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Sun Y, Wei H, Yu W, Gao H, Li J, Li X, Zhang H, Zhang H, Miao S, Zhao L, Yang R, Xu J, Lu Y, Wei F, Zhou H, Gao D, Jin Y, Zhang L. The actin-binding protein drebrin disrupts NF2-LATS kinases complex assembly to facilitate liver tumorigenesis. Hepatology 2025; 81:1433-1451. [PMID: 39325963 DOI: 10.1097/hep.0000000000001063] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/18/2024] [Accepted: 07/17/2024] [Indexed: 09/28/2024]
Abstract
BACKGROUND AND AIMS The Hippo signaling has emerged as a crucial regulator of tissue homeostasis, regeneration, and tumorigenesis, representing a promising therapeutic target. Neurofibromin 2 (NF2), a component of Hippo signaling, is directly linked to human cancers but has been overlooked as a target for cancer therapy. APPROACH AND RESULTS Through a high-content RNA interference genome-wide screen, the actin-binding protein Drebrin (DBN1) has been identified as a novel modulator of YAP localization. Further investigations have revealed that DBN1 directly interacts with NF2, disrupting the activation of large tumor suppressor kinases (LATS1/2) by competing with LATS kinases for NF2 binding. Consequently, DBN1 knockout considerably promotes YAP nuclear exclusion and repression of target gene expression, thereby preventing cell proliferation and liver tumorigenesis. We identified three lysine residues (K238, K248, and K252) essential for DBN1-NF2 interaction and developed a mutant DBN1 (DBN1-3K mut ) that is defective in NF2 binding and incompetent to trigger NF2-dependent YAP activation and tumorigenesis both in vitro and in vivo. Furthermore, BTP2, a DBN1 inhibitor, successfully restored NF2-LATS kinase binding and elicited potent antitumor activity. The combination of sorafenib and BTP2 exerted synergistic inhibitory effects against HCC. CONCLUSIONS Our study identifies a novel DBN1-NF2-LATS axis, and pharmacological inhibition of DBN1 represents a promising alternative intervention targeting the Hippo pathway in cancer treatment.
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Affiliation(s)
- Yang Sun
- Sheng Yushou Center of Cell Biology and Immunology, Department of Genetics and Developmental Science, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Minghang, Shanghai, China
| | - Henan Wei
- State Key Laboratory of Cell Biology, Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China
| | - Wentao Yu
- State Key Laboratory of Cell Biology, Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China
| | - Haoran Gao
- State Key Laboratory of Cell Biology, Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China
| | - Jinhui Li
- HuidaGene Therapeutics Co., Ltd., Shanghai, China
| | - Xiaoyu Li
- State Key Laboratory of Cell Biology, Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China
| | - Haijiao Zhang
- Sheng Yushou Center of Cell Biology and Immunology, Department of Genetics and Developmental Science, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Minghang, Shanghai, China
| | - Haoen Zhang
- State Key Laboratory of Cell Biology, Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China
| | - Sen Miao
- Department of Pathology, Affiliated Hospital of Jining Medical University, Jining, China
| | - Lihua Zhao
- Department of Pathology, Affiliated Hospital of Jining Medical University, Jining, China
| | - Ruizeng Yang
- Sheng Yushou Center of Cell Biology and Immunology, Department of Genetics and Developmental Science, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Minghang, Shanghai, China
| | - Jinjin Xu
- Sheng Yushou Center of Cell Biology and Immunology, Department of Genetics and Developmental Science, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Minghang, Shanghai, China
| | - Yi Lu
- State Key Laboratory of Cell Biology, Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China
| | - Fang Wei
- Sheng Yushou Center of Cell Biology and Immunology, Department of Genetics and Developmental Science, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Minghang, Shanghai, China
| | - Hu Zhou
- State Key Laboratory of Drug Research, Department of Analytical Chemistry, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, China
- University of Chinese Academy of Sciences, Beijing, China
| | - Daming Gao
- State Key Laboratory of Cell Biology, Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China
| | - Yunyun Jin
- Sheng Yushou Center of Cell Biology and Immunology, Department of Genetics and Developmental Science, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Minghang, Shanghai, China
- Department of Emergency and Critical Care Medicine, Shanghai Pudong New Area People's Hospital, Shanghai, China
| | - Lei Zhang
- Sheng Yushou Center of Cell Biology and Immunology, Department of Genetics and Developmental Science, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Minghang, Shanghai, China
- State Key Laboratory of Cell Biology, Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai, China
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Tang PC, Um S, Mayfield AB, Bracho OR, Castillo CD, Dinh CT, Dykxhoorn DM, Liu XZ. Interactions among Merlin, Arkadia, and SKOR2 mediate NF2-associated human Schwann cell proliferation. Stem Cell Res Ther 2025; 16:163. [PMID: 40188079 PMCID: PMC11972516 DOI: 10.1186/s13287-025-04281-x] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/09/2024] [Accepted: 03/19/2025] [Indexed: 04/07/2025] Open
Abstract
BACKGROUND NF2-related Schwannomatosis (previously referred to as Neurofibromatosis Type 2, or NF2) is a genetic-associated disease resulting from mutations in the gene, NF2. NF2 encodes the Merlin protein, which acts as a tumor suppressor. Bilateral vestibular schwannoma (VS) is a hallmark of NF2. Although the exactly molecular mechanism mediating NF2-driven schwannomatosis is not fully understood, it is known that defective Merlin protein functionality leads to abnormal cell proliferation. METHODS Herein, we utilized a human induced pluripotent stem cell (hiPSC)-based Schwann cell (SC) model to investigate the role of Merlin in human SCs. SCs were derived from hiPSCs carrying a NF2 mutation (c.191 T > C; p. L64P), its isogenic wild-type control cell line, and a NF2 patient-derived hiPSC line. Phenotypes were determined via immunocytochemistry and various bioassays. Different proteins interacting with Merlin in wild-type and NF2 mutation SCs were identified using co-immunoprecipitation followed by mass spectrometry. RESULTS SC derived from NF2L64P hiPSCs showed significantly higher proliferation and abnormal morphology compared to NF2WT SCs. Phenotypes that could be restored by wildtype NF2 overexpression. Interactome profiling of Merlin (NF2) in SCs derived from NF2WT- and NF2L64P- hiPCSs identified differential protein binding levels. Among identified proteins, we validated the interaction among Merlin, an E3 ubiquitin ligase (Arkadia), and a SKI family co-repressor (SKOR2). This complex plays a significant role for this interaction in SC proliferation. Our findings were further validated by SCs derived from the patient-derived hiPSCs carrying a deletion in the chromosome 22 which spans the NF2 gene. CONCLUSIONS Our results presented a hiPSC-derived SC system for SC-related disease modeling and established a new model in which Merlin interacts with Arkadia and SKOR2. This interaction is required for the proper cell proliferation in human SCs.
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Affiliation(s)
- Pei-Ciao Tang
- Department of Otolaryngology, University of Miami Miller School of Medicine, Miami, FL, 33136, USA.
- The Interdisciplinary Stem Cell Institute, University of Miami Miller School of Medicine, Miami, FL, 33136, USA.
| | - Seyoung Um
- Department of Biochemistry and Molecular Biology, University of Miami Miller School of Medicine, Miami, FL, 33136, USA
| | | | - Olena R Bracho
- Department of Otolaryngology, University of Miami Miller School of Medicine, Miami, FL, 33136, USA
| | - Christian Del Castillo
- Department of Otolaryngology, University of Miami Miller School of Medicine, Miami, FL, 33136, USA
| | - Christine T Dinh
- Department of Otolaryngology, University of Miami Miller School of Medicine, Miami, FL, 33136, USA
- Sylvester Comprehensive Cancer Center, Miami, FL, 33136, USA
| | - Derek M Dykxhoorn
- Department of Human Genetics, Dr. John T Macdonald Foundation, University of Miami Miller School of Medicine, Miami, FL, 33136, USA
| | - Xue Zhong Liu
- Department of Otolaryngology, University of Miami Miller School of Medicine, Miami, FL, 33136, USA.
- Department of Human Genetics, Dr. John T Macdonald Foundation, University of Miami Miller School of Medicine, Miami, FL, 33136, USA.
- The Interdisciplinary Stem Cell Institute, University of Miami Miller School of Medicine, Miami, FL, 33136, USA.
