumors express AMF and components of bone microenvironment express AMFR. Thus, a novel outlook suggests a cellular link and cross-talk between musculoskeletal tumors and the skeletal milieu are regulated by AMF-AMFR signaling. This review will highlight the pharmacological need for AMF and AMFR inhibitors as unmet medical needs for patients with malignant musculoskeletal tumors.
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Nakajima K, Raz A. Amplification of autocrine motility factor and its receptor in multiple myeloma and other musculoskeletal tumors. J Bone Oncol 2020; 23:100308. [PMID: 32714781 PMCID: PMC7378681 DOI: 10.1016/j.jbo.2020.100308] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/14/2020] [Revised: 07/01/2020] [Accepted: 07/02/2020] [Indexed: 11/30/2022] Open
Abstract
This study is a large scale cohort of the patients with malignant musculoskeletal tumors to determine the expression levels of Autocrine Motility factor (AMF) and its receptor (AMFR). We Visualization of amplified Autocrine motility factor (AMF) and its receptor (AMFR) in musculoskeletal tumors. A novel software aimed at analyzing numerous cell-to-cell and ligand-to-receptor interactions was developed, which lead to visualization of bone tumor microenvironment. Autocrine motility factor (AMF: GPI) and its receptor AMFR (AMF Receptor: gp78) regulate the metastatic process. Here, we have tested the expression levels of AMF, AMFR, and AMF × AMFR in 1348 patients with musculoskeletal tumor. The results depicted here identified that multiple myeloma highly express AMF × AMFR value as compared with normal bone samples (p < 0.00001). To visualize the AMF × AMFR autocrine amplification in multiple myeloma microenvironment, we have developed a novel software aimed at analyzing numerous cell-to-cell and ligand-to-receptor interactions, i.e., Environmentome. It has led to the identification that myeloma-associated interactions with normal bone cells including osteoblast, osteoclast, immunological components, and others in a paracrine manner. In conclusion, the data showed that AMF × AMFR amplification is a clinical manifestation in bone microenvironment of multiple myeloma.
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Affiliation(s)
- Kosei Nakajima
- Division of Translational Research, Exploratory Oncology Research & Clinical Trial Center, National Cancer Center Research Institute. 5-1-1 Tsukiji, Chuo-Ku, Tokyo 104-0045, Japan.,Division of Veterinary Oncology and Surgery, Faculty of Veterinary Medicine, Imabari Campus, Okayama University of Science, 1-3 Ikoinooka, Imabari, Ehime 794-8555, Japan
| | - Avraham Raz
- Departments of Oncology and Pathology, School of Medicine, Wayne State University and Barbara Ann Karmanos Cancer Institute, 4100 John R St, Detroit, MI 48201, United States
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7
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Rosa N, Sneyers F, Parys JB, Bultynck G. Type 3 IP 3 receptors: The chameleon in cancer. INTERNATIONAL REVIEW OF CELL AND MOLECULAR BIOLOGY 2020; 351:101-148. [PMID: 32247578 DOI: 10.1016/bs.ircmb.2020.02.003] [Citation(s) in RCA: 17] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
Inositol 1,4,5-trisphosphate (IP3) receptors (IP3Rs), intracellular calcium (Ca2+) release channels, fulfill key functions in cell death and survival processes, whose dysregulation contributes to oncogenesis. This is essentially due to the presence of IP3Rs in microdomains of the endoplasmic reticulum (ER) in close proximity to the mitochondria. As such, IP3Rs enable efficient Ca2+ transfers from the ER to the mitochondria, thus regulating metabolism and cell fate. This review focuses on one of the three IP3R isoforms, the type 3 IP3R (IP3R3), which is linked to proapoptotic ER-mitochondrial Ca2+ transfers. Alterations in IP3R3 expression have been highlighted in numerous cancer types, leading to dysregulations of Ca2+ signaling and cellular functions. However, the outcome of IP3R3-mediated Ca2+ transfers for mitochondrial function is complex with opposing effects on oncogenesis. IP3R3 can either suppress cancer by promoting cell death and cellular senescence or support cancer by driving metabolism, anabolic processes, cell cycle progression, proliferation and invasion. The aim of this review is to provide an overview of IP3R3 dysregulations in cancer and describe how such dysregulations alter critical cellular processes such as proliferation or cell death and survival. Here, we pose that the IP3R3 isoform is not only linked to proapoptotic ER-mitochondrial Ca2+ transfers but might also be involved in prosurvival signaling.
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Affiliation(s)
- Nicolas Rosa
- KU Leuven, Laboratory of Molecular and Cellular Signaling, Department of Cellular and Molecular Medicine and Leuven Kanker Instituut (LKI), Leuven, Belgium
| | - Flore Sneyers
- KU Leuven, Laboratory of Molecular and Cellular Signaling, Department of Cellular and Molecular Medicine and Leuven Kanker Instituut (LKI), Leuven, Belgium
| | - Jan B Parys
- KU Leuven, Laboratory of Molecular and Cellular Signaling, Department of Cellular and Molecular Medicine and Leuven Kanker Instituut (LKI), Leuven, Belgium
| | - Geert Bultynck
- KU Leuven, Laboratory of Molecular and Cellular Signaling, Department of Cellular and Molecular Medicine and Leuven Kanker Instituut (LKI), Leuven, Belgium.
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8
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Energy Metabolism in Cancer: The Roles of STAT3 and STAT5 in the Regulation of Metabolism-Related Genes. Cancers (Basel) 2020; 12:cancers12010124. [PMID: 31947710 PMCID: PMC7016889 DOI: 10.3390/cancers12010124] [Citation(s) in RCA: 52] [Impact Index Per Article: 10.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/27/2019] [Revised: 12/03/2019] [Accepted: 12/12/2019] [Indexed: 12/21/2022] Open
Abstract
A central characteristic of many types of cancer is altered energy metabolism processes such as enhanced glucose uptake and glycolysis and decreased oxidative metabolism. The regulation of energy metabolism is an elaborate process involving regulatory proteins such as HIF (pro-metastatic protein), which reduces oxidative metabolism, and some other proteins such as tumour suppressors that promote oxidative phosphorylation. In recent years, it has been demonstrated that signal transducer and activator of transcription (STAT) proteins play a pivotal role in metabolism regulation. STAT3 and STAT5 are essential regulators of cytokine- or growth factor-induced cell survival and proliferation, as well as the crosstalk between STAT signalling and oxidative metabolism. Several reports suggest that the constitutive activation of STAT proteins promotes glycolysis through the transcriptional activation of hypoxia-inducible factors and therefore, the alteration of mitochondrial activity. It seems that STAT proteins function as an integrative centre for different growth and survival signals for energy and respiratory metabolism. This review summarises the functions of STAT3 and STAT5 in the regulation of some metabolism-related genes and the importance of oxygen in the tumour microenvironment to regulate cell metabolism, particularly in the metabolic pathways that are involved in energy production in cancer cells.
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9
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Escobar-Henriques M, Joaquim M. Mitofusins: Disease Gatekeepers and Hubs in Mitochondrial Quality Control by E3 Ligases. Front Physiol 2019; 10:517. [PMID: 31156446 PMCID: PMC6533591 DOI: 10.3389/fphys.2019.00517] [Citation(s) in RCA: 57] [Impact Index Per Article: 9.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/20/2019] [Accepted: 04/11/2019] [Indexed: 02/06/2023] Open
Abstract
Mitochondria are dynamic organelles engaged in quality control and aging processes. They constantly undergo fusion, fission, transport, and anchoring events, which empower mitochondria with a very interactive behavior. The membrane remodeling processes needed for fusion require conserved proteins named mitofusins, MFN1 and MFN2 in mammals and Fzo1 in yeast. They are the first determinants deciding on whether communication and content exchange between different mitochondrial populations should occur. Importantly, each cell possesses hundreds of mitochondria, with a different severity of mitochondrial mutations or dysfunctional proteins, which potentially spread damage to the entire network. Therefore, the degree of their merging capacity critically influences cellular fitness. In turn, the mitochondrial network rapidly and dramatically changes in response to metabolic and environmental cues. Notably, cancer or obesity conditions, and stress experienced by neurons and cardiomyocytes, for example, triggers the downregulation of mitofusins and thus fragmentation of mitochondria. This places mitofusins upfront in sensing and transmitting stress. In fact, mitofusins are almost entirely exposed to the cytoplasm, a topology suitable for a critical relay point in information exchange between mitochondria and their cellular environment. Consistent with their topology, mitofusins are either activated or repressed by cytosolic post-translational modifiers, mainly by ubiquitin. Ubiquitin is a ubiquitous small protein orchestrating multiple quality control pathways, which is covalently attached to lysine residues in its substrates, or in ubiquitin itself. Importantly, from a chain of events also mediated by E1 and E2 enzymes, E3 ligases perform the ultimate and determinant step in substrate choice. Here, we review the ubiquitin E3 ligases that modify mitofusins. Two mitochondrial E3 enzymes—March5 and MUL1—one ligase located to the ER—Gp78—and finally three cytosolic enzymes—MGRN1, HUWE1, and Parkin—were shown to ubiquitylate mitofusins, in response to a variety of cellular inputs. The respective outcomes on mitochondrial morphology, on contact sites to the endoplasmic reticulum and on destructive processes, like mitophagy or apoptosis, are presented. Ultimately, understanding the mechanisms by which E3 ligases and mitofusins sense and bi-directionally signal mitochondria-cytosolic dysfunctions could pave the way for therapeutic approaches in neurodegenerative, cardiovascular, and obesity-linked diseases.
