DOCSLIB.ORG

Expression Profiling of RE1-Silencing Transcription Factor (REST), REST

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Expression Profiling of RE1-Silencing Transcription Factor (REST), REST JBUON 2016; 21(4): 964-972 ISSN: 1107-0625, online ISSN: 2241-6293 • www.jbuon.com E-mail: [email protected] ORIGINAL ARTICLE Expression profiling of RE1-silencing transcription factor (REST), REST corepressor 1 (RCOR1), and Synapsin 1 (SYN1) genes in human gliomas Musteyde Yucebas1, Sunde Yilmaz Susluer1, Hasan Onur Caglar2, Tugce Balci1, Z. Ozlem Dogan Sigva1, Taner Akalin3, Nezih Oktar4, Tayfun Dalbasti4, Cigir Biray Avci1, Cumhur Gunduz1 1Ege University Medical Faculty, Department of Medical Biology, Bornova, Izmir; 2Ege University, Health Science Institute, Department of Stem Cell, Bornova, Izmir; 3Ege University Medical Faculty, Department of Pathology, Bornova, Izmir; 4Ege University, Medical Faculty, Department of Neurosurgery, Bornova, Izmir, Turkey Summary Purpose: The repressor element 1 (RE-1) silencing tran- Results: Means of relative expression for REST were as scription factor (REST) is a transcription factor which re- follows: 0.7898, 0.7606, and 0.7318 in DA, AO, and GBM presses the expression of neuronal differentiation-related groups, respectively. For RCOR1, expression means in DA, genes including SYN1 gene. CoREST, encoded by RCOR1 AO, and GBM groups were 0.7203, 0.7334, and 0.7230, re- gene, binds to the REST protein for remodeling of chroma- spectively. SYN1 expression means were as follows: 0.3936, tin structure. Although there is a relation among REST, 0.3192, and 0.3197 in DA, AO, and GBM groups, respective- RCOR1, and SYN1 genes, the role of these genes in glioma ly. Neither gain nor loss of copy numbers were detected for tumors is still unclear. In this study, expressions of REST, REST and RCOR1 genes in all groups. Copy loss for SYN1 RCOR1, and SYN1 genes were detected in primary cultures was detected in primary culture of a DA case. REST and derived from tumor samples of diffuse astrocytoma (DA), CoREST presented positive precipitation pattern on pro- anaplastic oligodendroglioma (AO), and glioblastoma mul- moter region of SYN1 gene. tiforme (GBM) cases. Conclusions: Expressions of REST and RCOR1 genes may Methods: Expression profiles were analysed by RT-qPCR downregulate SYN1 expression in gliomas. Low expression and the copy number variations were examined with qPCR pattern of SYN1 may maintain cancer stem-like phenotype in primary cultures. ChIP assay was performed to show which contributes to development of gliomas. binding characteristics of REST and CoREST proteins on promoter region of SYN1 gene. Key words: gene expression, gliomas, RCOR1, REST, SYN1 Introduction Gliomas are central nervous system tumors toma (grade IV) [1]. Classification system of glio- that originate from glial cells such as astrocytes or mas is based on the presence of necrosis in tumor oligodendrocytes which maintain and support the tissue, mitotic activity, and vascular or micro-vas- physiology of neuron cells. According to World cular proliferation [1]. Histopathological features Health Organization, gliomas are classified as as- of gliomas are associated with tumor recurrence, trocytoma (grade II), oligoastrcytoma (grade II), therapy response, and median survival of patients. oligodendroglioma (grade II), anaplastic astrocy- Glioblastoma tumors exhibit resistance to therapy toma/oligodendroglioma (grade III) and glioblas- and tumor recurrence after therapy. Besides, medi- Correspondence to: Sunde Yilmaz Susluer, PhD. Department of Medical Biology, Faculty of Medicine, Ege University, Izmir, Turkey, Tel: +90 232 390 2260, Fax: +90 232 342 05 42, E-mail: [email protected] Received: 26/12/2015; Accepted: 11/01/2016 REST, RCOR1 and SYN1 in gliomas 965 an survival for glioblastoma patients ranges from regulate the expression of the SYN1 gene. SYN1 14 to 16 months [2,3]. Studies indicate that gene gene expression is up-regulated during brain de- mutations, co-deletions, and extra-chromosomes