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Tang PC, Um S, Mayfield AB, Bracho OR, Castillo CD, Dinh CT, Dykxhoorn DM, Liu XZ. Interactions among Merlin, Arkadia, and SKOR2 mediate NF2-associated Schwann cell proliferation in human. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2024:2024.09.24.614711. [PMID: 39386608 PMCID: PMC11463491 DOI: 10.1101/2024.09.24.614711] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Subscribe] [Scholar Register] [Indexed: 10/12/2024]
Abstract
NF2-Related Schwannomatosis (previously referred to as Neurofibromatosis Type 2, or NF2) is a genetic-associated disease resulting from mutations in the gene, NF2. NF2 encodes the merlin protein, which acts as a tumor suppressor. Bilateral vestibular schwannoma (VS) is a hallmark of NF2. Although the exactly molecular mechanism mediating NF2-driven schwannomatosis remain unclear, it is known that defective Merlin protein functionality leads to abnormal cell proliferation. Herein, we utilized a human induced pluripotent stem cell (hiPSC)-based Schwann cell (SC) model to investigate the role of merlin in human SCs. SCs were derived from hiPSCs carrying a NF2 mutation (c.191 T > C; p. L64P), its isogenic wild-type control cell line, and a NF2 patient-derived hiPSC line. NF2 mutant SCs showed abnormal cellular morphology and proliferation. Proteomic analyses identified novel interaction partners for Merlin - Arkadia and SKOR2. Our results established a new model in which merlin interacts with Arkadia and SKOR2 and this interaction is required for the proper activation of the SMAD-dependent pathway in TGFβ signaling.
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Affiliation(s)
- Pei-Ciao Tang
- Equal contribution: Pei-Ciao Tang and Seyoung Um
- Lead contact: Pei-Ciao Tang
- Department of Otolaryngology, University of Miami Miller School of Medicine, Miami, FL, 33136, United States
| | - Seyoung Um
- Equal contribution: Pei-Ciao Tang and Seyoung Um
- Department of Biochemistry and Molecular Biology, University of Miami Miller School of Medicine, Miami, FL, 33136, United States
| | | | - Olena R. Bracho
- Department of Otolaryngology, University of Miami Miller School of Medicine, Miami, FL, 33136, United States
| | - Christian Del Castillo
- Department of Otolaryngology, University of Miami Miller School of Medicine, Miami, FL, 33136, United States
| | - Christine T. Dinh
- Department of Otolaryngology, University of Miami Miller School of Medicine, Miami, FL, 33136, United States
- Sylvester Comprehensive Cancer Center, Miami, FL, 33136, United States
| | - Derek M. Dykxhoorn
- Dr. John T Macdonald Foundation Department of Human Genetics, University of Miami Miller School of Medicine, Miami, FL, 33136, United States
| | - Xue Zhong Liu
- Department of Otolaryngology, University of Miami Miller School of Medicine, Miami, FL, 33136, United States
- Dr. John T Macdonald Foundation Department of Human Genetics, University of Miami Miller School of Medicine, Miami, FL, 33136, United States
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Nourbakhsh A, Gosstola NC, Fernandez-Valle C, Dykxhoorn DM, Liu XZ. Characterization of UMi031-A-2 inducible pluripotent stem cell line with a neurofibromatosis type 2-associated mutation. Stem Cell Res 2021; 55:102474. [PMID: 34352618 DOI: 10.1016/j.scr.2021.102474] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/05/2021] [Revised: 06/28/2021] [Accepted: 07/16/2021] [Indexed: 11/17/2022] Open
Abstract
The UMi031-A-2 hiPSC line contains a CRISPR-induced homozygous, Neurofibromatosis Type 2 (NF2) mutation (L64P (CTG > CCG)) in the NF2 gene that encodes a merlin tumor suppressor. This line was generated from an unaffected iPSC line using CRISPR technology and characterized for pluripotency and karyotypic stability. The c.191 T > C variant in NF2 is associated with a syndromic nervous system tumor disorder leading to the development of bilateral vestibular schwannomas. Once differentiated into Schwann cells, UMi031-A-2 can serve as a resource for the analysis of signaling pathways deregulated upon merlin defects and provide a pre-clinical platform for testing therapies for NF2 schwannomas.
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Affiliation(s)
- Aida Nourbakhsh
- Department of Otolaryngology, University of Miami Miller, School of Medicine, Miami, FL 33136, USA
| | - Nicholas C Gosstola
- Department of Otolaryngology, University of Miami Miller, School of Medicine, Miami, FL 33136, USA
| | - Cristina Fernandez-Valle
- Burnett School of Biomedical Science, College of Medicine, University of Central Florida, Orlando, FL 32827, USA
| | - Derek M Dykxhoorn
- John P. Hussman Institute for Human Genomics, John T Macdonald Foundation, Department of Human Genetics, University of Miami Miller, School of Medicine, Miami, FL 33136, USA
| | - Xue Zhong Liu
- Department of Otolaryngology, University of Miami Miller, School of Medicine, Miami, FL 33136, USA; John P. Hussman Institute for Human Genomics, John T Macdonald Foundation, Department of Human Genetics, University of Miami Miller, School of Medicine, Miami, FL 33136, USA.
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Khan AB, Gadot R, Shetty A, Bayley JC, Hadley CC, Cardenas MF, Jalali A, Harmanci AS, Harmanci AO, Wheeler DA, Klisch TJ, Patel AJ. Identification of novel fusion transcripts in meningioma. J Neurooncol 2020; 149:219-230. [PMID: 32949309 DOI: 10.1007/s11060-020-03599-1] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/30/2020] [Accepted: 08/08/2020] [Indexed: 12/24/2022]
Abstract
INTRODUCTION Meningiomas are the most common primary intracranial tumor. Recent next generation sequencing analyses have elaborated the molecular drivers of this disease. We aimed to identify and characterize novel fusion genes in meningiomas. METHODS We performed a secondary analysis of our RNA sequencing data of 145 primary meningioma from 140 patients to detect fusion genes. Semi-quantitative rt-PCR was performed to confirm transcription of the fusion genes in the original tumors. Whole exome sequencing was performed to identify copy number variations within each tumor sample. Comparative RNA seq analysis was performed to assess the clonality of the fusion constructs within the tumor. RESULTS We detected six fusion events (NOTCH3-SETBP1, NF2-SPATA13, SLC6A3-AGBL3, PHF19-FOXP2 in two patients, and ITPK1-FBP2) in five out of 145 tumor samples. All but one event (NF2-SPATA13) led to extremely short reading frames, making these events de facto null alleles. Three of the five patients had a history of childhood radiation. Four out of six fusion events were detected in expression type C tumors, which represent the most aggressive meningioma. We validated the presence of the RNA transcripts in the tumor tissue by semi-quantitative RT PCR. All but the two PHF19-FOXP2 fusions demonstrated high degrees of clonality. CONCLUSIONS Fusion genes occur infrequently in meningiomas and are more likely to be found in tumors with greater degree of genomic instability (expression type C) or in patients with history of cranial irradiation.
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Affiliation(s)
- A Basit Khan
- Department of Neurosurgery, Baylor College of Medicine, 7200 Cambridge 9th Floor, Houston, TX, 77030, USA
| | - Ron Gadot
- Department of Neurosurgery, Baylor College of Medicine, 7200 Cambridge 9th Floor, Houston, TX, 77030, USA
| | - Arya Shetty
- Department of Neurosurgery, Baylor College of Medicine, 7200 Cambridge 9th Floor, Houston, TX, 77030, USA
| | - James C Bayley
- Department of Neurosurgery, Baylor College of Medicine, 7200 Cambridge 9th Floor, Houston, TX, 77030, USA
| | - Caroline C Hadley
- Department of Neurosurgery, Baylor College of Medicine, 7200 Cambridge 9th Floor, Houston, TX, 77030, USA
| | - Maria F Cardenas
- Department of Molecular and Human Genetics, Human Genome Sequencing Center, Baylor College of Medicine, Houston, TX, 77030, USA
| | - Ali Jalali
- Department of Neurosurgery, Baylor College of Medicine, 7200 Cambridge 9th Floor, Houston, TX, 77030, USA
| | - Akdes S Harmanci
- School of Biomedical Informatics, Center for Computational Systems Medicine, University of Texas Health Science Center At Houston, Houston, TX, 77030, USA
| | - Arif O Harmanci
- School of Biomedical Informatics, Center for Computational Systems Medicine, University of Texas Health Science Center At Houston, Houston, TX, 77030, USA
| | - David A Wheeler
- Department of Molecular and Human Genetics, Human Genome Sequencing Center, Baylor College of Medicine, Houston, TX, 77030, USA
| | - Tiemo J Klisch
- Texas Children's Hospital, Jan and Dan Duncan Neurological Research Institute, Houston, TX, 77030, USA.
- Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX, 77030, USA.
| | - Akash J Patel
- Department of Neurosurgery, Baylor College of Medicine, 7200 Cambridge 9th Floor, Houston, TX, 77030, USA.
- Texas Children's Hospital, Jan and Dan Duncan Neurological Research Institute, Houston, TX, 77030, USA.
- Department of Otolaryngology-Head and Neck Surgery, Baylor College of Medicine, Houston, TX, USA.