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Affiliation(s)
- Mafalda Escobar-Henriques
- Center for Molecular Medicine Cologne (CMMC), Institute for Genetics, Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases (CECAD), University of Cologne, Cologne, Germany
| | - Mariana Joaquim
- Center for Molecular Medicine Cologne (CMMC), Institute for Genetics, Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases (CECAD), University of Cologne, Cologne, Germany
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Joshi V, Upadhyay A, Chhangani D, Amanullah A, Sharan RN, Mishra A. Gp78 involvement in cellular proliferation: Can act as a promising modulator for cell cycle regulatory proteins? J Cell Physiol 2018; 233:6352-6368. [PMID: 29741771 DOI: 10.1002/jcp.26618] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/30/2017] [Accepted: 03/28/2018] [Indexed: 11/07/2022]
Abstract
In cells, protein synthesis and degradation are normal processes, which are tightly regulated by various cellular metabolic pathways. Cellular protein quality control (PQC) mechanisms always present a continuous and rigorous check over all intracellular proteins before they can participate in various cellular physiological processes with the help of PQC pathways like autophagy and ubiquitin proteasome system (UPS). The UPS employs few selective E3 ubiquitin ligases for the intracellular degradation of cyclin-dependent kinase inhibitor 1B (p27Kip1 ) that tightly controls cell cycle progression. But, the complex mechanistic interactions and the interplay between E3 ubiquitin ligases involved in the functional regulation as well as expression of p27 are not well known. Here, we demonstrate that cell surface glycoprotein Gp78, a putative E3 ubiquitin ligase, is involved in the stabilization of intracellular steady-state levels of p27. Transient overexpression of Gp78 increases the accumulation of p27 in cells in the form of massive inclusions like structures, which could be due to its cumulative increased stability in cells. We have also monitored how under stress condition, E3 ubiquitin ligase Gp78 regulates endogenous levels of p27 in cells. ER stress treatment generates a marginal increase in Gp78 endogenous levels, and this elevation effect was prominent for intracellular accumulation of p27 in cells. Taken together, our current findings suggest a valuable multifactorial regulatory mechanism and linkage of p27 with UPS pathway.
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Affiliation(s)
- Vibhuti Joshi
- Cellular and Molecular Neurobiology Unit, Indian Institute of Technology Jodhpur, Rajasthan, India
| | - Arun Upadhyay
- Cellular and Molecular Neurobiology Unit, Indian Institute of Technology Jodhpur, Rajasthan, India
| | - Deepak Chhangani
- Cellular and Molecular Neurobiology Unit, Indian Institute of Technology Jodhpur, Rajasthan, India
| | - Ayeman Amanullah
- Cellular and Molecular Neurobiology Unit, Indian Institute of Technology Jodhpur, Rajasthan, India
| | - Rajesh N Sharan
- Radiation and Molecular Biology Unit, Department of Biochemistry, North-Eastern Hill University, Shillong, Meghalaya, India
| | - Amit Mishra
- Cellular and Molecular Neurobiology Unit, Indian Institute of Technology Jodhpur, Rajasthan, India
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11
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Joshi V, Upadhyay A, Kumar A, Mishra A. Gp78 E3 Ubiquitin Ligase: Essential Functions and Contributions in Proteostasis. Front Cell Neurosci 2017; 11:259. [PMID: 28890687 PMCID: PMC5575403 DOI: 10.3389/fncel.2017.00259] [Citation(s) in RCA: 37] [Impact Index Per Article: 4.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/04/2017] [Accepted: 08/09/2017] [Indexed: 11/26/2022] Open
Abstract
As per the requirement of metabolism and fitness, normal cellular functions are controlled by several proteins, and their interactive molecular and signaling events at multiple levels. Protein quality control (PQC) mechanisms ensure the correct folding and proper utilization of these proteins to avoid their misfolding and aggregation. To maintain the optimum environment of complex proteome PQC system employs various E3 ubiquitin ligases for the selective degradation of aberrant proteins. Glycoprotein 78 (Gp78) is an E3 ubiquitin ligase that prevents multifactorial deleterious accumulation of different misfolded proteins via endoplasmic reticulum-associated degradation (ERAD). However, the precise role of Gp78 under stress conditions to avoid bulk misfolded aggregation is unclear, which can act as a crucial resource to establish the dynamic nature of the proteome. Present article systematically explains the detailed molecular characterization of Gp78 and also addresses its various cellular physiological functions, which could be crucial to achieving protein homeostasis. Here, we comprehensively represent the current findings of Gp78, which shows its PQC roles in different physiological functions and diseases; and thereby propose novel opportunities to better understand the unsolved questions for therapeutic interventions linked with different protein misfolding disorders.
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Affiliation(s)
- Vibhuti Joshi
- Cellular and Molecular Neurobiology Unit, Indian Institute of Technology JodhpurJodhpur, India
| | - Arun Upadhyay
- Cellular and Molecular Neurobiology Unit, Indian Institute of Technology JodhpurJodhpur, India
| | - Amit Kumar
- Centre for Biosciences and Biomedical Engineering, Indian Institute of Technology IndoreIndore, India
| | - Amit Mishra
- Cellular and Molecular Neurobiology Unit, Indian Institute of Technology JodhpurJodhpur, India
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Stiles JK, Meade JC, Kucerova Z, Lyn D, Thompson W, Zakeri Z, Whittaker J. Trypanosoma bruceiinfection induces apoptosis and up-regulates neuroleukin expression in the cerebellum. ANNALS OF TROPICAL MEDICINE AND PARASITOLOGY 2016. [DOI: 10.1080/00034983.2001.11813699] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/28/2022]
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13
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Lincet H, Icard P. How do glycolytic enzymes favour cancer cell proliferation by nonmetabolic functions? Oncogene 2014; 34:3751-9. [PMID: 25263450 DOI: 10.1038/onc.2014.320] [Citation(s) in RCA: 140] [Impact Index Per Article: 12.7] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/27/2014] [Revised: 08/23/2014] [Accepted: 08/23/2014] [Indexed: 12/16/2022]
Abstract
Cancer cells enhance their glycolysis, producing lactate, even in the presence of oxygen. Glycolysis is a series of ten metabolic reactions catalysed by enzymes whose expression is most often increased in tumour cells. HKII and phosphoglucose isomerase (PGI) have mainly an antiapoptotic effect; PGI and glyceraldehyde-3-phosphate dehydrogenase activate survival pathways (Akt and so on); phosphofructokinase 1 and triose phosphate isomerase participate in cell cycle activation; aldolase promotes epithelial mesenchymal transition; PKM2 enhances various nuclear effects such as transcription, stabilisation and so on. This review outlines the multiple non-glycolytic roles of glycolytic enzymes, which are essential for promoting cancer cells' survival, proliferation, chemoresistance and dissemination.
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Affiliation(s)
- H Lincet
- 1] Locally Aggressive Cancer Biology and Therapy Unit (BioTICLA), Caen, France [2] Normandie University, Caen, France [3] François-Baclesse Centre for Cancer, Caen, France
| | - P Icard
- 1] Locally Aggressive Cancer Biology and Therapy Unit (BioTICLA), Caen, France [2] Ecole Polytechnique, Laboratoire d'Informatique, Palaiseau, France
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Marzook H, Li DQ, Nair VS, Mudvari P, Reddy SDN, Pakala SB, Santhoshkumar TR, Pillai MR, Kumar R. Metastasis-associated protein 1 drives tumor cell migration and invasion through transcriptional repression of RING finger protein 144A. J Biol Chem 2012; 287:5615-26. [PMID: 22184113 PMCID: PMC3285335 DOI: 10.1074/jbc.m111.314088] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/14/2011] [Revised: 12/16/2011] [Indexed: 01/01/2023] Open
Abstract
Metastasis-associated protein 1 (MTA1), a component of the nucleosome-remodeling and histone deacetylase complex, is widely up-regulated in human cancers and significantly correlated with tumor invasion and metastasis, but the mechanisms involved remain largely unknown. Here, we report that MTA1 transcriptionally represses the expression of RING finger protein 144A (RNF144A), an uncharacterized gene whose protein product possesses potential E3 ubiquitin ligase activity, by recruiting the histone deacetylase 2 (HDAC2) and CCAAT/enhancer-binding protein α (c/EBPα) co-repressor complex onto human RNF144A promoter. Furthermore, an inverse correlation between the expression levels of MTA1 and RNF144A was demonstrated in publicly available breast cancer microarray datasets and the MCF10 breast cancer progression model system. To address functional aspects of MTA1 regulation of RNF144A, we demonstrate that RNF144A is a novel suppressor of cancer migration and invasion, two requisite steps of metastasis in vivo, and knockdown of endogenous RNF144A by small interfering RNAs accelerates the migration and invasion of MTA1-overexpressing cells. These results suggest that RNF144A is partially responsible for MTA1-mediated migration and invasion and that MTA1 overexpression in highly metastatic cancer cells drives cell migration and invasion by, at least in part, interfering with the suppressive function of RNF144A through transcriptional repression of RNF144A expression. Together, these findings provide novel mechanistic insights into regulation of tumor progression and metastasis by MTA1 and highlight a previously unrecognized role of RNF144A in MTA1-driven cancer cell migration and invasion.