velopment and this gene is expressed in adult are related with WHO grading and the prognosis neural tissue. Repression of the REST gene with of gliomas [4]. second-generation antisense oligonucleotides in- The repressor element 1 (RE-1) silencing tran- creases SYN1 expression in liver cells [21]. SYN1 scription factor (REST) is a transcription factor promoter has REST binding motifs and its expres- consisting of DNA binding and repressor domains sion is directly downregulated by REST protein [5,6]. DNA binding domain of REST binds to RE-1 [22]. Although there is a link among REST, RCOR1, sequence on promoter region of target gene, re- and SYN1 genes, interplay of these genes in glio- sulting in repression of expression [6]. It is known ma tumors is still unclear. that REST protein suppresses several neural dif- In this study, expressions of REST, RCOR1, ferentiation-related genes in the non-neuronal and SYN1 genes were analyzed in primary cell cells, while REST expression is down-regulated in cultures derived from tumor samples of DA, AO, neuronal precursor cells [5,7-11]. In carcinogen- and GBM cases. In order to demonstrate the cor- esis, REST can act as a tumor suppressor or an relation between copy number status and expres- oncogene. A splice variant of REST gene in small sion values, RT-qPCR was used. To show the bind- cell lung cancer cells (SCLC) causes production ing features of REST and CoREST transcription of truncated mature mRNA and this mRNA vari- factor on promoter region of SYN1 gene, chroma- ant down-regulates function of wild type REST tin immunoprecipitation assay was performed for protein [12]. Contrary to its tumor suppressor glioma derived primary cultures. function in SCLC, REST can act as an oncogene in neuroblastoma and medulloblastoma tumors and over-expression of REST has been identified Methods for these tumors [13,14]. Single nucleotide poly- Patients morphisms of REST leading to amino acid sub- stitutions are observed in primary and secondary Forty-nine patients with brain tumor (18 female, glioblastomas [15]. Furthermore, these polymor- 31 male) were recruited into the study. Informed con- sent was obtained from each patient and the study was phisms in REST are detected in other gliomas approved by the Local Ethics Committee of Ege Uni- such as astrocytoma, oligoastrocytoma, oligoden- versity, School of Medicine. The distribution of tumors drogliomas [15]. In addition to mutations, ampli- according to histopathology was as follows: 11 DA fication of REST is detected in glioblastomas [15]. (22.5%), 8 AO (16.3%), and 30 GBM (61.2%). The mean REST expression is found in GBM specimens and age was 53.00 ± 13.92 years. Demographic and clini- maintains self-renewal and tumor-initiating ca- cal features of cases are presented in Table 1. Prima- pacity of GBM stem-like cells. High REST expres- ry cell cultures of the tumor tissues were cultured in sion also induces aggressive properties of GBM BIOAMF-1 basal medium (Biological Industries, Israel) tumors in in vivo studies [16,17]. containing to 10,000U/ml penicillin, 10mg/ml strepto- Repressor domain of REST protein (RD2) in- mycin, 2 mM L-glutamine at 37oC in a humidified 95% teracts with REST corepressor 1 which is encoded air and 5% CO2 atmosphere. Expanded cell cultures of by the RCOR1 gene. REST and CoREST are assem- the third passage were used. bled with the histone demethylase LSD1/KDM1A and the histone deacetylases 1 and 2 (HDAC1/2) Isolation of total RNA and genomic DNA proteins to modify chromatin structure [6]. Chro- In order to detect mRNA expression of REST, matin repression by this complex leads to down- RCOR1, and SYN1 genes, total RNA were isolated from regulation of gene expression. For histone modi- primary tumor cell cultures. High Pure RNA Isolation fication, a complex consisting of both REST and Kit (Roche Applied Science, Mannheim, Germany) was CoREST may interact with different proteins such used to isolate total RNA from primary tumor cell cul- as SUMO and human positive coactivator 4 (PC4) tures according