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Alcantara KMM, Garcia RL. MicroRNA‑92a promotes cell proliferation, migration and survival by directly targeting the tumor suppressor gene NF2 in colorectal and lung cancer cells. Oncol Rep 2019; 41:2103-2116. [PMID: 30816526 PMCID: PMC6412542 DOI: 10.3892/or.2019.7020] [Citation(s) in RCA: 21] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/05/2018] [Accepted: 02/05/2019] [Indexed: 01/29/2023] Open
Abstract
Inactivation of the tumor suppressor protein Merlin leads to the development of benign nervous system tumors in neurofibromatosis type 2 (NF2). Documented causes of Merlin inactivation include deleterious mutations in the encoding neurofibromin 2 gene (NF2) and aberrant Merlin phosphorylation leading to proteasomal degradation. Rare somatic NF2 mutations have also been detected in common human malignancies not associated with NF2, including colorectal and lung cancer. Furthermore, tumors without NF2 mutations and with unaltered NF2 transcript levels, but with low Merlin expression, have been reported. The present study demonstrated that NF2 is also regulated by microRNAs (miRNAs) through direct interaction with evolutionarily conserved miRNA response elements (MREs) within its 3′-untranslated region (3′UTR). Dual-Luciferase assays in human colorectal carcinoma (HCT116) and lung adenocarcinoma (A549) cells revealed downregulation of NF2 by miR-92a-3p via its wild-type 3′UTR, but not NF2−3′UTR with mutated miR-92a-3p MRE. HCT116 cells overexpressing miR-92a-3p exhibited significant downregulation of endogenous NF2 mRNA and protein levels, which was rescued by co-transfection of a target protector oligonucleotide specific for the miR-92a-3p binding site within NF2−3′UTR. miR-92a-3p overexpression in HCT116 and A549 cells promoted migration, proliferation and resistance to apoptosis, as well as altered F-actin organization compared with controls. Knockdown of NF2 by siRNA phenocopied the oncogenic effects of miR-92a overexpression on HCT116 and A549 cells. Collectively, the findings of the present study provide functional proof of the unappreciated role of miRNAs in NF2 regulation and tumor progression, leading to enhanced oncogenicity.
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Affiliation(s)
- Krizelle Mae M Alcantara
- National Institute of Molecular Biology and Biotechnology, University of the Philippines, Diliman, Quezon City 1101, Philippines
| | - Reynaldo L Garcia
- National Institute of Molecular Biology and Biotechnology, University of the Philippines, Diliman, Quezon City 1101, Philippines
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Proctor DT, Ramachandran S, Lama S, Sutherland GR. Towards Molecular Classification of Meningioma: Evolving Treatment and Diagnostic Paradigms. World Neurosurg 2018; 119:366-373. [PMID: 30138732 DOI: 10.1016/j.wneu.2018.08.019] [Citation(s) in RCA: 39] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/27/2018] [Revised: 08/01/2018] [Accepted: 08/02/2018] [Indexed: 01/20/2023]
Abstract
Meningioma, a common primary brain tumor in adults, is graded based on World Health Organization criteria that rely on histology alone. This approach is unable to determine conclusively which tumors, especially benign or atypical, will recur. Molecular characterization of meningioma has identified genetic biomarkers that can predict tumor behavior. Only a few genetic changes are known to classify >85% of all meningioma and clinical trials using targeted therapy to genetic subtypes of meningioma are under way. Immunotherapy is also being trialed in treating high-grade and recurrent meningioma. This review summarizes recent developments characterizing meningioma using genetic and immunologic biomarkers and how these molecular tools may be integrated into existing care together with current World Health Organization grading to improve diagnosis, prognosis, and therapy.
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Affiliation(s)
- Dustin T Proctor
- Project neuroArm, Department of Clinical Neurosciences, Cumming School of Medicine, University of Calgary, Calgary, Alberta, Canada; Hotchkiss Brain Institute, University of Calgary, Calgary, Alberta, Canada; Arnie Charbonneau Cancer Institute, University of Calgary, Calgary, Alberta, Canada
| | - Sudheesh Ramachandran
- Project neuroArm, Department of Clinical Neurosciences, Cumming School of Medicine, University of Calgary, Calgary, Alberta, Canada; Hotchkiss Brain Institute, University of Calgary, Calgary, Alberta, Canada; Arnie Charbonneau Cancer Institute, University of Calgary, Calgary, Alberta, Canada
| | - Sanju Lama
- Project neuroArm, Department of Clinical Neurosciences, Cumming School of Medicine, University of Calgary, Calgary, Alberta, Canada; Hotchkiss Brain Institute, University of Calgary, Calgary, Alberta, Canada; Arnie Charbonneau Cancer Institute, University of Calgary, Calgary, Alberta, Canada
| | - Garnette R Sutherland
- Project neuroArm, Department of Clinical Neurosciences, Cumming School of Medicine, University of Calgary, Calgary, Alberta, Canada; Hotchkiss Brain Institute, University of Calgary, Calgary, Alberta, Canada; Arnie Charbonneau Cancer Institute, University of Calgary, Calgary, Alberta, Canada.
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NF2/Merlin Inactivation and Potential Therapeutic Targets in Mesothelioma. Int J Mol Sci 2018; 19:ijms19040988. [PMID: 29587439 PMCID: PMC5979333 DOI: 10.3390/ijms19040988] [Citation(s) in RCA: 70] [Impact Index Per Article: 10.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/28/2018] [Revised: 03/19/2018] [Accepted: 03/19/2018] [Indexed: 12/14/2022] Open
Abstract
The neurofibromatosis type 2 (NF2) gene encodes merlin, a tumor suppressor protein frequently inactivated in schwannoma, meningioma, and malignant mesothelioma (MM). The sequence of merlin is similar to that of ezrin/radixin/moesin (ERM) proteins which crosslink actin with the plasma membrane, suggesting that merlin plays a role in transducing extracellular signals to the actin cytoskeleton. Merlin adopts a distinct closed conformation defined by specific intramolecular interactions and regulates diverse cellular events such as transcription, translation, ubiquitination, and miRNA biosynthesis, many of which are mediated through Hippo and mTOR signaling, which are known to be closely involved in cancer development. MM is a very aggressive tumor associated with asbestos exposure, and genetic alterations in NF2 that abrogate merlin’s functional activity are found in about 40% of MMs, indicating the importance of NF2 inactivation in MM development and progression. In this review, we summarize the current knowledge of molecular events triggered by NF2/merlin inactivation, which lead to the development of mesothelioma and other cancers, and discuss potential therapeutic targets in merlin-deficient mesotheliomas.
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Abstract
Neurofibromatosis (NF) encompasses a group of distinct genetic disorders in which affected children and adults are prone to the development of benign and malignant tumors of the nervous system. The purpose of this review is to discuss the spectrum of CNS tumors arising in individuals with NF type 1 (NF1) and NF type 2 (NF2), their pathogenic etiologies, and the rational treatment options for people with these neoplasms. This article is a review of preclinical and clinical data focused on the treatment of the most common CNS tumors encountered in children and adults with NF1 and NF2. Although children with NF1 are at risk for developing low-grade gliomas of the optic pathway and brainstem, individuals with NF2 typically manifest low-grade tumors affecting the cranial nerves (vestibular schwannomas), meninges (meningiomas), and spinal cord (ependymomas). With the identification of the NF1 and NF2 genes, molecularly targeted therapies are beginning to emerge, as a result of a deeper understanding of the mechanisms underlying NF1 and NF2 protein function. As we enter into an era of precision oncology, a more comprehensive awareness of the factors that increase the risk of developing CNS cancers in affected individuals, coupled with a greater appreciation of the cellular and molecular determinants that maintain tumor growth, will undoubtedly yield more effective therapies for these cancer predisposition syndromes.
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Affiliation(s)
- Jian Campian
- All authors: Washington University School of Medicine, St. Louis, MO
| | - David H Gutmann
- All authors: Washington University School of Medicine, St. Louis, MO
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Merlin inhibits Wnt/β-catenin signaling by blocking LRP6 phosphorylation. Cell Death Differ 2016; 23:1638-47. [PMID: 27285107 DOI: 10.1038/cdd.2016.54] [Citation(s) in RCA: 29] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/01/2015] [Revised: 05/04/2016] [Accepted: 05/10/2016] [Indexed: 12/20/2022] Open
Abstract
Merlin, encoded by the NF2 gene, is a tumor suppressor that acts by inhibiting mitogenic signaling and is mutated in Neurofibromatosis type II (NF2) disease, although its molecular mechanism is not fully understood. Here, we observed that Merlin inhibited Wnt/β-catenin signaling by blocking phosphorylation of LRP6, which is necessary for Wnt signal transduction, whereas mutated Merlin in NF2 patients did not. Treatment with Wnt3a enhanced phosphorylation of Ser518 in Merlin via activation of PAK1 in a PIP2-dependent manner. Phosphorylated Merlin dissociated from LRP6, allowing for phosphorylation of LRP6. Tissues from NF2 patients exhibited higher levels of β-catenin, and proliferation of RT4-D6P2T rat schwannoma cells was significantly reduced by treatment with chemical inhibitors of Wnt/β-catenin signaling. Taken together, our findings suggest that sustained activation of Wnt/β-catenin signaling due to abrogation of Merlin-mediated inhibition of LRP6 phosphorylation may be a cause of NF2 disease.