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Affiliation(s)
- Hezlin Marzook
- From the Cancer Research Program, Rajiv Gandhi Centre for Biotechnology, Thiruvananthapuram 695014, India and
| | - Da-Qiang Li
- the Department of Biochemistry and Molecular Biology, School of Medicine and Health Sciences, George Washington University, Washington, D. C. 20037
| | - Vasudha S. Nair
- the Department of Biochemistry and Molecular Biology, School of Medicine and Health Sciences, George Washington University, Washington, D. C. 20037
| | - Prakriti Mudvari
- the Department of Biochemistry and Molecular Biology, School of Medicine and Health Sciences, George Washington University, Washington, D. C. 20037
| | - Sirigiri Divijendra Natha Reddy
- the Department of Biochemistry and Molecular Biology, School of Medicine and Health Sciences, George Washington University, Washington, D. C. 20037
| | - Suresh B. Pakala
- the Department of Biochemistry and Molecular Biology, School of Medicine and Health Sciences, George Washington University, Washington, D. C. 20037
| | - T. R. Santhoshkumar
- From the Cancer Research Program, Rajiv Gandhi Centre for Biotechnology, Thiruvananthapuram 695014, India and
| | - M. Radhakrishna Pillai
- From the Cancer Research Program, Rajiv Gandhi Centre for Biotechnology, Thiruvananthapuram 695014, India and
| | - Rakesh Kumar
- From the Cancer Research Program, Rajiv Gandhi Centre for Biotechnology, Thiruvananthapuram 695014, India and
- the Department of Biochemistry and Molecular Biology, School of Medicine and Health Sciences, George Washington University, Washington, D. C. 20037
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Fairbank M, St-Pierre P, Nabi IR. The complex biology of autocrine motility factor/phosphoglucose isomerase (AMF/PGI) and its receptor, the gp78/AMFR E3 ubiquitin ligase. MOLECULAR BIOSYSTEMS 2009; 5:793-801. [PMID: 19603112 DOI: 10.1039/b820820b] [Citation(s) in RCA: 64] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/21/2022]
Abstract
Phosphoglucose isomerase (PGI) is a glycolytic enzyme that exhibits a dual function as an extracellular cytokine, under the name autocrine motility factor (AMF). Its cell surface receptor, gp78/AMFR, is also localized to the endoplasmic reticulum where it functions as an E3 ubiquitin ligase. Expression of both AMF/PGI and gp78/AMFR is associated with cancer and, in this review, we will discuss various aspects of the biology of this ligand-receptor complex and its role in tumor progression.
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Affiliation(s)
- Maria Fairbank
- Department of Cellular and Physiological Sciences, University of British Columbia, Vancouver, BC V6P 5V8, Canada
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16
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Repiso A, Andrés R, Climent F, Ureña JM. Expression Patterns in Mouse Embryos of Neuroleukin/Glucose-6-Phosphate Isomerase and Autocrine Motility Factor Receptor. Anat Histol Embryol 2008; 37:380-2. [DOI: 10.1111/j.1439-0264.2008.00865.x] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022]
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17
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Funasaka T, Raz A. The role of autocrine motility factor in tumor and tumor microenvironment. Cancer Metastasis Rev 2008; 26:725-35. [PMID: 17828376 DOI: 10.1007/s10555-007-9086-7] [Citation(s) in RCA: 54] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 10/22/2022]
Abstract
Autocrine motility factor (AMF) is a tumor-secreted cytokine and is abundant at tumor sites, where it may affect the process of tumor growth and metastasis. AMF is a multifunctional protein capable of affecting cell migration, invasion, proliferation, and survival, and possesses phosphoglucose isomerase activity and can catalyze the step in glycolysis and gluconeogenesis. Here, we review the role of AMF and tumor environment on malignant processes. The outcome of metastasis depends on multiple interactions between tumor cells and homeostatic mechanisms, therefore elucidation of the tumor/host interactions in the tumor microenvironment is essential in the development of new prevention and treatment strategies. Such knowledge might provide clues to develop new future therapeutic approaches for human cancers.
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Affiliation(s)
- Tatsuyoshi Funasaka
- Tumor Progression and Metastasis Program, Barbara Ann Karmanos Cancer Institute, School of Medicine, Wayne State University, Detroit, MI 48201, USA
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18
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Funasaka T, Hu H, Hogan V, Raz A. Down-regulation of phosphoglucose isomerase/autocrine motility factor expression sensitizes human fibrosarcoma cells to oxidative stress leading to cellular senescence. J Biol Chem 2007; 282:36362-9. [PMID: 17925402 DOI: 10.1074/jbc.m706301200] [Citation(s) in RCA: 26] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
Phosphoglucose isomerase/autocrine motility factor (PGI/AMF) is a housekeeping gene product present in all cells, is an essential enzyme of catabolic glycolysis and anabolic gluconeogenesis, and regulates tumor cell growth and metastasis. Because glycolytic enzyme up-regulation of expression contributes to glycolytic flux, leading to increased of cell growth and a resistance to cellular stress of normal fibroblasts whereas down-regulation of PGI/AMF leads to mesenchymal-to-epithelial transition in tumor cells, we examined the involvement of PGI/AMF in overcoming cellular senescence in cancer cells. PGI/AMF cellular expression in HT1080 human fibrosarcoma was down-regulated by small interfering RNA methodology, which resulted in an increased sensitivity to oxidative stress and oxidative stress-induced cellular senescence. Signaling analysis revealed that the senescence pathway involving p21 cyclin-dependent kinase inhibitor was up-regulated in PGI/AMF knockdown cells and that superoxide dismutase is the upstream regulator protein of p21-mediated cellular senescence. A specific inhibitor of PGI/AMF induced cellular senescence and p21 expression in tumor cells exposed to an oxidative stress environment. Taken together, the results presented here suggest that PGI/AMF is involved in oxidative stress-induced cellular senescence and should bring novel insights into the control of cellular growth leading to a new methodology for cancer treatment.
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Affiliation(s)
- Tatsuyoshi Funasaka
- Tumor Progression and Metastasis Program, Karmanos Cancer Institute, Wayne State University, School of Medicine, Detroit, Michigan 48201, USA
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19
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Funasaka T, Hu H, Yanagawa T, Hogan V, Raz A. Down-regulation of phosphoglucose isomerase/autocrine motility factor results in mesenchymal-to-epithelial transition of human lung fibrosarcoma cells. Cancer Res 2007; 67:4236-43. [PMID: 17483335 DOI: 10.1158/0008-5472.can-06-3935] [Citation(s) in RCA: 57] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022]
Abstract
Phosphoglucose isomerase (PGI) is one of the glycolytic enzymes and is a multifunctional enzyme that functions in glucose metabolism inside the cell while acting as a cytokine outside the cell, with properties that include autocrine motility factor (AMF) regulating tumor cell motility. Although there are many studies indicating that PGI/AMF has been implicated in progression of metastasis, no direct studies of the significance of exogenous PGI/AMF on tumor progression have been reported. Here, we report on the mesenchymal-to-epithelial transition (MET), which is the reverse phenomenon of the epithelial-to-mesenchymal transition that is associated with loss of cell polarity, loss of epithelia markers, and enhancement of cell motility essential for tumor cell invasion and metastasis. Mesenchymal human fibrosarcoma HT1080 cells, which have naturally high levels of endogenous and exogenous PGI/AMF, were stably transfected with PGI/AMF small interfering RNA (siRNA). The siRNA targeting human PGI/AMF down-regulated the endogenous PGI/AMF expression and completely extinguished the secretion of PGI/AMF in a human fibrosarcoma HT1080, whereas the control siRNA showed no effects. The PGI/AMF siRNA caused cells to change shape dramatically and inhibited cell motility and invasion markedly. Suppression of PGI/AMF led to a contact-dependent inhibition of cell growth. Those PGI/AMF siRNA-transfected cells showed epithelial phenotype. Furthermore, tumor cells with PGI/AMF deficiency lost their abilities to form tumor mass. This study identifies that MET in HT1080 human lung fibrosarcoma cells was initiated by down-regulation of the housekeeping gene product/cytokine PGI/AMF, and the results depicted here suggest a novel therapeutic target/modality for mesenchymal cancers.