to manufacturer’s instructions. Total [18,19]. All three members of the CoREST family RNA was converted to cDNA with using Transcriptor proteins are expressed in brain tissue and this ex- First Strand cDNA Synthesis Kit (Roche Diagnostics, pression profile is observed at common territories Mannheim, Germany). cDNA samples were used for of brain. However, interactions of CoREST with gene expression analysis. For detection of copy number HDAC proteins in histone remodeling complex variations of REST, RCOR1, and SYN1 genes, genom- may show specific affinity depending on CoREST ic DNA samples were used. High Pure PCR Template family members [20]. REST and CoREST can also Preparation Kit for Genomic DNA (Roche Diagnostics, JBUON 2016; 21(4): 965 966 REST, RCOR1 and SYN1 in gliomas Table 1. Demographic and clinical features of glioma cases Gender Case number Age, years, mean Standard deviation Min age Max age (years) (years) F 18 49.78 15.40 15 73 M 31 54.87 12.87 30 79 Total 49 53.00 13.92 15 79 Diagnosis Grade Case number % Diffuse astrocytoma II 11 22.5 Anaplastic oligodendroglioma III 8 16.3 Glioblastoma IV 30 61.2 Total 49 100 F: female, M: male Mannheim, Germany) was used to obtain DNA samples pionChIP Antibody Kit (Sabiosciences, Germany-QIA- according to manufacturer’s instructions. GEN GmbH) was used as positive and negative control. Assays were performed for each antibody for 24 cases. Detection of gene expression and copy number variation Champion ChIP-One Day Kit (Sabiosciences, Germa- ny-QIAGEN GmbH) was used for ChIP assay. Exper- For the detection of gene expression, 2.6 µl cDNA imental steps were performed according to the man- were mixed with 7.4 µl Light Cycler PCR Master Mix ufacturer’s instructions. Primary tumor cell cultures and sterile bidistilled water was added to PCR mix. The obtained from tumor tissues were cultured until their expression studies of REST, RCOR1, and SYN1 genes numbers reached to 4-6x106. Chromatin structures in were performed with Real Time Online qRT-PCR ac- cell lysates were sheared with Sonicator S-4000 (Mis- cording to the Taqman protocol.
Load more

Recommended publications
  • A Computational Approach for Defining a Signature of Β-Cell Golgi Stress in Diabetes Mellitus
    Page 1 of 781 Diabetes A Computational Approach for Defining a Signature of β-Cell Golgi Stress in Diabetes Mellitus Robert N. Bone1,6,7, Olufunmilola Oyebamiji2, Sayali Talware2, Sharmila Selvaraj2, Preethi Krishnan3,6, Farooq Syed1,6,7, Huanmei Wu2, Carmella Evans-Molina 1,3,4,5,6,7,8* Departments of 1Pediatrics, 3Medicine, 4Anatomy, Cell Biology & Physiology, 5Biochemistry & Molecular Biology, the 6Center for Diabetes & Metabolic Diseases, and the 7Herman B. Wells Center for Pediatric Research, Indiana University School of Medicine, Indianapolis, IN 46202; 2Department of BioHealth Informatics, Indiana University-Purdue University Indianapolis, Indianapolis, IN, 46202; 8Roudebush VA Medical Center, Indianapolis, IN 46202. *Corresponding Author(s): Carmella Evans-Molina, MD, PhD ([email protected]) Indiana University School of Medicine, 635 Barnhill Drive, MS 2031A, Indianapolis, IN 46202, Telephone: (317) 274-4145, Fax (317) 274-4107 Running Title: Golgi Stress Response in Diabetes Word Count: 4358 Number of Figures: 6 Keywords: Golgi apparatus stress, Islets, β cell, Type 1 diabetes, Type 2 diabetes 1 Diabetes Publish Ahead of Print, published online August 20, 2020 Diabetes Page 2 of 781 ABSTRACT The Golgi apparatus (GA) is an important site of insulin processing and granule maturation, but whether GA organelle dysfunction and GA stress are present in the diabetic β-cell has not been tested. We utilized an informatics-based approach to develop a transcriptional signature of β-cell GA stress using existing RNA sequencing and microarray datasets generated using human islets from donors with diabetes and islets where type 1(T1D) and type 2 diabetes (T2D) had been modeled ex vivo. To narrow our results to GA-specific genes, we applied a filter set of 1,030 genes accepted as GA associated.