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Mao B, Gao Y, Bai Y, Yuan Z. Hippo signaling in stress response and homeostasis maintenance. Acta Biochim Biophys Sin (Shanghai) 2015; 47:2-9. [PMID: 25476206 DOI: 10.1093/abbs/gmu109] [Citation(s) in RCA: 38] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/19/2022] Open
Abstract
Co-ordination of cell proliferation, differentiation, and apoptosis maintains tissue development and homeostasis under normal or stress conditions. Recently, the highly conserved Hippo signaling pathway, discovered in Drosophila melanogaster and mammalian system, has been implicated as a key regulator of organ size control. Importantly, emerging evidence suggests that Hippo pathway is involved in the responses to cellular stresses, including mechanic stress, DNA damage, and oxidative stress, to maintain homeostasis at the cellular and organic levels. The mutation or deregulation of the key components in the pathway will result in degenerative disorder, developmental defects, or tumorigenesis. The purpose of this review is to summarize the recent findings and discuss how Hippo pathway responds to cellular stress and regulates early development events, tissue homeostasis as well as tumorigenesis.
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Affiliation(s)
- Beibei Mao
- State Key Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China
| | - Yuhao Gao
- State Key Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China
| | - Yujie Bai
- Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China
| | - Zengqiang Yuan
- State Key Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China
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Park HW, Guan KL. Regulation of the Hippo pathway and implications for anticancer drug development. Trends Pharmacol Sci 2013; 34:581-9. [PMID: 24051213 DOI: 10.1016/j.tips.2013.08.006] [Citation(s) in RCA: 104] [Impact Index Per Article: 8.7] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/18/2013] [Revised: 08/12/2013] [Accepted: 08/20/2013] [Indexed: 12/22/2022]
Abstract
Research in the past decade has revealed key components of the Hippo tumor suppressor pathway and its critical role in organ size regulation and tumorigenesis. Recent progress has identified a wide range of upstream factors that control the Hippo pathway, which include cell-cell contact, various diffusible signals, and cognate receptors. Dysregulation of the Hippo pathway, caused by gene mutation or aberrant expression, promotes cell proliferation and tumorigenesis. Here, we discuss the current state of Hippo pathway research, primarily focusing on upstream regulators and protein-protein interactions as potential therapeutic targets. Consideration of pharmacological intervention of the Hippo pathway may provide novel avenues for future therapeutic treatment of human diseases, particularly in cancer.
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Affiliation(s)
- Hyun Woo Park
- Department of Pharmacology and Moores Cancer Center, University of California San Diego, La Jolla, CA 92093, USA
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14
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The characterisation of Pax3 expressant cells in adult peripheral nerve. PLoS One 2013; 8:e59184. [PMID: 23527126 PMCID: PMC3602598 DOI: 10.1371/journal.pone.0059184] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/29/2012] [Accepted: 02/12/2013] [Indexed: 12/25/2022] Open
Abstract
Pax3 has numerous integral functions in embryonic tissue morphogenesis and knowledge of its complex function in cells of adult tissue continues to unfold. Across a variety of adult tissue lineages, the role of Pax3 is principally linked to maintenance of the tissue’s resident stem/progenitor cell population. In adult peripheral nerves, Pax3 is reported to be expressed in nonmyelinating Schwann cells, however, little is known about the purpose of this expression. Based on the evidence of the role of Pax3 in other adult tissue stem and progenitor cells, it was hypothesised that the cells in adult peripheral nerve that express Pax3 may be peripheral glioblasts. Here, methods have been developed for identification and visualisation of Pax3 expressant cells in normal 60 day old mouse peripheral nerve that allowed morphological and phenotypic distinctions to be made between Pax3 expressing cells and other nonmyelinating Schwann cells. The distinctions described provide compelling support for a resident glioblast population in adult mouse peripheral nerve.
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15
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Laulajainen M, Melikova M, Muranen T, Carpén O, Grönholm M. Distinct overlapping sequences at the carboxy-terminus of merlin regulate its tumour suppressor and morphogenic activity. J Cell Mol Med 2013; 16:2161-75. [PMID: 22325036 PMCID: PMC3822986 DOI: 10.1111/j.1582-4934.2012.01525.x] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/14/2023] Open
Abstract
The Neurofibromatosis 2 (NF2) gene product merlin is a tumour suppressor, which in addition to inhibiting cell proliferation regulates cell morphology. The morphogenic properties of merlin may play a role in tumour suppression, as patient-derived tumour cells demonstrate cytoskeletal abnormalities. However, it is still unclear how these functions are linked. The N-terminal FERM-domain of merlin is highly homologous to the oncogenic protein ezrin, while the C-termini are less conserved, suggesting that the opposite effect of the proteins on proliferation could be mediated by their distinct C-terminal regions. In this study we characterize the role of the most C-terminal residues of merlin in the regulation of proliferation, cytoskeletal organization, phosphorylation and intramolecular associations. In addition to the two full-length merlin isoforms and truncating mutations found in patients, we focused on the evolutionally conserved C-terminal residues 545-547, also harbouring disease-causing mutations. We demonstrate that merlin induces cell extensions, which result from impaired retraction of protrusions rather than from increased formation of filopodia. The residues 538-568 were found particularly important for this morphogenic activity. The results further show that both merlin isoforms are able to equally inhibit proliferation, whereas C-terminal mutants affecting residues 545-547 are less effective in growth suppression. This study demonstrates that the C-terminus contains distinct but overlapping functional domains important for regulation of the morphogenic activity, intramolecular associations and cell proliferation.
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Affiliation(s)
- Minja Laulajainen
- Biomedicum Helsinki, Department of Pathology, University of Helsinki, Helsinki, Finland.
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16
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Cooper J, Li W, You L, Schiavon G, Pepe-Caprio A, Zhou L, Ishii R, Giovannini M, Hanemann CO, Long SB, Erdjument-Bromage H, Zhou P, Tempst P, Giancotti FG. Merlin/NF2 functions upstream of the nuclear E3 ubiquitin ligase CRL4DCAF1 to suppress oncogenic gene expression. Sci Signal 2011; 4:pt6. [PMID: 21878678 DOI: 10.1126/scisignal.2002314] [Citation(s) in RCA: 40] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/02/2022]
Abstract
Integrin-mediated activation of PAK (p21-activated kinase) causes phosphorylation and inactivation of the FERM (4.1, ezrin, radixin, moesin) domain-containing protein Merlin, which is encoded by the NF2 (neurofibromatosis type 2) tumor suppressor gene. Conversely, cadherin engagement inactivates PAK, thus leading to accumulation of unphosphorylated Merlin. Current models imply that Merlin inhibits cell proliferation by inhibiting mitogenic signaling at or near the plasma membrane. We have recently shown that the unphosphorylated, growth-inhibiting form of Merlin accumulates in the nucleus and binds to the E3 ubiquitin ligase CRL4(DCAF1) to suppress its activity. Depletion of DCAF1 blocks the hyperproliferation caused by inactivation of Merlin. Conversely, expression of a Merlin-insensitive DCAF1 mutant counteracts the antimitogenic effect of Merlin. Expression of Merlin or silencing of DCAF1 in Nf2-deficient cells induce an overlapping, tumor-suppressive program of gene expression. Mutations present in some tumors from NF2 patients disrupt Merlin's ability to interact with or inhibit CRL4(DCAF1). Lastly, depletion of DCAF1 inhibits the hyperproliferation of Schwannoma cells isolated from NF2 patients and suppresses the oncogenic potential of Merlin-deficient tumor cell lines. Current studies are aimed at identifying the substrates and mechanism of action of CRL4(DCAF1) and examining its role in NF2-dependent tumorigenesis in mouse models. We propose that Merlin mediates contact inhibition and suppresses tumorigenesis by translocating to the nucleus to inhibit CRL4(DCAF1).
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Affiliation(s)
- Jonathan Cooper
- Memorial Sloan-Kettering Cancer Center, New York, NY 10065, USA
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17
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Choy W, Kim W, Nagasawa D, Stramotas S, Yew A, Gopen Q, Parsa AT, Yang I. The molecular genetics and tumor pathogenesis of meningiomas and the future directions of meningioma treatments. Neurosurg Focus 2011; 30:E6. [DOI: 10.3171/2011.2.focus1116] [Citation(s) in RCA: 124] [Impact Index Per Article: 8.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/22/2023]
Abstract
Meningiomas are mostly benign, slow-growing tumors of the CNS that originate from arachnoidal cap cells. While monosomy 22 is the most frequent genetic abnormality found in meningiomas, a multitude of other aberrant chromosomal alterations, signaling pathways, and growth factors have been implicated in its pathogenesis. Losses on 22q12.2, a region encoding the tumor suppressor gene merlin, represent the most common genetic alterations in early meningioma formation. Malignant meningioma progression, however, is associated with more complex karyotypes and greater genetic instability. Cytogenetic studies of atypical and anaplastic meningiomas revealed gains and losses on chromosomes 9, 10, 14, and 18, with amplifications on chromosome 17. However, the specific gene targets in a majority of these chromosomal abnormalities remain elusive.
Studies have also implicated a myriad of aberrant signaling pathways involved with meningioma tumorigenesis, including those involved with proliferation, angiogenesis, and autocrine loops. Understanding these disrupted pathways will aid in deciphering the relationship between various genetic changes and their downstream effects on meningioma pathogenesis.