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Affiliation(s)
- Tatsuyoshi Funasaka
- Tumor Progression and Metastasis Program, Karmanos Cancer Institute, Wayne State University, School of Medicine, Detroit, Michigan 48201, USA
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20
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21
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Hatayama K, Higuchi H, Kimura M, Takeda M, Ono H, Watanabe H, Takagishi K. Histologic changes after meniscal repair using radiofrequency energy in rabbits. Arthroscopy 2007; 23:299-304. [PMID: 17349474 DOI: 10.1016/j.arthro.2006.10.017] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/15/2006] [Revised: 09/22/2006] [Accepted: 10/31/2006] [Indexed: 02/02/2023]
Abstract
PURPOSE Our purpose was to investigate histologic changes in the rabbit meniscus after meniscal repair via radiofrequency energy (RFE). METHODS Twenty Japanese white rabbits underwent bilateral knee arthrotomies, and a longitudinal tear was made in the avascular area of both medial menisci. On the right knees, RFE treatment (60 degrees C and 40 W) was performed on the femoral surface of the meniscal tear in monopolar mode. On the left knees, the meniscus was left untreated as a control. The rabbits were killed at 0, 1, 2, 4, or 12 weeks after surgery, and all medial menisci were examined histologically. The expression of autocrine motility factor in meniscal fibrochondrocytes was examined by immunohistochemical analysis. RESULTS Histologic examination at baseline showed fusion of collagen fibers in the tear. Failure of fusion was found in 2 of 4 menisci at 2 weeks and 1 of 4 menisci at 4 and 12 weeks. One week after surgery, the specimens showed an acellular area as a result of fibrochondrocyte death. The acellular area expanded deeper until 4 weeks and was reduced at 12 weeks. On the femoral surface of the RFE-treated area, fibroblast proliferation was found at 2 weeks, and fibroblasts had invaded into the meniscus tissue from the meniscal surface at 12 weeks. Immunohistochemical analysis showed that the expression of autocrine motility factor in RFE-treated menisci was significantly higher than that in control menisci from 1 to 12 weeks. CONCLUSIONS RFE treatment at 60 degrees C and 40 W fused the collagen fiber in the meniscal tear in rabbits just after surgery. After RFE treatment, an acellular area developed as a result of fibrochondrocyte damage. RFE caused fibroblast proliferation at 2 weeks. The acellular area was reduced by cell repopulation at 12 weeks. CLINICAL RELEVANCE RFE may induce fibroblast proliferation for meniscal repair.
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Affiliation(s)
- Kazuhisa Hatayama
- Department of Orthopaedic Surgery, Gunma University Faculty of Medicine, Maebashi, Japan.
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22
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Haga A, Tanaka N, Funasaka T, Hashimoto K, Nakamura KT, Watanabe H, Raz A, Nagase H. The Autocrine Motility Factor (AMF) and AMF-receptor Combination Needs Sugar Chain Recognition Ability and Interaction Using the C-terminal Region of AMF. J Mol Biol 2006; 358:741-53. [PMID: 16563432 DOI: 10.1016/j.jmb.2006.02.046] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/29/2005] [Revised: 02/14/2006] [Accepted: 02/16/2006] [Indexed: 11/21/2022]
Abstract
The autocrine motility factor (AMF) promotes cellular locomotion or invasion, and regulates tumor angiogenesis or ascites accumulation. These signals are triggered by binding between AMF and its receptor (AMFR), a glycoprotein on the cell surface. AMF has been identified as phosphohexose isomerase (PHI). Previous reports have suggested that the substrate-recognition of exo-PHI is significant for receptor binding. Crystallographic studies have shown that AMF consists of three domains, and that the substrate or inhibitor of PHI is stored between the large and small domains, corresponding to approximately residues 117-288. Here, site-directed mutagenesis was used to investigate 18 recombinant human AMF point mutants involving critical amino acid residues for substrate or enzyme inhibitor recognition or binding. Mutation of residues that interact with the phosphate group of the PHI substrate significantly reduced the cell motility-stimulating activity. Their binding capacities for AMFR were also lower than wild-type human AMF. Mutants that retained the enzymic activity showed the motility-stimulating effect and receptor binding and had sensitivity to a PHI inhibitor. Mutant AMFR lacking the N-sugar chain was expressed on the cell membrane but did not respond to AMF-stimulation, and N-glycosidase-treated AMFR did not compete with receptor binding of AMF. Furthermore, the AMF domains that contain the substrate storage domain and C-terminal region stimulate cell locomotion. These results suggest that the N-glyco side-chain of AMFR is a trigger and that interaction between the 117-C-terminal part of AMF and the extracellular core protein of AMFR is needed during AMF-AMFR interactions.
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Affiliation(s)
- Arayo Haga
- Gifu Pharmaceutical University, 5-6-1 Mitahora-Higashi, Gifu 502-8585, Japan.
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23
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Comparative analysis of lymphatic metastasis — Associated genes in mouse hepatocellular carcinoma cell lines with different metastatic potential. Chin J Cancer Res 2006. [DOI: 10.1007/s11670-006-0026-9] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/23/2022] Open
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24
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Kikkert M, Hassink G, Wiertz E. The role of the ubiquitination machinery in dislocation and degradation of endoplasmic reticulum proteins. Curr Top Microbiol Immunol 2006; 300:57-93. [PMID: 16573237 DOI: 10.1007/3-540-28007-3_4] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/30/2022]
Abstract
Ubiquitination is essential for the dislocation and degradation of proteins from the endoplasmic reticulum (ER). How exactly this is regulated is unknown at present. This review provides an overview of ubiquitin-conjugating enzymes (E2s) and ubiquitin ligases (E3s) with a role in the degradation of ER proteins. Their structure and functions are described, as well as their mutual interactions. Substrate specificity and functional redundancy of E3 ligases are discussed, and other components of the ER degradation machinery that may associate with the ubiquitination system are reviewed.
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Affiliation(s)
- M Kikkert
- Department of Medical Microbiology, Leiden University Medical Center (LUMC), Albinusdreef 2, 2333 ZA Leiden, The Netherlands.
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25
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Abstract
Relatively small genomes and high replication rates allow viruses and bacteria to accumulate mutations. This continuously presents the host immune system with new challenges. On the other side of the trenches, an increasingly well-adjusted host immune response, shaped by coevolutionary history, makes a pathogen's life a rather complicated endeavor. It is, therefore, no surprise that pathogens either escape detection or modulate the host immune response, often by redirecting normal cellular pathways to their advantage. For the purpose of this chapter, we focus mainly on the manipulation of the class I and class II major histocompatibility complex (MHC) antigen presentation pathways and the ubiquitin (Ub)-proteasome system by both viral and bacterial pathogens. First, we describe the general features of antigen presentation pathways and the Ub-proteasome system and then address how they are manipulated by pathogens. We discuss the many human cytomegalovirus (HCMV)-encoded immunomodulatory genes that interfere with antigen presentation (immunoevasins) and focus on the HCMV immunoevasins US2 and US11, which induce the degradation of class I MHC heavy chains by the proteasome by catalyzing their export from the endoplasmic reticulum (ER)-membrane into the cytosol, a process termed ER dislocation. US2- and US11-mediated subversion of ER dislocation ensures proteasomal degradation of class I MHC molecules and presumably allows HCMV to avoid recognition by cytotoxic T cells, whilst providing insight into general aspects of ER-associated degradation (ERAD) which is used by eukaryotic cells to purge their ER of defective proteins. We discuss the similarities and differences between the distinct pathways co-opted by US2 and US11 for dislocation and degradation of human class I MHC molecules and also a putatively distinct pathway utilized by the murine herpes virus (MHV)-68 mK3 immunoevasin for ER dislocation of murine class I MHC. We speculate on the implications of the three pathogen-exploited dislocation pathways to cellular ER quality control. Moreover, we discuss the ubiquitin (Ub)-proteasome system and its position at the core of antigen presentation as proteolysis and intracellular trafficking rely heavily on Ub-dependent processes. We add a few examples of manipulation of the Ub-proteasome system by pathogens in the context of the immune system and such diverse aspects of the host-pathogen relationship as virus budding, bacterial chromosome integration, and programmed cell death, to name a few. Finally, we speculate on newly found pathogen-encoded deubiquitinating enzymes (DUBs) and their putative roles in modulation of host-pathogen interactions.
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Affiliation(s)
- Joana Loureiro
- Whitehead Institute, 9 Cambridge Center, Cambridge, Massachusetts, USA
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26
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Gong W, Jiang Y, Wang L, Wei D, Yao J, Huang S, Fang S, Xie K. Expression of Autocrine Motility Factor Correlates with the Angiogenic Phenotype of and Poor Prognosis for Human Gastric Cancer. Clin Cancer Res 2005; 11:5778-83. [PMID: 16115916 DOI: 10.1158/1078-0432.ccr-05-0214] [Citation(s) in RCA: 16] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022]
Abstract
Autocrine motility factor (AMF) is a cytokine known to regulate tumor cell motility. Recent studies have extended its role to many other aspects of cancer biology. In the present study, we examined the level of AMF expression and its relationship with vascular endothelial growth factor (VEGF) expression and the angiogenic phenotype in human gastric cancer and their effect on survival. The AMF and VEGF expression level and tumor microvessel density (MVD) status in archived tissue specimens from 86 resected gastric cancer cases were determined. AMF expression was significantly higher in both primary tumors and lymph node metastases than in adjacent normal gastric mucosa and normal gastric mucosa from individuals without gastric cancer. In univariate survival analyses, strong AMF expression was associated with inferior survival (P = 0.028). In a Cox proportional hazards model, strong AMF expression (P = 0.019) was independently prognostic of poor survival. Strong AMF expression in the lymph node metastases was associated with poor survival (P = 0.011). Furthermore, AMF expression in the primary tumors was directly correlated with VEGF expression and MVD status. We found the first clinical evidence that AMF expression is directly correlated with VEGF expression and MVD status and predicts clinical outcome in patients with gastric cancer, supporting the hypothesis that the AMF/AMF receptor pathway plays an important role in multiple aspects of cancer biology.