    [Show full text]
  • Supplemental Information
    Supplemental information Dissection of the genomic structure of the miR-183/96/182 gene. Previously, we showed that the miR-183/96/182 cluster is an intergenic miRNA cluster, located in a ~60-kb interval between the genes encoding nuclear respiratory factor-1 (Nrf1) and ubiquitin-conjugating enzyme E2H (Ube2h) on mouse chr6qA3.3 (1). To start to uncover the genomic structure of the miR- 183/96/182 gene, we first studied genomic features around miR-183/96/182 in the UCSC genome browser (http://genome.UCSC.edu/), and identified two CpG islands 3.4-6.5 kb 5’ of pre-miR-183, the most 5’ miRNA of the cluster (Fig. 1A; Fig. S1 and Seq. S1). A cDNA clone, AK044220, located at 3.2-4.6 kb 5’ to pre-miR-183, encompasses the second CpG island (Fig. 1A; Fig. S1). We hypothesized that this cDNA clone was derived from 5’ exon(s) of the primary transcript of the miR-183/96/182 gene, as CpG islands are often associated with promoters (2). Supporting this hypothesis, multiple expressed sequences detected by gene-trap clones, including clone D016D06 (3, 4), were co-localized with the cDNA clone AK044220 (Fig. 1A; Fig. S1). Clone D016D06, deposited by the German GeneTrap Consortium (GGTC) (http://tikus.gsf.de) (3, 4), was derived from insertion of a retroviral construct, rFlpROSAβgeo in 129S2 ES cells (Fig. 1A and C). The rFlpROSAβgeo construct carries a promoterless reporter gene, the β−geo cassette - an in-frame fusion of the β-galactosidase and neomycin resistance (Neor) gene (5), with a splicing acceptor (SA) immediately upstream, and a polyA signal downstream of the β−geo cassette (Fig.
    [Show full text]
  • WO 2019/079361 Al 25 April 2019 (25.04.2019) W 1P O PCT
    (12) INTERNATIONAL APPLICATION PUBLISHED UNDER THE PATENT COOPERATION TREATY (PCT) (19) World Intellectual Property Organization I International Bureau (10) International Publication Number (43) International Publication Date WO 2019/079361 Al 25 April 2019 (25.04.2019) W 1P O PCT (51) International Patent Classification: CA, CH, CL, CN, CO, CR, CU, CZ, DE, DJ, DK, DM, DO, C12Q 1/68 (2018.01) A61P 31/18 (2006.01) DZ, EC, EE, EG, ES, FI, GB, GD, GE, GH, GM, GT, HN, C12Q 1/70 (2006.01) HR, HU, ID, IL, IN, IR, IS, JO, JP, KE, KG, KH, KN, KP, KR, KW, KZ, LA, LC, LK, LR, LS, LU, LY, MA, MD, ME, (21) International Application Number: MG, MK, MN, MW, MX, MY, MZ, NA, NG, NI, NO, NZ, PCT/US2018/056167 OM, PA, PE, PG, PH, PL, PT, QA, RO, RS, RU, RW, SA, (22) International Filing Date: SC, SD, SE, SG, SK, SL, SM, ST, SV, SY, TH, TJ, TM, TN, 16 October 2018 (16. 10.2018) TR, TT, TZ, UA, UG, US, UZ, VC, VN, ZA, ZM, ZW. (25) Filing Language: English (84) Designated States (unless otherwise indicated, for every kind of regional protection available): ARIPO (BW, GH, (26) Publication Language: English GM, KE, LR, LS, MW, MZ, NA, RW, SD, SL, ST, SZ, TZ, (30) Priority Data: UG, ZM, ZW), Eurasian (AM, AZ, BY, KG, KZ, RU, TJ, 62/573,025 16 October 2017 (16. 10.2017) US TM), European (AL, AT, BE, BG, CH, CY, CZ, DE, DK, EE, ES, FI, FR, GB, GR, HR, HU, ΓΕ , IS, IT, LT, LU, LV, (71) Applicant: MASSACHUSETTS INSTITUTE OF MC, MK, MT, NL, NO, PL, PT, RO, RS, SE, SI, SK, SM, TECHNOLOGY [US/US]; 77 Massachusetts Avenue, TR), OAPI (BF, BJ, CF, CG, CI, CM, GA, GN, GQ, GW, Cambridge, Massachusetts 02139 (US).