Despite advancements in our understanding of meningioma pathogenesis, the conventional treatments, including surgery, radiotherapy, and stereotactic radiosurgery, have remained largely stagnant. Surgery and radiation therapy are curative in the majority of lesions, yet treatment remains challenging for meningiomas that are recurrent, aggressive, or refractory to conventional treatments. Future therapies will include combinations of targeted molecular agents as a result of continued progress in the understanding of genetic and biological changes associated with meningiomas.
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Affiliation(s)
| | - Won Kim
- 1Department of Neurological Surgery, and
| | | | | | - Andrew Yew
- 1Department of Neurological Surgery, and
| | - Quinton Gopen
- 2Division of Otolaryngology, University of California Los Angeles; and
| | - Andrew T. Parsa
- 3Department of Neurological Surgery, University of California, San Francisco, California
| | - Isaac Yang
- 1Department of Neurological Surgery, and
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18
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Sosa-García B, Gunduz V, Vázquez-Rivera V, Cress WD, Wright G, Bian H, Hinds PW, Santiago-Cardona PG. A role for the retinoblastoma protein as a regulator of mouse osteoblast cell adhesion: implications for osteogenesis and osteosarcoma formation. PLoS One 2010; 5:e13954. [PMID: 21085651 PMCID: PMC2978706 DOI: 10.1371/journal.pone.0013954] [Citation(s) in RCA: 28] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/14/2010] [Accepted: 10/11/2010] [Indexed: 11/18/2022] Open
Abstract
The retinoblastoma protein (pRb) is a cell cycle regulator inactivated in most human cancers. Loss of pRb function results from mutations in the gene coding for pRb or for any of its upstream regulators. Although pRb is predominantly known as a cell cycle repressor, our data point to additional pRb functions in cell adhesion. Our data show that pRb regulates the expression of a wide repertoire of cell adhesion genes and regulates the assembly of the adherens junctions required for cell adhesion. We conducted our studies in osteoblasts, which depend on both pRb and on cell-to-cell contacts for their differentiation and function. We generated knockout mice in which the RB gene was excised specifically in osteoblasts using the cre-lox P system and found that osteoblasts from pRb knockout mice did not assemble adherens junction at their membranes. pRb depletion in wild type osteoblasts using RNAi also disrupted adherens junctions. Microarrays comparing pRb-expressing and pRb-deficient osteoblasts showed that pRb controls the expression of a number of cell adhesion genes, including cadherins. Furthermore, pRb knockout mice showed bone abnormalities consistent with osteoblast adhesion defects. We also found that pRb controls the function of merlin, a well-known regulator of adherens junction assembly, by repressing Rac1 and its effector Pak1. Using qRT-PCR, immunoblots, co-immunoprecipitation assays, and immunofluorescent labeling, we observed that pRb loss resulted in Rac1 and Pak1 overexpression concomitant with merlin inactivation by Pak1, merlin detachment from the membrane, and adherens junction loss. Our data support a pRb function in cell adhesion while elucidating the mechanism for this function. Our work suggests that in some tumor types pRb inactivation results in both a loss of cell cycle control that promotes initial tumor growth as well as in a loss of cell-to-cell contacts, which contributes to later stages of metastasis.
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Affiliation(s)
| | - Volkan Gunduz
- Molecular Oncology Research Institute, Tufts Medical Center, Boston, Massachusetts, United States of America
| | | | - W. Douglas Cress
- Molecular Oncology and Thoracic Oncology Departments, H. Lee Moffitt Cancer Center and Research Institute, Tampa, Florida, United States of America
| | - Gabriela Wright
- Molecular Oncology and Thoracic Oncology Departments, H. Lee Moffitt Cancer Center and Research Institute, Tampa, Florida, United States of America
| | - Haikuo Bian
- Molecular Oncology and Thoracic Oncology Departments, H. Lee Moffitt Cancer Center and Research Institute, Tampa, Florida, United States of America
| | - Philip W. Hinds
- Molecular Oncology Research Institute, Tufts Medical Center, Boston, Massachusetts, United States of America
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19
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Microtubule-mediated transport of the tumor-suppressor protein Merlin and its mutants. Proc Natl Acad Sci U S A 2010; 107:7311-6. [PMID: 20368450 DOI: 10.1073/pnas.0907389107] [Citation(s) in RCA: 33] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/18/2022] Open
Abstract
The neurofibromatosis type 2 (NF2) tumor-suppressor protein Merlin is a member of the ERM family of proteins that links the cytoskeleton to the plasma membrane. In humans, mutations in the NF2 gene cause neurofibromatosis type-2 (NF2), a cancer syndrome characterized by the development of tumors of the nervous system. Previous reports have suggested that the subcellular distribution of Merlin is critical to its function, and that several NF2 mutants that lack tumor-suppressor activity present improper localization. Here we used a Drosophila cell culture model to study the distribution and mechanism of intracellular transport of Merlin and its mutants. We found that Drosophila Merlin formed cytoplasmic particles that move bidirectionally along microtubules. A single NF2-causing amino acid substitution in the FERM domain dramatically inhibited Merlin particle movement. Surprisingly, the presence of this immotile Merlin mutant also inhibited trafficking of the WT protein. Analysis of the movement of WT protein using RNAi and pull-downs showed that Merlin particles are associated with and moved by microtubule motors (kinesin-1 and cytoplasmic dynein), and that binding of motors and movement is regulated by Merlin phosphorylation. Inhibition of Merlin transport by expression of the dominant-negative mutant or depletion of kinesin-1 results in increased nuclear accumulation of the transcriptional coactivator Yorkie. These results demonstrate the requirement of microtubule-dependent transport for Merlin function.
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20
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Li W, You L, Cooper J, Schiavon G, Pepe-Caprio A, Zhou L, Ishii R, Giovannini M, Hanemann CO, Long SB, Erdjument-Bromage H, Zhou P, Tempst P, Giancotti FG. Merlin/NF2 suppresses tumorigenesis by inhibiting the E3 ubiquitin ligase CRL4(DCAF1) in the nucleus. Cell 2010; 140:477-90. [PMID: 20178741 DOI: 10.1016/j.cell.2010.01.029] [Citation(s) in RCA: 262] [Impact Index Per Article: 17.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/22/2009] [Revised: 08/07/2009] [Accepted: 01/16/2010] [Indexed: 11/29/2022]
Abstract
Current models imply that the FERM domain protein Merlin, encoded by the tumor suppressor NF2, inhibits mitogenic signaling at or near the plasma membrane. Here, we show that the closed, growth-inhibitory form of Merlin accumulates in the nucleus, binds to the E3 ubiquitin ligase CRL4(DCAF1), and suppresses its activity. Depletion of DCAF1 blocks the promitogenic effect of inactivation of Merlin. Conversely, enforced expression of a Merlin-insensitive mutant of DCAF1 counteracts the antimitogenic effect of Merlin. Re-expression of Merlin and silencing of DCAF1 implement a similar, tumor-suppressive program of gene expression. Tumor-derived mutations invariably disrupt Merlin's ability to interact with or inhibit CRL4(DCAF1). Finally, depletion of DCAF1 inhibits the hyperproliferation of Schwannoma cells from NF2 patients and suppresses the oncogenic potential of Merlin-deficient tumor cell lines. We propose that Merlin suppresses tumorigenesis by translocating to the nucleus to inhibit CRL4(DCAF1).
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Affiliation(s)
- Wei Li
- Memorial Sloan-Kettering Cancer Center, New York, NY 10065, USA
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21
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Sughrue ME, Yeung AH, Rutkowski MJ, Cheung SW, Parsa AT. Molecular biology of familial and sporadic vestibular schwannomas: implications for novel therapeutics. J Neurosurg 2009; 114:359-66. [PMID: 19943731 DOI: 10.3171/2009.10.jns091135] [Citation(s) in RCA: 34] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/17/2022]
Abstract
Vestibular schwannomas (VSs) are benign tumors arising from the sheath of cranial nerve VIII. The pathogenesis underlying most familial and sporadic VSs has been linked to a mutation in a single gene, the neurofibromin 2 (NF2) gene located on chromosome 22, band q11-13.1. In this review, the authors summarized what is known about the epidemiology of NF2 mutations and patients with VSs. The authors also discuss the function of the NF2 gene product, merlin, and describe the known and hypothetical effects of genetic mutations that lead to merlin dysfunction on a broad variety of cellular and histological end points. A better understanding of the molecular pathobiology of VSs may lead to novel therapeutics to augment current modalities of treatment while minimizing morbidity.
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Affiliation(s)
- Michael E Sughrue
- Department of Otolaryngology-Head and Neck Surgery, University of California at San Francisco, 94143-0350, USA
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22
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Sauvageot CM, Kesari S, Stiles CD. Molecular pathogenesis of adult brain tumors and the role of stem cells. Neurol Clin 2008; 25:891-924, vii. [PMID: 17964020 DOI: 10.1016/j.ncl.2007.07.014] [Citation(s) in RCA: 28] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2022]
Abstract
Primary brain tumors consist of neoplasms with varied molecular defects, morphologic phenotypes, and clinical outcomes. The genetic and signaling abnormalities involved in tumor initiation and progression of the most prevalent adult primary brain tumors, including gliomas, meningiomas, and medulloblastomas, are described in this article. The current understanding of the cell-of-origin of these neoplasms is reviewed, which suggests that the malignant phenotype is propelled by cells with stem-like qualities. A comprehensive understanding of the molecular basis of transformation and the cell-of-origin of these neoplasms will enable the formulation of more targeted treatment alternatives that could improve survival and quality of life.