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Affiliation(s)
- Weida Gong
- Department of Neurosurgery, University of Texas M.D. Anderson Cancer Center, Houston, Texas 77030, USA
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27
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Song B, Tang JW, Wang B, Cui XN, Hou L, Sun L, Mao LM, Zhou CH, Du Y, Wang LH, Wang HX, Zheng RS, Sun L. Identify lymphatic metastasis-associated genes in mouse hepatocarcinoma cell lines using gene chip. World J Gastroenterol 2005; 11:1463-72. [PMID: 15770722 PMCID: PMC4305688 DOI: 10.3748/wjg.v11.i10.1463] [Citation(s) in RCA: 40] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: In order to obtain lymphogenous metastasis-associated genes, we compared the transcriptional profiles of mouse hepatocarcinoma cell lines Hca-F with highly lymphatic metastasis potential and Hca-P with low lymphatic metastasis potential.
METHODS: Total RNA was isolated from Hca-F and Hca-P cells and synthesized into double-stranded cDNA. In vitro transcription double-stranded cDNA was labeled with biotin (i.e., biotin-labeled cRNA, used as the probe). The cRNA probes hybridized with Affymetrix GeneChip® MOE430A (containing 22690 transcripts, including 14500 known mouse genes and 4371 ESTs) respectively and the signals were scanned by the GeneArray Scanner. The results were then analyzed by bioinformatics.
RESULTS: Out of the 14500 known genes investigated, 110 (0.8%) were up regulated at least 23 fold. Among the total 4371 ESTs, 17 ESTs (0.4%) (data were not presented) were up regulated at least 23 fold. According to the Gene Ontology and TreeView analysis, the 110 genes were further classified into two groups: differential biological process profile and molecular function profile.
CONCLUSION: Using high-throughput gene chip method, a large number of genes and their cellular functions about angiogenesis, cell adhesion, signal transduction, cell motility, transport, microtubule-based process, cytoskeleton organization and biogenesis, cell cycle, transcription, chaperone activity, motor activity, protein kinase activity, receptor binding and protein binding might be involved in the process of lymphatic metastasis and deserve to be used as potential candidates for further investigation. Cyclin D1, Fosl1, Hsp47, EGFR and AR, and Cav-1 are selected as the possible candidate genes of the metastatic phenotype, which need to be validated in later experiments. ESTs (data were not presented) might indicate novel genes associated with lymphatic metastasis. Validating the function of these genes is helpful to identify the key or candidate gene/pathway responsible for lymphatic metastasis, which might be used as the diagnostic markers and the therapeutic targets for lymphatic metastasis.
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Affiliation(s)
- Bo Song
- Department of Pathology, Dalian Medical University, Dalian 116027, Liaoning Province, China
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Registre M, Goetz JG, St Pierre P, Pang H, Lagacé M, Bouvier M, Le PU, Nabi IR. The gene product of the gp78/AMFR ubiquitin E3 ligase cDNA is selectively recognized by the 3F3A antibody within a subdomain of the endoplasmic reticulum. Biochem Biophys Res Commun 2004; 320:1316-22. [PMID: 15303277 DOI: 10.1016/j.bbrc.2004.06.089] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/19/2022]
Abstract
The receptor for the autocrine motility factor/phosphoglucose isomerase cytokine (gp78 or AMFR) has been extensively characterized using the 3F3A monoclonal antibody. Cloning of AMFR identified a seven-transmembrane domain G-protein-coupled receptor ubiquitin E3 ligase whose identity as AMFR was based on prior expression cloning with the 3F3A mAb that generated a truncated sequence. We show here that the gp78/AMFR gene product is indeed recognized by the 3F3A mAb. The FLAG-taggedAMFR immunoprecipitated with an anti-FLAG antibody was recognized by the 3F3A mAb in Western blot analysis and cells transfected with AMFR exhibit increased labeling with the 3F3A mAb. The 3F3A mAb does not however recognize higher molecular weight isoforms of AMFR. 3F3A labeling colocalizes with tagged AMFR in a peripheral ER network but does not recognize FLAG- or GFP-tagged AMFR localized to a perinuclear ER domain that likely corresponds to misfolded forms of the protein retained in the ER. These data indicate that 3F3A antibody binding is highly specific for a subpopulation of AMFR localized to an ER subdomain. Coexpression of AMFR-GFP and a lumenal ER-targeted RFP presented extensive colocalization in living cells andAMFR-GFP is concentrated in a basal ER network morphologically similar to that labeled by the 3F3A mAb in fixed cells. The3F3A anti-AMFR mAb therefore selectively recognizes a subpopulation of expressed AMFR localized to a subdomain of the ER.
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Affiliation(s)
- Marilyn Registre
- Département de pathologie et biologie cellulaire, Université de Montréal, Que., Canada
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29
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Yanagawa T, Watanabe H, Takeuchi T, Fujimoto S, Kurihara H, Takagishi K. Overexpression of autocrine motility factor in metastatic tumor cells: possible association with augmented expression of KIF3A and GDI-beta. J Transl Med 2004; 84:513-22. [PMID: 14968121 DOI: 10.1038/labinvest.3700057] [Citation(s) in RCA: 37] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/09/2022] Open
Abstract
Autocrine motility factor (AMF), which is identical to phosphohexose isomerase (PHI)/glucose-6-phosphate isomerase (GPI), a ubiquitous enzyme essential for glycolysis, neuroleukin (NLK), a neurotrophic growth factor, and maturation factor (MF) mediating the differentiation of human myeloid cells, enhances the motility and metastatic ability of tumor cells. AMF/PHI activity is elevated in the serum or urine in patients with malignant tumors. Here, we constructed an amf/phi/nlk/mf gene using adenovirus vector and transfected into two tumor cell lines. Overexpression of AMF/PHI/NLK/MF enhanced AMF secretion into the culture media in both tumor cell lines. However, upregulation of motility and metastatic ability was found only in metastatic fibrosarcoma cells expressing an AMF receptor, gp78, and was not found in gp78-undetectable osteosarcoma cells. Thus, not only serum AMF activity but also gp78-expression in tumor cells may be required for metastasis-related motility induction. With the use of microarray analyses, we detected two augmented genes, rho GDP dissociation inhibitor beta and kinesin motor 3A, as well as AMF itself. The RNA message and protein expression of these two molecules was confirmed to be upregulated, suggesting a possible association with AMF-induced signaling for cell motility and metastasis.
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Affiliation(s)
- Takashi Yanagawa
- Department of Orthopaedic Surgery, Institute for Molecular and Cellular Regulation, Gunma University, 3-39-22 Showa, Maebashi, Gunma 371-8511, Japan
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Haga A, Funasaka T, Niinaka Y, Raz A, Nagase H. Autocrine motility factor signaling induces tumor apoptotic resistance by regulations Apaf-1 and Caspase-9 apoptosome expression. Int J Cancer 2004; 107:707-14. [PMID: 14566819 DOI: 10.1002/ijc.11449] [Citation(s) in RCA: 64] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2022]
Abstract
Autocrine motility factor (AMF) is a cytokine that regulates locomotion and metastasis of tumor cells. It is well known that expression levels of AMF secretion and its receptor (AMF R) are closely related to tumor malignancy and rheumatoid arthritis. We have established that AMF signaling induced anti-apoptotic activity and that human fibrosarcoma HT-1080 line that secreted high levels of AMF were resistant to drug-induced apoptosis. These cells did not express the apoptotic protease activating factor-1 (Apaf-1) and Caspase-9 genes that encode for the proteins that form the "apoptosome" complex. The disappearance of the Apaf-1 and Caspase-9 gene was recovered by a cellular signaling inhibitor of protein kinase C, phosphatidylinositol 3-phosphate kinase and mitogen-activated protein kinase of the in vitro cultured human fibrosarcoma HT-1080 line. Treatment with these inhibitors favored apoptotic cell death induced by anti-cancer drugs of the murine ascites Ehrlich line. Apoptotic resistance of tumor cells allows them to escape death from cancer chemotherapy, so an understanding of malignant anti-apoptotic activities is important. Antibodies against AMF induced Ehrlich ascites apoptosis in vitro, and effectively aided in vivo apoptosis induced by anti-cancer drugs. The results might indicate a novel route by which tumor cells protect themselves with products, such as AMF, and proliferate despite various stresses and chemical insults; AMF regulates expression of Apaf-1 and caspase-9 genes via a complex signaling pathway and indirectly regulates formation of the apoptosome.
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Affiliation(s)
- Arayo Haga
- Department of Hygienics, Gifu Pharmaceutical University, Gifu, Japan.