    [Show full text]
  • Supporting Information
    Supporting Information The deubiquitinase USP38 promotes NHEJ repair through regulation of HDAC1 activity and regulates cancer cell response to genotoxic insults Yongfeng Yang1,2, Chuanzhen Yang1，2, Tingting Li3, Shuyu Yu1,2,Tingting Gan4, Jiazhi Hu4, Jun Cui5,6, and Xiaofeng Zheng1,2,* 1 State Key Laboratory of Protein and Plant Gene Research, School of Life Sciences, Peking University, Beijing, China. 2Department of Biochemistry and Molecular Biology, School of Life Sciences, Peking University, Beijing, China. 3State Key Laboratory of Proteomics, National Center of Biomedical Analysis, Institute of Basic Medical Sciences, Beijing, China. 4Department of Cell Biology, School of Life Sciences, Peking University, Beijing, China. 5Key Laboratory of Gene Engineering of the Ministry of Education, State Key Laboratory of Biocontrol, School of Life Sciences, Sun Yat-sen University, Guangzhou, China. 6Collaborative Innovation Center of Cancer Medicine, Sun Yat-sen University, Guangzhou, China. *To whom correspondence should be addressed. Xiaofeng Zheng, School of Life Sciences, Peking University, Beijing 100871, China. Tel: +86 10-6275-5712; Email: [email protected] Supplementary Materials and Methods: Antibodies and reagents Mouse monoclonal anti-Flag (F3165, RRID: AB_259529) antibodies, anti-HA (H9658, RRID: AB_260092) antibodies, and etoposide (E1383) were purchased from Sigma-Aldrich. Mouse monoclonal anti-Myc (M047-3, RRID: AB_591112), anti-histidine (D291–3, RRID: AB_10597733), and rabbit polyclonal anti-β-actin (PM053, RRID: AB_10598196) antibodies were purchased from MBL. Mouse monoclonal anti-H3 (BE3015) antibodies were purchased from EASYBIO. Mouse monoclonal anti-γH2AX (05–636, RRID: AB_309864) antibodies and MG132 (2772605) were purchased from Millipore. Mouse monoclonal anti-HDAC1 (sc-81598, RRID: AB_2118083) antibodies were purchased from Santa Cruz Biotechnology.
    [Show full text]
  • Screen for Multi-SUMO–Binding Proteins Reveals a Multi-SIM–Binding Mechanism for Recruitment of the Transcriptional Regulator ZMYM2 to Chromatin
    Screen for multi-SUMO–binding proteins reveals a multi-SIM–binding mechanism for recruitment of the transcriptional regulator ZMYM2 to chromatin Elisa Aguilar-Martineza, Xi Chena, Aaron Webbera, A. Paul Moulda, Anne Seifertb, Ronald T. Hayb, and Andrew D. Sharrocksa,1 aFaculty of Life Sciences, University of Manchester, Manchester M13 9PT, United Kingdom; and bCentre for Gene Regulation and Expression, University of Dundee, Dundee DD1 5EH, United Kingdom Edited by James L. Manley, Columbia University, New York, NY, and approved July 17, 2015 (received for review May 20, 2015) Protein SUMOylation has emerged as an important regulatory human proteins containing two or more motifs corresponding event, particularly in nuclear processes such as transcriptional to the extended negatively charged amino acid-dependent control and DNA repair. In this context, small ubiquitin-like modifier SUMOylation motif (NDSM) (13) Thus, there is a huge poten- (SUMO) often provides a binding platform for the recruitment of tial for widespread multi-SUMOylation of proteins to occur. proteins via their SUMO-interacting motifs (SIMs). Recent discoveries Indeed, several of these proteins have been shown to be point to an important role for multivalent SUMO binding through SUMOylated on multiple sites, including megakaryoblastic leu- multiple SIMs in the binding partner as exemplified by poly- kemia (translocation) 1 (MKL1) (14), CREB-binding protein SUMOylation acting as a binding platform for ubiquitin E3 ligases (CBP) (15), and PEA3/ETV4 (16). Furthermore, two recent such as ring finger protein 4. Here, we have investigated whether proteomic studies emphasize the potential for more widespread other types of protein are recruited through multivalent SUMO multi-SUMOylation as they found that a large proportion of all interactions.