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Affiliation(s)
- Claire M Sauvageot
- Department of Cancer Biology, Dana-Farber Cancer Institute, 44 Binney Street, Boston, MA 02115-6084, USA
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23
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Ahronowitz I, Xin W, Kiely R, Sims K, MacCollin M, Nunes FP. Mutational spectrum of the NF2 gene: a meta-analysis of 12 years of research and diagnostic laboratory findings. Hum Mutat 2007; 28:1-12. [PMID: 16983642 DOI: 10.1002/humu.20393] [Citation(s) in RCA: 68] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/07/2022]
Abstract
The NF2 tumor suppressor gene on chromosome 22 is a member of the protein 4.1 family of cytoskeletal elements. A number of single- and multiple-tumor phenotypes have been linked to alterations of NF2 since its characterization in 1993. We present a meta-analysis of 967 constitutional and somatic NF2 alterations from 93 published reports, along with 59 additional unpublished events identified in our laboratory and 115 alterations identified in clinical samples submitted to the Massachusetts General Hospital (MGH) Neurogenetics DNA Diagnostic Laboratory. In total, these sources defined 1,070 small genetic changes detected primarily by exon scanning, 42 intragenic changes of one whole exon or larger, and 29 whole gene deletions and gross chromosomal rearrangements. Constitutional single-exon events (N=422) were significantly more likely to be nonsense or splice site changes than somatic events (N=533), which favored frameshift changes (chi(2) test; P<0.001). Somatic events also differed markedly between tumors of different pathology, most significantly in the tendency of somatic events in meningiomas to lie within the 5' FERM domain of the transcript (Fisher's exact test; P<0.01 in comparison to schwannomas) with a complete absence of mutations in exons 14 and 15. There was no statistically significant difference in mutation type or exon distribution between published constitutional events and those found by the clinical laboratory. Less than 10% of all published and unpublished small alterations are nontruncating (N=63) and these changes are clustered in exons 2 and 3, suggesting that this region may be especially crucial to tumor suppressor activity in the protein.
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Affiliation(s)
- Iris Ahronowitz
- Molecular Neurofibromatosis Laboratory, Massachusetts General Hospital, Charlestown, Massachusetts 02129, USA
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24
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Neff BA, Welling DB, Akhmametyeva E, Chang LS. The molecular biology of vestibular schwannomas: dissecting the pathogenic process at the molecular level. Otol Neurotol 2007; 27:197-208. [PMID: 16436990 DOI: 10.1097/01.mao.0000180484.24242.54] [Citation(s) in RCA: 53] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/27/2022]
Abstract
OBJECTIVE The goal of this article was to review concisely what is currently known about the tumorigenesis of vestibular schwannomas. BACKGROUND Recent advances in molecular biology have led to a better understanding of the cause of vestibular schwannomas. Mutations in the neurofibromatosis type 2 tumor suppressor gene (NF2) have been identified in these tumors. In addition, the interactions of merlin, the protein product of the NF2 gene, and other cellular proteins are beginning to give us a better idea of NF2 function and the pathogenesis of vestibular schwannomas. METHODS Review of the relevant basic science studies at our institution as well as the basic science and clinical literature. RESULTS The clinical characteristics of vestibular schwannomas and neurofibromatosis type 2 syndromes are reviewed and related to alterations in the NF2 gene. Studies demonstrating our current understanding of tumor developmental pathways are highlighted. In addition, methods of clinical and genetic screening for neurofibromatosis type 2 disease are outlined. Avenues for the development of potential future research and therapies are discussed. CONCLUSION Great strides have been made to identify why vestibular schwannomas develop at the molecular level. Continued research is needed to find targeted therapies with which to treat these tumors.
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Affiliation(s)
- Brian A Neff
- Department of Otolaryngology, The Ohio State University College of Medicine and Children's Hospital, Columbus, Ohio, USA
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25
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Baser ME, Kuramoto L, Woods R, Joe H, Friedman JM, Wallace AJ, Ramsden RT, Olschwang S, Bijlsma E, Kalamarides M, Papi L, Kato R, Carroll J, Lázaro C, Joncourt F, Parry DM, Rouleau GA, Evans DGR. The location of constitutional neurofibromatosis 2 (NF2) splice site mutations is associated with the severity of NF2. J Med Genet 2006; 42:540-6. [PMID: 15994874 PMCID: PMC1736092 DOI: 10.1136/jmg.2004.029504] [Citation(s) in RCA: 69] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/04/2022]
Abstract
Neurofibromatosis 2 (NF2) patients with constitutional splice site NF2 mutations have greater variability in disease severity than NF2 patients with other types of mutations; the cause of this variability is unknown. We evaluated genotype-phenotype correlations, with particular focus on the location of splice site mutations, using mutation and clinical information on 831 patients from 528 NF2 families with identified constitutional NF2 mutations. The clinical characteristics examined were age at onset of symptoms of NF2 and number of intracranial meningiomas, which are the primary indices of the severity of NF2. Two regression models were used to analyse genotype-phenotype correlations. People with splice site mutations in exons 1-5 had more severe disease than those with splice site mutations in exons 11-15. This result is compatible with studies showing that exons 2 and 3 are required for self-association of the amino terminal of the NF2 protein in vitro, and that deletions of exons 2 and 3 in transgenic and knockout mouse models of NF2 cause a high prevalence of Schwann cell derived tumours.
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26
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Golovnina K, Blinov A, Akhmametyeva EM, Omelyanchuk LV, Chang LS. Evolution and origin of merlin, the product of the Neurofibromatosis type 2 (NF2) tumor-suppressor gene. BMC Evol Biol 2005; 5:69. [PMID: 16324214 PMCID: PMC1315344 DOI: 10.1186/1471-2148-5-69] [Citation(s) in RCA: 30] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/18/2005] [Accepted: 12/02/2005] [Indexed: 11/10/2022] Open
Abstract
Background Merlin, the product of the Neurofibromatosis type 2 (NF2) tumor suppressor gene, belongs to the ezrin-radixin-moesin (ERM) subgroup of the protein 4.1 superfamily, which links cell surface glycoproteins to the actin cytoskeleton. While merlin's functional activity has been examined in mammalian and Drosophila models, little is understood about its evolution, diversity, and overall distribution among different taxa. Results By combining bioinformatic and phylogenetic approaches, we demonstrate that merlin homologs are present across a wide range of metazoan lineages. While the phylogenetic tree shows a monophyletic origin of the ERM family, the origin of the merlin proteins is robustly separated from that of the ERM proteins. The derivation of merlin is thought to be in early metazoa. We have also observed the expansion of the ERM-like proteins within the vertebrate clade, which occurred after its separation from Urochordata (Ciona intestinalis). Amino acid sequence alignment reveals the absence of an actin-binding site in the C-terminal region of all merlin proteins from various species but the presence of a conserved internal binding site in the N-terminal domain of the merlin and ERM proteins. In addition, a more conserved pattern of amino acid residues is found in the region containing the so-called "Blue Box," although some amino acid substitutions in this region exist in the merlin sequences of worms, fish, and Ciona. Examination of sequence variability at functionally significant sites, including the serine-518 residue, the phosphorylation of which modulates merlin's intra-molecular association and function as a tumor suppressor, identifies several potentially important sites that are conserved among all merlin proteins but divergent in the ERM proteins. Secondary structure prediction reveals the presence of a conserved α-helical domain in the central to C-terminal region of the merlin proteins of various species. The conserved residues and structures identified correspond to the important sites highlighted by the available crystal structures of the merlin and ERM proteins. Furthermore, analysis of the merlin gene structures from various organisms reveals the increase of gene length during evolution due to the expansion of introns; however, a reduction of intron number and length appears to occur in the merlin gene of the insect group. Conclusion Our results demonstrate a monophyletic origin of the merlin proteins with their root in the early metazoa. The overall similarity among the primary and secondary structures of all merlin proteins and the conservation of several functionally important residues suggest a universal role for merlin in a wide range of metazoa.