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Tsutsumi S, Yanagawa T, Shimura T, Fukumori T, Hogan V, Kuwano H, Raz A. Regulation of cell proliferation by autocrine motility factor/phosphoglucose isomerase signaling. J Biol Chem 2003; 278:32165-72. [PMID: 12783864 DOI: 10.1074/jbc.m304537200] [Citation(s) in RCA: 48] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/18/2022] Open
Abstract
Autocrine motility factor (AMF)/phosphoglucose isomerase (PGI; EC 5.3.1.9) is a housekeeping cytosolic enzyme that plays a key role in both glycolysis and gluconeogenesis pathways. AMF/PGI is also a multifunctional protein that displays cytokine properties, eliciting mitogenic, motogenic, and differentiation activities, and has been implicated in tumor progression and metastasis. Because little is known about AMF/PGI-dependent signaling in general and during tumorigenesis in particular, we sought to study its effect on the cell cycle. To elucidate the functional role of PGI, we stably transfected its cDNA into NIH/3T3 and BALB/c 3T3-A31 fibroblasts. Ectopic overexpression of PGI results in the acquisition of a transformed phenotype associated with an acceleration of G1 to S cell cycle transition. These were manifested by up-regulation of cyclin D1 expression and cyclin-dependent kinase activity and down-regulation of the cyclin-dependent kinase inhibitor p27Kip1. The reduced p27Kip1 protein expression level in PGI-overexpressing cells could be restored to control levels by treatment with proteasome inhibitor. PGI-overexpressing cells also exhibited elevated expression of Skp2 involved in p27Kip1 ubiquitination and elevation in the levels of retinoblastoma protein hyperphosphorylation. Thus, we may conclude that the overexpression of AMF/PGI enhances cell proliferation together with up-regulation of cyclin/cyclin-dependent kinase activities and down-regulation of p27Kip1, whereas the induction of 3T3 fibroblast transformation by PGI is regulated by the retinoblastoma protein pathway.
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Affiliation(s)
- Soichi Tsutsumi
- Tumor Progression and Metastasis, Karmanos Cancer Institute, The Department of Pathology, Wayne State University, School of Medicine, Detroit, Michigan 48201, USA
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Abstract
The presence of activated oncogenes and/or inactivated tumor suppressor genes may result in constitutive activation of multiple transcription factors. This may be especially true in the early stages of tumor development. At advanced stages, however, uncontrolled tumor growth and the consequent development of a stress microenvironment, such as hypoxia, acidosis, and free radical overproduction, may further alter the activity of these transcription factors. Abnormal activation of and interplay between these factors lead to aberrant expression of multiple metastasis-related proteins and confer a tremendous survival and growth advantage to emerging metastatic variants. Understanding the expression and regulation of these molecules may shed more light on the biology of cancer metastasis as well as suggest new preventive and therapeutic approaches.
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Affiliation(s)
- Keping Xie
- Department of Gastrointestinal Medical Oncology, The University of Texas M. D. Anderson Cancer Center, Houston, Texas 77030, USA
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Onishi Y, Tsukada K, Yokota J, Raz A. Overexpression of autocrine motility factor receptor (AMFR) in NIH3T3 fibroblasts induces cell transformation. Clin Exp Metastasis 2003; 20:51-8. [PMID: 12650607 DOI: 10.1023/a:1022594503657] [Citation(s) in RCA: 19] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/12/2022]
Abstract
Autocrine motility factor receptor (AMFR) is a cell surface glycoprotein of 78000 molecular weight (gp78), regulating cell motility signaling in vitro and metastasis in vivo. To test whether AMFR could be a common mediator of transformation and oncogenic itself, we transfected NIH3T3 fibroblast cells with expression vectors carrying the full-length cDNA for mouse AMFR and evaluated the effects of increased AMFR on transforming potential. The cells stably expressing high levels of AMFR as a result of transfection displayed a complete morphological change and acquired the ability to grow even in low serum. Furthermore, they were anchorage-independent for growth in soft agar and more motile in phagokinetic track assay. Interestingly, the enhanced expression of AMFR produced tumors in nude mice. Our findings provide a direct evidence that overexpression of the AMFR is associated with the acquisition of a transformation phenotype.
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Affiliation(s)
- Yasuharu Onishi
- Tumor Progression and Metastasis Program, Karmanos Cancer Institute, Detroit, Michigan 48201, USA
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Fang S, Lorick KL, Jensen JP, Weissman AM. RING finger ubiquitin protein ligases: implications for tumorigenesis, metastasis and for molecular targets in cancer. Semin Cancer Biol 2003; 13:5-14. [PMID: 12507552 DOI: 10.1016/s1044-579x(02)00095-0] [Citation(s) in RCA: 105] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/24/2022]
Abstract
Covalent modification of proteins with ubiquitin regulates almost all aspects of eukaryotic cellular function. Ubiquitin protein ligases (E3s) play central regulatory roles in that they provide substrate specificity to this process and therefore, represent attractive molecular targets for disease therapy. We summarize recent advances in our understanding of RING finger and RING finger-related E3s with emphasis on BRCA1 and the tumor autocrine motility factor receptor (gp78), as well as discuss the potential for components of the ubiquitin pathway for proteasomal degradation as molecular targets.
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Affiliation(s)
- Shengyun Fang
- Regulation of Protein Function Laboratory, Center for Cancer Research, National Cancer Institute at Frederick, Building 560, Room 22-95, 1050 Boyles Street, Frederick, MD 21702, USA
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Niizeki H, Kobayashi M, Horiuchi I, Akakura N, Chen J, Wang J, Hamada JI, Seth P, Katoh H, Watanabe H, Raz A, Hosokawa M. Hypoxia enhances the expression of autocrine motility factor and the motility of human pancreatic cancer cells. Br J Cancer 2002; 86:1914-9. [PMID: 12085186 PMCID: PMC2375441 DOI: 10.1038/sj.bjc.6600331] [Citation(s) in RCA: 74] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/28/2001] [Revised: 03/19/2002] [Accepted: 03/27/2002] [Indexed: 12/16/2022] Open
Abstract
The incidence of distant metastases is higher in the tumours with low oxygen pressure than in those with high oxygen pressure. It is well known that hypoxia induces the transcription of various genes involved in angiogenesis and anaerobic metabolism necessary for the growth of tumour cells in vivo, suggesting that hypoxia may also induce the transcription of metastasis-associated genes. We sought to identify the metastasis-associated genes differentially expressed in tumour cells under hypoxic conditions with the use of a DNA microarray system. We found that hypoxia enhanced the expression of autocrine motility factor mRNA in various cancer cells and also enhanced the random motility of pancreatic cancer cells. Autocrine motility factor inhibitors abrogated the increase of motility under hypoxic conditions. In order to explore the roles of hypoxia-inducible factor-1alpha, we established hypoxia-inducible factor-1alpha-transfectants and dominant negative hypoxia-inducible factor-1alpha-transfectants. Transfection with hypoxia-inducible factor-1alpha and dominant-negative hypoxia-inducible factor-1alpha enhanced and suppressed the expression of autocrine motility factor/phosphohexase isomerase/neuroleukin mRNA and the random motility, respectively. These results suggest that hypoxia may promote the metastatic potential of cancer cells through the enhanced autocrine motility factor/phosphohexase isomerase/neuroleukin mRNA expression and that the disruption of the hypoxia-inducible factor-1 pathway may be an effective treatment for metastasis.
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Affiliation(s)
- H Niizeki
- Division of Cancer Pathobiology, Institute for Genetic Medicine, Hokkaido University, Kita-15, Nishi-7, Kita-ku, Sapporo 060-0815, Japan
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36
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Zhi J, Sommerfeldt DW, Rubin CT, Hadjiargyrou M. Differential expression of neuroleukin in osseous tissues and its involvement in mineralization during osteoblast differentiation. J Bone Miner Res 2001; 16:1994-2004. [PMID: 11697795 DOI: 10.1359/jbmr.2001.16.11.1994] [Citation(s) in RCA: 24] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/18/2022]
Abstract
Osteoblast differentiation is a multistep process that involves critical spatial and temporal regulation of cellular processes marked by the presence of a large number of differentially expressed molecules. To identify key functional molecules, we used differential messenger RNA (mRNA) display and compared RNA populations isolated from the defined transition phases (proliferation, matrix formation, and mineralization) of the MC3T3-E1 osteoblast-like cell line. Using this approach, a complementary DNA (cDNA) fragment was isolated and identified as neuroleukin (NLK), a multifunctional cytokine also known as autocrine motility factor (AMF), phosphoglucose isomerase (PGI; phosphohexose isomerase [PHI]), and maturation factor (MF). Northern analysis showed NLK temporal expression during MC3T3-E1 cell differentiation with a 3.5-fold increase during matrix formation and mineralization. Immunocytochemical studies revealed the presence of NLK in MC3T3-E1 cells as well as in the surrounding matrix, consistent with a secreted molecule. In contrast, the NLK receptor protein was detected primarily on the cell membrane. In subsequent studies, a high level of NLK expression was identified in osteoblasts and superficial articular chondrocytes in bone of 1-, 4-, and 8-month-old normal mice, as well as in fibroblasts, proliferating chondrocytes, and osteoblasts within a fracture callus. However, NLK was not evident in hypertrophic chondrocytes or osteocytes. In addition, treatment of MC3T3 cells with 6-phosphogluconic acid (6PGA; a NLK inhibitor) resulted in diminishing alkaline phosphatase (ALP) activity and mineralization in MC3T3-E1 cells, especially during the matrix formation stage of differentiating cells. Taken together, these data show specific expression of NLK in discrete populations of bone and cartilage cells and suggest a possible role for this secreted protein in bone development and regeneration.