    [Show full text]
  • Whole Transcriptomic Expression Differences in EBV Immortalized Versus Primary B-Cells
    W&M ScholarWorks Undergraduate Honors Theses Theses, Dissertations, & Master Projects 12-2010 Whole Transcriptomic Expression Differences in EBV Immortalized versus Primary B-Cells Dolores Huberts College of William and Mary Follow this and additional works at: https://scholarworks.wm.edu/honorstheses Part of the Biology Commons Recommended Citation Huberts, Dolores, "Whole Transcriptomic Expression Differences in EBV Immortalized versus Primary B- Cells" (2010). Undergraduate Honors Theses. Paper 347. https://scholarworks.wm.edu/honorstheses/347 This Honors Thesis is brought to you for free and open access by the Theses, Dissertations, & Master Projects at W&M ScholarWorks. It has been accepted for inclusion in Undergraduate Honors Theses by an authorized administrator of W&M ScholarWorks. For more information, please contact [email protected]. Whole Transcriptomic Expression Differences in EBV Immortalized versus Primary B-Cells A thesis submitted in partial fulfillment of the requirement for the degree of Bachelor of Science with Honors in Biology from the College of William and Mary in Virginia By Dolores Huberts Accepted for Honors ________________________________________ Lizabeth A. Allison, Director ________________________________________ Matthew Wawersik ________________________________________ Drew LaMar ________________________________________ Beverly Sher Williamsburg, Virginia December 17, 2010 ABSTRACT The Epstein–Barr Virus (EBV) is a human gamma herpes virus that infects more than 90% of the human population worldwide. It is commonly known in the US as the cause of Infectious Mononucleosis, and around the world as the cause of nasopharyngeal carcinoma and malignant lymphomas such as non-Hodgkin lymphoma, endemic Burkett’s lymphoma and Hodgkin lymphoma. Additionally, the EBV is used to immortalize cells to create cell lines for in-vitro studies.
    [Show full text]
  • Characterization of the Zinc Finger Proteins ZMYM2 and ZMYM4 As
    www.nature.com/scientificreports OPEN Characterization of the zinc fnger proteins ZMYM2 and ZMYM4 as novel B-MYB binding proteins Hannah Cibis, Abhiruchi Biyanee, Wolfgang Dörner, Henning D. Mootz & Karl-Heinz Klempnauer✉ B-MYB, a highly conserved member of the MYB transcription factor family, is expressed ubiquitously in proliferating cells and plays key roles in important cell cycle-related processes, such as control of G2/M- phase transcription, cytokinesis, G1/S-phase progression and DNA-damage reponse. Deregulation of B-MYB function is characteristic of several types of tumor cells, underlining its oncogenic potential. To gain a better understanding of the functions of B-MYB we have employed afnity purifcation coupled to mass spectrometry to discover novel B-MYB interacting proteins. Here we have identifed the zinc-fnger proteins ZMYM2 and ZMYM4 as novel B-MYB binding proteins. ZMYM4 is a poorly studied protein whose initial characterization reported here shows that it is highly SUMOylated and that its interaction with B-MYB is stimulated upon induction of DNA damage. Unlike knockdown of B-MYB, which causes G2/M arrest and defective cytokinesis in HEK293 cells, knockdown of ZMYM2 or ZMYM4 have no obvious efects on the cell cycle of these cells. By contrast, knockdown of ZMYM2 strongly impaired the G1/S-phase progression of HepG2 cells, suggesting that ZMYM2, like B-MYB, is required for entry into S-phase in these cells. Overall, our work identifes two novel B-MYB binding partners with possible functions in the DNA-damage response and the G1/S-transition. Te highly conserved MYB proto-oncogene family member B-MYB is ubiquitously expressed in proliferat- ing cells where it acts as an essential cell cycle-regulated transcription factor1,2.
    [Show full text]
  • VHL Inactivation Without Hypoxia Is Sufficient to Achieve Genome Hypermethylation
    bioRxiv preprint doi: https://doi.org/10.1101/093310; this version posted December 12, 2016. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. VHL inactivation without hypoxia is sufficient to achieve genome hypermethylation Artem V. Artemov1*, Nadezhda Zhigalova1, Svetlana Zhenilo1, Alexander M. Mazur1 and Egor B. Prokhortchouk1 1 Institute of Bioengineering, Research Center of Biotechnology RAS, Moscow, Russian Federation * [email protected] Abstract VHL inactivation is a key oncogenic event for renal carcinomas. In normoxia, VHL suppresses HIF1a-mediated response to hypoxia. It has previously been shown that hypoxic conditions inhibit TET-dependent hydroxymethylation of cytosines and cause DNA hypermethylation at gene promoters. In this work, we performed VHL inactivation by CRISPR/Cas9 and studied its effects on gene expression and DNA methylation. We showed that even without hypoxia, VHL inactivation leads to hypermethylation of the genome which mainly occurred in AP-1 and TRIM28 binding sites. We also observed promoter hypermethylation of several transcription regulators associated with decreased gene expression. Keywords DNA methylation; VHL; hypoxia; HIF1a; JUN; FOS Introduction Sequencing of cancer genomes was initially aimed to find cancer drivers, or genes, that, once mutated, give a selective advantage to a cancer cell, such as increased proliferation, suppression of apoptosis or the ability to avoid immune response. VHL is a key oncosuppressor gene for kidney cancer. Inactivation of the VHL gene is the most common event in renal carcinomas, accounting for 50{70% of sporadic cases (Scelo et al. 2014; Cancer Genome Atlas Research Network 2013; Thomas et al.