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Affiliation(s)
- Kseniya Golovnina
- Institute of Cytology and Genetics, Russian Academy of Sciences, 10 Lavrent'ev Ave., 630090, Novosibirsk, Russia
| | - Alexander Blinov
- Institute of Cytology and Genetics, Russian Academy of Sciences, 10 Lavrent'ev Ave., 630090, Novosibirsk, Russia
| | - Elena M Akhmametyeva
- Center for Childhood Cancer, Children's Research Institute, Children's Hospital and Department of Pediatrics, The Ohio State University, 700 Children's Drive, Columbus, OH 43205-2696, USA
| | - Leonid V Omelyanchuk
- Institute of Cytology and Genetics, Russian Academy of Sciences, 10 Lavrent'ev Ave., 630090, Novosibirsk, Russia
| | - Long-Sheng Chang
- Center for Childhood Cancer, Children's Research Institute, Children's Hospital and Department of Pediatrics, The Ohio State University, 700 Children's Drive, Columbus, OH 43205-2696, USA
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27
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Oh MK, Scoles DR, Pulst SM. DNA microarray analysis of immediate response to EGF treatment in rat schwannoma cells. BIOTECHNOL BIOPROC E 2005. [DOI: 10.1007/bf02989827] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/01/2022]
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Abstract
Meningiomas are common central nervous system tumors that originate from the meningeal coverings of the brain and the spinal cord. Most meningiomas are slowly growing benign tumors that histologically correspond to World Health Organization (WHO) grade I. However, certain rare histological variants (clear cell, chordoid, papillary, and rhabdoid), as well as atypical (WHO grade II) and anaplastic (WHO grade III) meningiomas show a more aggressive biological behavior and are clinically associated with a high risk of local recurrence and a less favorable prognosis. This review summarizes the most important features of meningioma pathology and provides an up-to-date overview about the molecular mechanisms involved in meningioma initiation and progression. Current data indicate that meningioma initiation is closely linked to the inactivation of one or more members of the highly conserved protein 4.1 superfamily, including the neurofibromatosis type 2 gene product merlin/schwannomin, protein 4.IB (DAL-1) and protein 4.1R. The genetic alterations in atypical meningiomas are complex and involve losses on 1p, 6q, 10, 14q and 18q, as well as gains on multiple chromosomes. The relevant genes are still unknown. Anaplastic meningiomas show even more complex genetic alterations, including frequent alteration of the CDKN2A, p14ARF, and CDKN2B tumor suppressor genes at 9p21, as well as gene amplification on 17q23. A better understanding of the molecular mechanisms involved in meningioma pathogenesis may not only lead to the identification of novel diagnostic and prognostic marker but will also facilitate the development of new pathogenesis-based therapeutic strategies.
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Affiliation(s)
- Arie Perry
- Division of Neuropathology, Washington University School of Medicine, St Louis, MO 63110-1093, USA.
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Rong R, Surace EI, Haipek CA, Gutmann DH, Ye K. Serine 518 phosphorylation modulates merlin intramolecular association and binding to critical effectors important for NF2 growth suppression. Oncogene 2004; 23:8447-54. [PMID: 15378014 DOI: 10.1038/sj.onc.1207794] [Citation(s) in RCA: 96] [Impact Index Per Article: 4.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/08/2022]
Abstract
The neurofibromatosis 2 (NF2) tumor suppressor protein, merlin, functions as a negative growth regulator; however, the molecular mechanisms that underlie merlin regulation remain elusive. Recent studies have implicated merlin phosphorylation in regulating merlin subcellular localization and growth suppression. P21-activated kinase (PAK), a downstream target of Rac1/Cdc42, directly phosphorylates merlin at Serine 518. In this report, we show that PAK2 directly phosphorylates wild-type merlin, whereas merlin truncation mutants with impaired GST-amino-terminal domain (N-term or NTD)/GST-carboxy-terminal domain (C-term or CTD) intramolecular association exhibit impaired S518 phosphorylation. We directly demonstrate that PAK2 phosphorylation impairs merlin N-term/C-term binding in vitro and in vivo. Lastly, we show that PAK2 phosphorylation impairs the ability of merlin to bind to two interacting proteins, CD44 and hepatocyte growth factor-regulated tyrosine kinase substrate (HRS), both critical for merlin growth suppression. These observations suggest that merlin S518 phosphorylation directly modulates merlin intramolecular and intermolecular associations important for the ability of merlin to function as a tumor suppressor.
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Affiliation(s)
- Rong Rong
- Department of Pathology and Laboratory Medicine, Emory University School of Medicine, 615 Michael Street, Atlanta, GA 30322, USA
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Surace EI, Haipek CA, Gutmann DH. Effect of merlin phosphorylation on neurofibromatosis 2 (NF2) gene function. Oncogene 2004; 23:580-7. [PMID: 14724586 DOI: 10.1038/sj.onc.1207142] [Citation(s) in RCA: 82] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/09/2022]
Abstract
The neurofibromatosis 2 (NF2) tumor suppressor gene product, merlin, belongs to the ezrin-radixin-moesin (ERM) subgroup of the Protein 4.1 family, which links cell surface glycoproteins to the actin cytoskeleton. Previous studies have suggested that phosphorylation of merlin, similar to other ERM proteins, may regulate its function. To determine whether merlin phosphorylation has functional consequences for merlin suppression of cell growth and motility, we generated doxycycline-regulatable RT4 schwannoma cell lines that inducibly express full-length merlin with mutations at two potential phosphorylation sites (amino-acid residues S518 and T576). Whereas a mutation at S518 that mimics constitutive phosphorylation (S518D) abrogates the ability of merlin to suppress cell growth and motility, the S518A merlin mutant, which mimics nonphosphorylated merlin, functions equivalently to wild-type merlin. Similar mutations involving T576, the analogous phosphorylation site in ERM proteins important for regulating their function, had no effect. In contrast to other functionally inactive missense merlin mutants, the regulated overexpression of S518D merlin resulted in dramatic changes in cell shape and the elaboration of filopodial extensions. These results provide the first direct demonstration that the S518D merlin mutation, which mimics merlin phosphorylation, impairs not only merlin growth and motility suppression but also leads to an acquisition of a novel phenotype previously ascribed to ERM proteins.
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Affiliation(s)
- Ezequiel I Surace
- Department of Neurology, Washington University School of Medicine, Box 8111, 660 S Euclid Avenue, St Louis, MO 63110, USA
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Oh MK, Scoles DR, Haipek C, Strand AD, Gutmann DH, Olson JM, Pulst SM. Genetic heterogeneity of stably transfected cell lines revealed by expression profiling with oligonucleotide microarrays. J Cell Biochem 2003; 90:1068-78. [PMID: 14624465 DOI: 10.1002/jcb.10712] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/11/2022]
Abstract
Large-scale gene expression measurements with oligonucleotide microarrays have contributed tremendously to biological research. However, to distinguish between relevant expression changes and falsely identified positives, the source and magnitude of errors must be understood. Here, we report a source of biological variability in microarray experiments with stably transfected cell lines. Mouse embryonic fibroblast (MEF/3T3) and rat schwannoma (RT4) cell lines were generated to provide regulatable schwannomin expression. The expression levels of 29 samples from five different mouse embryonic fibroblast clonal cell lines and 18 samples from 3 RT4 cell lines were monitored with oligonucleotide microarrays. Using hierarchical clustering, we determined that the changes in gene expression induced by schwannomin overexpression were subtle when compared with those detected as a consequence of clonal selection during generation of the cell lines. The hierarchical clustering implies that significant alterations of gene expression were introduced during the transfection and selection processes. A total of 28 genes were identified by Kruskal-Wallis rank test that showed significant variation between clonal lines. Most of them were related to cytoskeletal function and signaling pathways. Based on these analyses, we recommend that replications of experiments with several selected cell lines are necessary to assess biological effects of induced gene expression.
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Affiliation(s)
- Min-Kyu Oh
- Rose Moss Neurogenetics Laboratory for Parkinson and Related Diseases, CSMS Burns and Allen Research Institute, Division of Neurology, Cedars-Sinai Medical Center, 8700 Beverly Boulevard, Los Angeles, California 90048, USA
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Sun CX, Robb VA, Gutmann DH. Protein 4.1 tumor suppressors: getting a FERM grip on growth regulation. J Cell Sci 2002; 115:3991-4000. [PMID: 12356905 DOI: 10.1242/jcs.00094] [Citation(s) in RCA: 131] [Impact Index Per Article: 5.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/30/2022] Open
Abstract
Members of the Protein 4.1 superfamily have highly conserved FERM domains that link cell surface glycoproteins to the actin cytoskeleton. Within this large and constantly expanding superfamily, at least five subgroups have been proposed. Two of these subgroups, the ERM and prototypic Protein 4.1 molecules, include proteins that function as tumor suppressors. The ERM subgroup member merlin/schwannomin is inactivated in the tumor-predisposition syndrome neurofibromatosis 2 (NF2), and the prototypic 4.1 subgroup member, Protein 4.1B, has been implicated in the molecular pathogenesis of breast, lung and brain cancers. This review focuses on what is known of mechanisms of action and critical protein interactions that may mediate the unique growth inhibitory signals of these two Protein 4.1 tumor suppressors. On the basis of insights derived from studying the NF2 tumor suppressor, we propose a model for merlin growth regulation in which CD44 links growth signals from plasma membrane to the nucleus by interacting with ERM proteins and merlin.