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Affiliation(s)
- J Zhi
- Department of Biomedical Engineering, State University of New York at Stony Brook, 11794-2580, USA
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37
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Funasaka T, Haga A, Raz A, Nagase H. Tumor autocrine motility factor is an angiogenic factor that stimulates endothelial cell motility. Biochem Biophys Res Commun 2001; 285:118-28. [PMID: 11437381 DOI: 10.1006/bbrc.2001.5135] [Citation(s) in RCA: 42] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
Abstract
Autocrine motility factor (AMF) is a type of tumor-secreted cytokine which primarily stimulates tumor cell motility via receptor-mediated signaling pathways, and is thought to be connected to tumor progression and metastasis. Using in vivo models, we showed that critical neovascularization responded to a biological amount of AMF. This angiogenic activity was fixed by specific inhibitors against AMF. AMF stimulated in vitro motility of human umbilical vein endothelial cells (HUVECs), inducing the expression of cell surface AMF receptor localizing a single predominant perinuclear pattern closely correlated with its motile ability. AMF also elicited the formation of tube-like structures mimicking angiogenesis when HUVECs were grown in three-dimensional type I collagen gels. We further immunohistochemically detected AMF receptors on the surrounding sites of newborn microvessels. These findings suggest that AMF is a possible tumor progressive angiogenic factor which may act in a paracrine manner for the endothelial cells in the clinical neoplasm, and it will be a new target for antiangiogenic treatment.
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Affiliation(s)
- T Funasaka
- Department of Hygienics, Gifu Pharmaceutical University, Gifu, 502, Japan
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38
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Funasaka T, Haga A, Raz A, Nagase H. Tumor autocrine motility factor is an angiogenic factor hat stimulates endothelial cell motility. Biochem Biophys Res Commun 2001; 284:1116-25. [PMID: 11414699 DOI: 10.1006/bbrc.2001.4912] [Citation(s) in RCA: 24] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/28/2023]
Abstract
Autocrine motility factor (AMF) is a type of tumor-secreted cytokine that primarily stimulates tumor cell motility via receptor-mediated signaling pathways and is thought to be connected to tumor progression and metastasis. Using in vivo models, we showed that critical neovascularization responded to a biological amount of AMF. This angiogenic activity was fixed by specific inhibitors against AMF. AMF stimulated in vitro motility of human umbilical vein endothelial cells (HUVECs), inducing the expression of cell surface AMF receptor localizing a single predominant perinuclear pattern closely correlated with its motile ability. AMF also elicited the formation of tube-like structures mimicking angiogenesis when HUVECs were grown in three-dimensional type I collagen gels. We further immunohistochemically detected AMF receptors on the surrounding sites of newborn microvessels. These findings suggest that AMF is a possible tumor progressive angiogenic factor which may act in a paracrine manner for the endothelial cells in the clinical neoplasm, and it will be a new target for anti-angiogenic treatment.
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Affiliation(s)
- T Funasaka
- Department of Hygienics, Gifu Pharmaceutical University, Gifu, 502, Japan
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39
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Rotsch C, Jacobson K, Condeelis J, Radmacher M. EGF-stimulated lamellipod extension in adenocarcinoma cells. Ultramicroscopy 2001; 86:97-106. [PMID: 11215638 DOI: 10.1016/s0304-3991(00)00102-9] [Citation(s) in RCA: 37] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
Abstract
The extension of lamellipodia has been triggered by the application of epidermal growth factor (EGF). We have used an atomic force microscope (AFM) to investigate this lamellipodial extension. During extension we could detect an increase in height from about 500 nm for the stable lamellipodium to typical values of 600-800 nm for the extending lamellipodium. The AFM was also used to determine the mechanical properties of the lamellipodium where we found a decrease of the elastic modulus by a factor of 1.4 at the same location within the same cell. Both findings are consistent with the cortical expansion hypothesis, suggesting that severing of actin filaments, leading to a swelling of the cytoskeleton, generates the protrusive force during lamellipodial extension.
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Affiliation(s)
- C Rotsch
- Lehrstuhl für Angewandte Physik, Ludwig-Maximilians Universität München, Germany
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40
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Abstract
Genetic instability and an accumulation of genetic and epigenetic changes during tumor progression lead to an increasingly aggressive and treatment-resistant phenotype, and ultimately metastasis. In recent years it has become well established that angiogenesis, the process by which new vasculature is formed from pre-existing vessels, is an essential component to primary tumor growth and distant metastasis. A greater understanding of the complex multitude of factors involved in tumor angiogenesis and metastasis is fundamental to the development of potential therapeutics to treat malignant disease. As highlighted throughout this review, angiogenesis and metastasis share many common cellular and molecular features. We will briefly discuss the pertinent genes involved in the regulation of angiogenesis and metastasis.
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Affiliation(s)
- C P Webb
- Van Andel Research Institute, Grand Rapids, MI 49503, USA.
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41
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Abstract
E-cadherin and the associated catenin complex have been recognised as performing a key role in cell adhesion. Loss of cell adhesion is seen as a key step in the cascade leading to tumour metastasis. The ability of both extra- and intracellular factors to regulate E-cadherin-mediated cell adhesion in physiological processes has provided insight into both the interactions of the E-cadherin-catenin complex, and possible mechanisms utilised by tumours in the process of metastasis. The interaction of the E-cadherin-catenin complex with various regulating factors, their effect on cell signalling pathways, and the relationship with the metastatic potential of tumours are reviewed.
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Affiliation(s)
- I R Beavon
- Department of Anatomical Pathology, School of Pathology, South African Institute for Medical Research and University of the Witwatersrand, Johannesburg, South Africa.
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42
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Haga A, Niinaka Y, Raz A. Phosphohexose isomerase/autocrine motility factor/neuroleukin/maturation factor is a multifunctional phosphoprotein. BIOCHIMICA ET BIOPHYSICA ACTA 2000; 1480:235-44. [PMID: 11004567 DOI: 10.1016/s0167-4838(00)00075-3] [Citation(s) in RCA: 69] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/23/2022]
Abstract
Phosphohexose isomerase (PHI) is a member of the ectoenzyme/exoenzyme family and plays a key role in both glycolysis and gluconeogenesis pathways. Upon secretion PHI acts as a cytokine with tumor autocrine motility factor (AMF), neuroleukin (NLK) and maturation factor (MF) functions. Signaling is initiated by its binding to a cell surface 78 kDa glycoprotein (gp78). However, since PHI protein is a 'leaderless' secretory protein, released from cells via a non-classical route(s), we questioned whether the molecule undergoes post-translation modification while retaining proper folding and maintaining intact enzymatic and motogenic activities. To address this, we have generated, expressed and isolated a recombinant human AMF (rhAMF). The rhAMF retained the biological activities of the native AMF, i.e., catalyzes phosphohexose isomerization and stimulated cell motility. Additionally, we show here that human PHI is phosphorylated at serine 185 by casein kinase II (CK II) and we provide experimental evidence suggesting that this phosphorylation is associated with secretion, thus providing insights for elucidating the intracellular signal transmission of cell response to stimulation by AMF/NLK/MF.
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Affiliation(s)
- A Haga
- Metastasis Research Program, Karmanos Cancer Institute, Detroit, MI 48201, USA
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43
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Abstract
The peptide sequence of autocrine motility factor (AMF), a tumor secreted cytokine that induces cell motility, corresponds to that of the previously identified cytokine/enzyme, neuroleukin/glucose-6-phosphate isomerase. Neuroleukin is a neurotrophic factor that promotes neuronal survival and sprouting at the neuromuscular junction. The AMF receptor (AMF-R) has been identified and shown to be highly expressed in malignant tumors with minimal expression in adjacent normal tissue. Neuroleukin mRNA is highly expressed in the cerebellum and we therefore undertook a developmental study of AMF-R expression in rat cerebellum. As determined by immunoblot, AMF-R is expressed at equivalent high levels in brain and cerebellum of postnatal day 5 (P5) and 12 (P12) rats and at significantly reduced levels in the adult. Coimmunofluorescence studies with MAP-2 and gamma-actin revealed that at P12, AMF-R was mainly localized to Purkinje and granule cells. Moreover, the premigratory cells of the external granular layer were also immunoreactive for AMF-R suggesting a role for AMF-R in granule cell migration during cerebellar development in the first two weeks after birth. In the adult, AMF-R distribution was similar to P12, although weaker, and was localized to Purkinje and granule cells. AMF-R labeling of GFAP positive glial processes could not be detected in cerebellar sections although in cerebellar primary cultures, both neurons and glial cells were labeled for AMF-R. In neurons, AMF-R labeling was present in the cell body, neurites and growth cones. These data indicate that regulation of the neurotrophic function of neuroleukin might be regulated spatially and temporally by expression of its receptor, AMF-R, in developing and adult cerebellum.
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Affiliation(s)
- N Leclerc
- Département de pathologie et biologie cellulaire, Université de Montréal, Québec, Canada
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44
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Shimizu K, Tani M, Watanabe H, Nagamachi Y, Niinaka Y, Shiroishi T, Ohwada S, Raz A, Yokota J. The autocrine motility factor receptor gene encodes a novel type of seven transmembrane protein. FEBS Lett 1999; 456:295-300. [PMID: 10456327 DOI: 10.1016/s0014-5793(99)00966-7] [Citation(s) in RCA: 97] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/28/2022]
Abstract
Autocrine motility factor receptor (AMFR) is a cell surface glycoprotein of molecular weight 78,000 (gp78), mediating cell motility signaling in vitro and metastasis in vivo. Here, we cloned the full-length cDNAs for both human and mouse AMFR genes. Both genes encode a protein of 643 amino acids containing a seven transmembrane domain, a RING-H2 motif and a leucine zipper motif and showed a 94.7% amino acid sequence identity to each other. Analysis of the amino acid sequence of AMFR with protein databases revealed no significant homology with all known seven transmembrane proteins, but a significant structural similarity to a hypothetical protein of Caenorhabditis elegans, F26E4.11. Thus, AMFR is a highly conserved gene which encodes a novel type of seven transmembrane protein.