    [Show full text]
  • REST Upregulates Gremlin to Modulate Diffuse Intrinsic Pontine Glioma Vasculature
    www.impactjournals.com/oncotarget/ Oncotarget, 2018, Vol. 9, (No. 4), pp: 5233-5250 Research Paper REST upregulates gremlin to modulate diffuse intrinsic pontine glioma vasculature Shavali Shaik1,*, Bridget Kennis1,*, Shinji Maegawa1, Keri Schadler1, Yang Yanwen1, Keri Callegari1, Rishi R. Lulla3, Stewart Goldman3, Javad Nazarian4, Veena Rajaram5, Jason Fangusaro3 and Vidya Gopalakrishnan1,2,6,7 1Department of Pediatrics, University of Texas, MD Anderson Cancer Center, Houston, Texas, USA 2Department of Molecular and Cellular Oncology, University of Texas, MD Anderson Cancer Center, Houston, Texas, USA 3Department of Pediatrics, Northwestern Feinberg School of Medicine, Northwestern University, Chicago, Illinois, USA 4Department of Integrative Systems Biology, George Washington University School of Medicine and Health Sciences, Washington, District of Columbia, USA 5Department of Pathology, University of Texas Southwestern Medical Center, Dallas, Texas, USA 6Center for Cancer Epigenetics, University of Texas, MD Anderson Cancer Center, Houston, Texas, USA 7Brain Tumor Center, University of Texas, MD Anderson Cancer Center, Houston, Texas, USA *These authors have contributed equally to this work Correspondence to: Vidya Gopalakrishnan, email: [email protected] Keywords: DIPG; REST; vasculature; gremlin; VEGFR2 Received: August 29, 2017 Accepted: December 16, 2017 Published: December 28, 2017 Copyright: Shaik et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License 3.0 (CC BY 3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. ABSTRACT Diffuse intrinsic pontine glioma (DIPG) is a highly aggressive glial tumor that occurs in children. The extremely poor median and 5-year survival in children afflicted with DIPG highlights the need for novel biology-driven therapeutics.
    [Show full text]
  • ZNF750 Interacts with KLF4 and RCOR1, KDM1A, and CTBP1/2 Chromatin Regulators to Repress Epidermal Progenitor Genes and Induce Differentiation Genes
    Downloaded from genesdev.cshlp.org on September 27, 2021 - Published by Cold Spring Harbor Laboratory Press ZNF750 interacts with KLF4 and RCOR1, KDM1A, and CTBP1/2 chromatin regulators to repress epidermal progenitor genes and induce differentiation genes Lisa D. Boxer,1,2 Brook Barajas,1 Shiying Tao,1 Jiajing Zhang,1 and Paul A. Khavari1,3 1Program in Epithelial Biology, Stanford University School of Medicine, Stanford, California 94305, USA; 2Department of Biology, Stanford University, Stanford, California 94305, USA; 3Veterans Affairs Palo Alto Healthcare System, Palo Alto, California 94304, USA ZNF750 controls epithelial homeostasis by inhibiting progenitor genes while inducing differentiation genes, a role underscored by pathogenic ZNF750 mutations in cancer and psoriasis. How ZNF750 accomplishes these dual gene regulatory impacts is unknown. Here, we characterized ZNF750 as a transcription factor that binds both the progenitor and differentiation genes that it controls at a CCNNAGGC DNA motif. ZNF750 interacts with the pluripotency transcription factor KLF4 and chromatin regulators RCOR1, KDM1A, and CTBP1/2 through conserved PLNLS sequences. ChIP-seq (chromatin immunoprecipitation [ChIP] followed by high-throughput sequencing) and gene depletion revealed that KLF4 colocalizes ~10 base pairs from ZNF750 at differentiation target genes to facilitate their activation but is unnecessary for ZNF750-mediated progenitor gene repression. In contrast, KDM1A colocalizes with ZNF750 at progenitor genes and facilitates their repression but is unnecessary for ZNF750-driven differentiation. ZNF750 thus controls differentiation in concert with RCOR1 and CTBP1/2 by acting with either KDM1A to repress progenitor genes or KLF4 to induce differentiation genes. [Keywords: stem cell; differentiation; ZNF750; KLF4; chromatin regulator] Supplemental material is available for this article.