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Affiliation(s)
- Chun-Xiao Sun
- Department of Neurology, Washington University School of Medicine, St Louis, MO 63110, USA
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Zhou CZ, Peng ZH, Zhang F, Qiu GQ, He L. Loss of heterozygosity on long arm of chromosome 22 in sporadic colorectal carcinoma. World J Gastroenterol 2002; 8:668-73. [PMID: 12174376 PMCID: PMC4656318 DOI: 10.3748/wjg.v8.i4.668] [Citation(s) in RCA: 28] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: The loss of heterozygosity (LOH) on tumor suppressor genes is believed to play a key role in carcinogenesis of colorectal cancer. In this study, we analyzed the LOH at 5 loci on the long arm of chromosome 22 in sporadic colorectal cancer to identify additional loci involved in colorectal tumorigenesis.
METHODS: Five polymorphic microsatellite markers were analyzed in 83 cases of colorectal and normal DNA by PCR. PCR products were eletrophoresed on an ABI 377 DNA sequencer; Genescan 3.1 and Genotype 2.1 software were used for LOH scanning and analysis. Comparison between LOH frequency and clinicopathological data were performed by χ² test. P < 0.05 was considered as statistically significant.
RESULTS: The average LOH frequency on chromosome 22q was 28.38%. The region between markers D22S280 and D22S274 (22q12.2-q13.33) exhibited relatively high LOH frequency. The two highest LOH loci with frequencies of 35.09% and 34.04% was identified on D22S280 (22q12.2-12.3) and D22S274 (22q13.32-13.33).8 cases showed LOH at all informative loci, suggesting that one chromosome 22q had been completely lost. On D22S274 locus, LOH frequency of rectal cancer was 50% (9/18), which was higher than that of proximal colon cancer (12%, 2/17) (P = 0.018). The frequency of distal colon cancer was 42% (5/12), also higher than that of proximal colon cancer. But there was no statistical significance. Putting both the tumors in distal colon and rectum together into consideration, the frequency, 47% (14/30), was higher than that of proximal colon cancer (P = 0.015), suggesting the mechanism of carcinogenisis was different in both groups.
CONCLUSIONS: This study provided evidence for the involvement of putative tumor suppressor genes related to the sporadic colorectal carcinoma on chromosome 22q. The tumor-suppressor-gene (s) might locate on the 22q12.2-12.3 and/or 22q13.32-13.33.
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Affiliation(s)
- Chong-Zhi Zhou
- Department of General Surgery, Shanghai First People Hospital, 85 Wujin Road, Shanghai 200080, China
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Gutmann DH, Giovannini M. Mouse models of neurofibromatosis 1 and 2. Neoplasia 2002; 4:279-90. [PMID: 12082543 PMCID: PMC1531708 DOI: 10.1038/sj.neo.7900249] [Citation(s) in RCA: 53] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/07/2002] [Accepted: 03/08/2002] [Indexed: 11/08/2022]
Abstract
The neurofibromatoses represent two of the most common inherited tumor predisposition syndromes affecting the nervous system. Individuals with neurofibromatosis 1 (NF1) are prone to the development of astrocytomas and peripheral nerve sheath tumors whereas those affected with neurofibromatosis 2 (NF2) develop schwannomas and meningiomas. The development of traditional homozygous knockout mice has provided insights into the roles of the NF1 and NF2 genes during development and in differentiation, but has been less instructive regarding the contribution of NF1 and NF2 dysfunction to the pathogenesis of specific benign and malignant tumors. Recent progress employing novel mouse targeting strategies has begun to illuminate the roles of the NF1 and NF2 gene products in the molecular pathogenesis of NF-associated tumors.
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Affiliation(s)
- David H Gutmann
- Department of Neurology, Washington University School of Medicine, Box 8111, 660 S. Euclid Avenue, St. Louis, MO 63110, USA.
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Uhlmann EJ, Apicelli AJ, Baldwin RL, Burke SP, Bajenaru ML, Onda H, Kwiatkowski D, Gutmann DH. Heterozygosity for the tuberous sclerosis complex (TSC) gene products results in increased astrocyte numbers and decreased p27-Kip1 expression in TSC2+/- cells. Oncogene 2002; 21:4050-9. [PMID: 12037687 DOI: 10.1038/sj.onc.1205435] [Citation(s) in RCA: 62] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/29/2001] [Revised: 11/15/2001] [Accepted: 11/26/2001] [Indexed: 11/09/2022]
Abstract
Tuberous sclerosis complex (TSC) is an autosomal dominant tumor predisposition syndrome characterized by benign proliferations (hamartomas). In the brain, individuals with TSC develop autism, mental retardation and seizures associated with focal cortical dysplasias, subependymal nodules, and subependymal giant cell astrocytomas (SEGAs). We hypothesize that dysregulated astrocyte function due to mutations in the tumor suppressor genes, TSC1 and TSC2, may contribute to the pathogenesis of these brain abnormalities. In this report, we demonstrate that mice heterozygous for a targeted defect in either the Tsc1 or Tsc2 genes(Tsc1+/- and Tsc2+/- mice) exhibit a 1.5-fold increase in the number of astrocytes in vivo. Whereas increased astrocyte numbers in vivo were suggestive of a proliferative advantage, Tsc2+/- primary astrocyte cultures did not show a cell-autonomous growth advantage, anchorage-independent growth, increased saturation density, or increased fluid-phase endocytosis compared to wild type astrocytes. Tsc2 null mouse embryonic fibroblasts (MEFs) however, did exhibit increased saturation density compared to Tsc2 wild type controls. In both Tsc2+/- astrocytes and Tsc2 null mouse embryonic fibroblasts, p27-Kip1 expression was decreased compared to wild type cells, and was reversed by tuberin re-expression in Tsc2-/- MEFs. In contrast, no change in endocytosis was observed upon tuberin re-expression in Tsc2-/- MEFs. Collectively, these results suggest Tsc heterozygosity may provide a non-cell-autonomous growth advantage for astrocytes that may involve p27-Kip1 expression.
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Affiliation(s)
- Erik J Uhlmann
- Department of Neurology, Washington University School of Medicine, St Louis, Missouri 63110, USA
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Shimizu T, Seto A, Maita N, Hamada K, Tsukita S, Tsukita S, Hakoshima T. Structural basis for neurofibromatosis type 2. Crystal structure of the merlin FERM domain. J Biol Chem 2002; 277:10332-6. [PMID: 11756419 DOI: 10.1074/jbc.m109979200] [Citation(s) in RCA: 88] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
Neurofibromatosis type 2 (NF2) is a dominantly inherited disease associated with the central nervous system. The NF2 gene product merlin is a tumor suppressor, and its mutation or inactivation causes this disease. We report here the crystal structure of the merlin FERM domain containing a 22-residue alpha-helical segment. The structure reveals that the merlin FERM domain consists of three subdomains displaying notable features of the electrostatic surface potentials, although the overall surface potentials similar to those of ezrin/radixin/moesin (ERM) proteins indicate electrostatic membrane association. The structure also is consistent with inactivation mechanisms caused by the pathogenic mutations associated with NF2.
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Affiliation(s)
- Toshiyuki Shimizu
- Structural Biology Laboratory, Nara Institute of Science and Technology and CREST, Japan
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Bashour AM, Meng JJ, Ip W, MacCollin M, Ratner N. The neurofibromatosis type 2 gene product, merlin, reverses the F-actin cytoskeletal defects in primary human Schwannoma cells. Mol Cell Biol 2002; 22:1150-7. [PMID: 11809806 PMCID: PMC134629 DOI: 10.1128/mcb.22.4.1150-1157.2002] [Citation(s) in RCA: 56] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Schwannoma tumors, which occur sporadically and in patients with neurofibromatosis, account for 8% of intracranial tumors and can only be treated by surgical removal. Most schwannomas have biallelic mutations in the NF2 tumor suppressor gene. We previously showed that schwannoma-derived Schwann cells exhibit membrane ruffling and aberrant cell spreading when plated onto laminin, indicative of fundamental F-actin cytoskeletal defects. Here we expand these observations to a large group of sporadic and NF2-related tumors and extend them to schwannomatosis-derived tumors. Mutation at NF2 correlated with F-actin abnormalities, but the extent of morphological change did not correlate with the type of NF2 mutation. We used a recently described molecular strategy, TAT-mediated protein transfer, to acutely introduce the NF2 protein, merlin, into primary human schwannoma cells in an attempt to reverse the cytoskeletal phenotype. Abnormal ruffling and cell spreading by cells with identified NF2 mutations were rapidly reversed by introduction of TAT-merlin. The effect is specific to TAT-merlin isoform 1, the growth-suppressive isoform of merlin. TAT-merlin isoform 2, a TAT-merlin mutant (L64P), and merlin lacking TAT were ineffective in reversing the cytoskeletal phenotype. Results show that merlin isoform 1 is sufficient to restore normal actin organization in NF2-deficient human tumor cells, demonstrating a key role for merlin in the NF2 phenotype. These results lay the foundation for epigenetic complementation studies in NF2 mouse models and possibly for experiments to evaluate the utility of merlin transduction into patients as protein therapy.
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Affiliation(s)
- Anne-Marie Bashour
- Department of Cell Biology, Neurobiology and Anatomy, University of Cincinnati College of Medicine, Cincinnati, Ohio 45267, USA
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