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Affiliation(s)
- K Shimizu
- Biology Division, National Cancer Center Research Institute, Tokyo, Japan
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45
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Abstract
The ability of tumours to metastasis is regarded as one of the hallmarks of malignancy. The process through which tumours evolve to achieve this has been termed the metastatic cascade. This cascade has been the subject of much investigation over many years. One of the vital events identified by these investigations is the reduction of adhesion between tumour cells facilitating invasion of the surrounding tissues and vascular channels, ultimately leading to the development of a distant metastasis. E-cadherin and its associated catenin complex have been identified as key molecules in cell adhesion. This review looks at the structure and interaction of the E-cadherin-catenin complex and the factors that appear to regulate E-cadherin expression and thus cell adhesion. From the data gathered, it has become possible to propose the hypothesis that the development of tumour hypoxia is the initiating factor that sets the tumour on the road to metastasis.
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Affiliation(s)
- I R Beavon
- Department of Anatomical Pathology, School of Pathology, South African Institute for Medical Research, Johannesburg, South Africa.
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46
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Sun YJ, Chou CC, Chen WS, Wu RT, Meng M, Hsiao CD. The crystal structure of a multifunctional protein: phosphoglucose isomerase/autocrine motility factor/neuroleukin. Proc Natl Acad Sci U S A 1999; 96:5412-7. [PMID: 10318897 PMCID: PMC21873 DOI: 10.1073/pnas.96.10.5412] [Citation(s) in RCA: 93] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/18/2022] Open
Abstract
Phosphoglucose isomerase (PGI) plays a central role in both the glycolysis and the gluconeogenesis pathways. We present here the complete crystal structure of PGI from Bacillus stearothermophilus at 2.3-A resolution. We show that PGI has cell-motility-stimulating activity on mouse colon cancer cells similar to that of endogenous autocrine motility factor (AMF). PGI can also enhance neurite outgrowth on neuronal progenitor cells similar to that observed for neuroleukin. The results confirm that PGI is neuroleukin and AMF. PGI has an open twisted alpha/beta structural motif consisting of two globular domains and two protruding parts. Based on this substrate-free structure, together with the previously published biological, biochemical, and modeling results, we postulate a possible substrate-binding site that is located within the domains' interface for PGI and AMF. In addition, the structure provides evidence suggesting that the top part of the large domain together with one of the protruding loops might participate in inducing the neurotrophic activity.
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Affiliation(s)
- Y J Sun
- Institute of Molecular Biology, Academia Sinica, Taipei, Taiwan 11529, Republic of China
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47
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Benlimame N, Le PU, Nabi IR. Localization of autocrine motility factor receptor to caveolae and clathrin-independent internalization of its ligand to smooth endoplasmic reticulum. Mol Biol Cell 1998; 9:1773-86. [PMID: 9658170 PMCID: PMC25416 DOI: 10.1091/mbc.9.7.1773] [Citation(s) in RCA: 94] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/11/2022] Open
Abstract
Autocrine motility factor receptor (AMF-R) is a cell surface receptor that is also localized to a smooth subdomain of the endoplasmic reticulum, the AMF-R tubule. By postembedding immunoelectron microscopy, AMF-R concentrates within smooth plasmalemmal vesicles or caveolae in both NIH-3T3 fibroblasts and HeLa cells. By confocal microscopy, cell surface AMF-R labeled by the addition of anti-AMF-R antibody to viable cells at 4 degreesC exhibits partial colocalization with caveolin, confirming the localization of cell surface AMF-R to caveolae. Labeling of cell surface AMF-R by either anti-AMF-R antibody or biotinylated AMF (bAMF) exhibits extensive colocalization and after a pulse of 1-2 h at 37 degreesC, bAMF accumulates in densely labeled perinuclear structures as well as fainter tubular structures that colocalize with AMF-R tubules. After a subsequent 2- to 4-h chase, bAMF is localized predominantly to AMF-R tubules. Cytoplasmic acidification, blocking clathrin-mediated endocytosis, results in the essentially exclusive distribution of internalized bAMF to AMF-R tubules. By confocal microscopy, the tubular structures labeled by internalized bAMF show complete colocalization with AMF-R tubules. bAMF internalized in the presence of a 10-fold excess of unlabeled AMF labels perinuclear punctate structures, which are therefore the product of fluid phase endocytosis, but does not label AMF-R tubules, demonstrating that bAMF targeting to AMF-R tubules occurs via a receptor-mediated pathway. By electron microscopy, bAMF internalized for 10 min is located to cell surface caveolae and after 30 min is present within smooth and rough endoplasmic reticulum tubules. AMF-R is therefore internalized via a receptor-mediated clathrin-independent pathway to smooth ER. The steady state localization of AMF-R to caveolae implicates these cell surface invaginations in AMF-R endocytosis.
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Affiliation(s)
- N Benlimame
- Département de Pathologie et Biologie Cellulaire, Université de Montréal, Montréal, Québec, Canada H3C 3J7
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48
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Tímár J, Tóvári J, Szekeres K, Kagawa D, Honn KV. Key determinants of the invasion mechanism of melanoma. Role for a new signaling pathway. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 1997; 407:303-10. [PMID: 9321968 DOI: 10.1007/978-1-4899-1813-0_45] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 02/05/2023]
Affiliation(s)
- J Tímár
- 1st Institute of Pathology & Experimental Cancer Research Semmelweis University of Medicine Budapest, Hungary
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49
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Young MR. Protein phosphatases-1 and -2A regulate tumor cell migration, invasion and cytoskeletal organization. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 1997; 407:311-8. [PMID: 9321969 DOI: 10.1007/978-1-4899-1813-0_46] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 02/05/2023]
Abstract
The role of protein kinase A (PKA) and protein phosphatases (PP) -1 and -2A in regulating the metastatic phenotype of Lewis lung carcinoma (LLC) cells was evaluated. The metastatic LLC-LN7 cells were more motile and invasive than were nonmetastatic LLC-C8 cells. Compared to the nonmetastatic cells, the LLC-LN7 cells had increased PKA activity and a deficiency in PP-2A. Nonmetastatic LLC-C8 cells became migratory and invasive when PP-1 and P-2A activities were inhibited with okadaic acid. This stimulation of LLC-C8 motility was tempered by PKA inhibition. Also examined was if the okadaic acid-stimulated LLC-C8 motility was associated with a change in the cytoskeletal organization to that typical of metastatic cells. Treatment of nonmetastatic LLC-C8 cells with okadaic acid caused a redistribution of F-actin toward the periphery of the cells, and eventually to a loss of the filamentous actin network. All of these effects were reversed upon removal of okadaic acid. Our results show that PP-1/2A maintain reduced motility and increased cytoskeletal organization within nonmetastatic LLC cells.
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Affiliation(s)
- M R Young
- Research Service (151-Z2) Hines V.A. Hospital, IL 60141, USA
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Nabi IR, Guay G, Simard D. AMF-R tubules concentrate in a pericentriolar microtubule domain after MSV transformation of epithelial MDCK cells. J Histochem Cytochem 1997; 45:1351-63. [PMID: 9313797 DOI: 10.1177/002215549704501004] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/05/2023] Open
Abstract
Autocrine motility factor receptor (AMF-R) is localized to an intracellular microtubule-associated membranous organelle, the AMF-R tubule. In well-spread untransformed MDCK epithelial cells, the microtubules originate from a broad perinuclear region and AMF-R tubules extend throughout the cytoplasm of the cells. In Moloney sarcoma virus (mos)-transformed MDCK (MSV-MDCK) cells, microtubules accumulate around the centrosome, forming a microtubule domain rich in stabilized detyrosinated microtubules. AMF-R tubules are quantitatively associated with this pericentriolar microtubule domain and the rough endoplasmic reticulum and lysosomes also co-distribute with the pericentriolar mass of microtubules. The Golgi apparatus is closely associated with the microtubule organizing center (MTOC) within the juxtanuclear mass of AMF-R tubules, and no co-localization of AMF-R tubules with the Golgi marker beta-COP could be detected by confocal microscopy. After nocodazole treatment and washout, microtubule nucleation occurs exclusively at the centrosome of MSV-MDCK cells, and only after microtubule extension to the cell periphery does the microtubule cytoskeleton reorganize to generate the pericentriolar microtubule domain after 30-60 min. AMF-R tubules dispersed by nocodazole treatment concentrate in the pericentriolar region in parallel with the reorganization of the microtubule cytoskeleton. MSV transformation of epithelial MDCK cells results in the stabilization of a pericentriolar microtubule domain responsible for the concentration and polarized distribution of AMF-R tubules.
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Affiliation(s)
- I R Nabi
- Département d'Anatomie, Université de Montréal, Québec, Canada
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