    [Show full text]
  • NRF1) Coordinates Changes in the Transcriptional and Chromatin Landscape Affecting Development and Progression of Invasive Breast Cancer
    Florida International University FIU Digital Commons FIU Electronic Theses and Dissertations University Graduate School 11-7-2018 Decipher Mechanisms by which Nuclear Respiratory Factor One (NRF1) Coordinates Changes in the Transcriptional and Chromatin Landscape Affecting Development and Progression of Invasive Breast Cancer Jairo Ramos [email protected] Follow this and additional works at: https://digitalcommons.fiu.edu/etd Part of the Clinical Epidemiology Commons Recommended Citation Ramos, Jairo, "Decipher Mechanisms by which Nuclear Respiratory Factor One (NRF1) Coordinates Changes in the Transcriptional and Chromatin Landscape Affecting Development and Progression of Invasive Breast Cancer" (2018). FIU Electronic Theses and Dissertations. 3872. https://digitalcommons.fiu.edu/etd/3872 This work is brought to you for free and open access by the University Graduate School at FIU Digital Commons. It has been accepted for inclusion in FIU Electronic Theses and Dissertations by an authorized administrator of FIU Digital Commons. For more information, please contact [email protected]. FLORIDA INTERNATIONAL UNIVERSITY Miami, Florida DECIPHER MECHANISMS BY WHICH NUCLEAR RESPIRATORY FACTOR ONE (NRF1) COORDINATES CHANGES IN THE TRANSCRIPTIONAL AND CHROMATIN LANDSCAPE AFFECTING DEVELOPMENT AND PROGRESSION OF INVASIVE BREAST CANCER A dissertation submitted in partial fulfillment of the requirements for the degree of DOCTOR OF PHILOSOPHY in PUBLIC HEALTH by Jairo Ramos 2018 To: Dean Tomás R. Guilarte Robert Stempel College of Public Health and Social Work This dissertation, Written by Jairo Ramos, and entitled Decipher Mechanisms by Which Nuclear Respiratory Factor One (NRF1) Coordinates Changes in the Transcriptional and Chromatin Landscape Affecting Development and Progression of Invasive Breast Cancer, having been approved in respect to style and intellectual content, is referred to you for judgment.
    [Show full text]
  • Than a Corepressor: the Role of Corest Proteins in Neurodevelopment
    Review Development More than a Corepressor: The Role of CoREST Proteins in Neurodevelopment Simon Maksour,1,2 Lezanne Ooi,1,3 and Mirella Dottori1,2 https://doi.org/10.1523/ENEURO.0337-19.2020 1Illawarra Health and Medical Research Institute, Wollongong, New South Wales 2522, Australia, 2School of Medicine, University of Wollongong, Wollongong, New South Wales 2522, Australia, and 3School of Chemistry and Molecular Bioscience, University of Wollongong, Wollongong, New South Wales 2522, Australia Abstract The molecular mechanisms governing normal neurodevelopment are tightly regulated by the action of tran- scription factors. Repressor element 1 (RE1) silencing transcription factor (REST) is widely documented as a regulator of neurogenesis that acts by recruiting corepressor proteins and repressing neuronal gene expres- sion in non-neuronal cells. The REST corepressor 1 (CoREST1), CoREST2, and CoREST3 are best described for their role as part of the REST complex. However, recent evidence has shown the proteins have the ability to repress expression of distinct target genes in a REST-independent manner. These findings indicate that each CoREST paralogue may have distinct and critical roles in regulating neurodevelopment and are more than simply “REST corepressors,” whereby they act as independent repressors orchestrating biological proc- esses during neurodevelopment. Key words: CoREST; differentiation; gene expression; neurodevelopment; REST; transcription factor Significance Statement The molecular mechanisms governing normal development of the brain are yet to be fully elucidated. The regulation of gene expression by transcription factors plays a significant role in the specification and matu- ration of neurons and glia. Repressor element 1 (RE1) silencing transcription factor (REST) has been well characterized as a transcriptional regulator of neurogenesis through the formation of a complex with the REST corepressor (CoREST) proteins.
    [Show full text]