Covalent Modification of a Cysteine Residue in the XPB Subunit of The
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XPB Induces C1D Expression to Counteract UV-Induced Apoptosis
Published OnlineFirst June 8, 2010; DOI: 10.1158/1541-7786.MCR-09-0467 Molecular DNA Damage and Cellular Stress Responses Cancer Research XPB Induces C1D Expression to Counteract UV-Induced Apoptosis Guang Li1, Juhong Liu2, Mones Abu-Asab1, Shibuya Masabumi3, and Yoshiro Maru4 Abstract Although C1D has been shown to be involved in DNA double-strand break repair, how C1D expression was induced and the mechanism(s) by which C1D facilitates DNA repair in mammalian cells remain poorly understood. We and others have previously shown that expression of xeroderma pigmentosum B (XPB) pro- tein efficiently compensated the UV irradiation–sensitive phenotype of 27-1 cells, which lack functional XPB. To further explore XPB-regulated genes that could be involved in UV-induced DNA repair, differential dis- play analysis of mRNA levels from CHO-9, 27-1, and 27-1 complemented with wild-type XPB was done and C1D gene was identified as one of the major genes whose expression was significantly upregulated by restoring XPB function. We found that XPB is essential to induce C1D transcription after UV irradiation. The increase in C1D expression effectively compensates for the UV-induced proteolysis of C1D and thus maintains cellular C1D level to cope with DNA damage inflicted by UV irradiation. We further showed that although insufficient to rescue 27-1 cells from UV-induced apoptosis by itself, C1D facilitates XPB DNA repair through direct interaction with XPB. Our findings provided direct evidence that C1D is associated with DNA repair complex and may promote repair of UV-induced DNA damage. Mol Cancer Res; 8(6); 885–95. -
Oncogenic H-Ras Up-Regulates Expression of ERCC1 to Protect Cells from Platinum-Based Anticancer Agents
[CANCER RESEARCH 64, 4849–4857, July 15, 2004] Oncogenic H-Ras Up-Regulates Expression of ERCC1 to Protect Cells from Platinum-Based Anticancer Agents Cha-Kyung Youn,1 Mi-Hwa Kim,1 Hyun-Ju Cho,1 Hong-Beum Kim,1 In-Youb Chang,1 Myung-Hee Chung,3 and Ho Jin You1,2 1Research Center for Proteineous Materials and 2Department of Pharmacology, School of Medicine, Chosun University, Gwangju, and 3Department of Pharmacology, School of Medicine, Seoul National University, Seoul, Korea ABSTRACT elucidated, evidence suggests that the activated Ras may contribute to cisplatin resistance by stimulating the DNA repair activity (9, 12, 13). Tumors frequently contain mutations in the ras genes, resulting in the Hence, there has been considerable interest in determining which constitutive activation of the Ras-activated signaling pathway. The acti- vation of Ras is involved not only in tumor progression but also in the proteins mediate the altered DNA repair capacity in activated Ras- development of resistance of the tumor cells to platinum-based chemo- containing cells. However, the downstream target genes of the onco- therapeutic agents. To investigate the potential mechanisms underlying genic Ras, which are involved in the enhancement of the DNA repair this resistance, we analyzed the effect of activated H-Ras on the expression activity, are unclear. of the nucleotide excision repair genes. Here we identified ERCC1, which Cisplatin is one of the most effective and widely used anticancer is one of the key enzymes involved in nucleotide excision repair, as being drugs for treating human solid tumors (14). However, its therapeutic markedly up-regulated by the activated H-Ras. -
P38 MAPK-Mediated Activation of NF-Kb by the Rhogef Domain of Bcr
Oncogene (2002) 21, 4601 – 4612 ª 2002 Nature Publishing Group All rights reserved 0950 – 9232/02 $25.00 www.nature.com/onc p38 MAPK-mediated activation of NF-kB by the RhoGEF domain of Bcr Malgorzata Korus1,2, Gwendolyn M Mahon1,2, Li Cheng1 and Ian P Whitehead*,1 1Department of Microbiology and Molecular Genetics, UMDNJ – New Jersey Medical School, Newark, New Jersey, NJ 07103, USA The oncogenic fusion protein p210 Bcr-Abl is causally et al., 1988; Fainstein et al., 1987; Konopka et al., associated with virtually all cases of chronic myelogenous 1984; Kurzrock et al., 1987; Walker et al., 1987). leukemia. The wild-type Bcr product has several Although animal models suggest that elevated levels of recognizable structural and functional motifs including Abl tyrosine kinase activity contribute to Bcr-Abl a domain that contains guanine nucleotide exchange mediated leukemogenesis (Lugo et al., 1990), the activity for Rho family GTPases (DH/PH domain). residues that distinguish the rearrangements associated Although this domain is retained within p210 Bcr-Abl, it with ALL (p185 Bcr-Abl) from the rearrangements has no known signaling activities in vivo. Here we report associated with CML (p210 Bcr-Abl) reside within the that a fragment of Bcr that encodes the isolated DH/PH Bcr protein. Thus, a complete understanding of the domain is a potent activator of the NF-kB transcription biological mechanisms that distinguish ALL from factor. Within the context of full length Bcr, this activity CML may depend upon the characterization of the is regulated by proximal flanking sequences that suppress signaling activities that reside within Bcr. -
Role of Excess Inorganic Pyrophosphate in Primer-Extension Genotyping Assays
Methods Role of Excess Inorganic Pyrophosphate in Primer-Extension Genotyping Assays Ming Xiao,1 Angie Phong,1 Kristen L. Lum,1 Richard A. Greene,2 Philip R. Buzby,2,3 and Pui-Yan Kwok1,4,5 1Cardiovascular Research Institute, University of California, San Francisco, San Francisco, California 94143-0130, USA; 2PerkinElmer Life and Analytical Sciences, Inc., Boston, Massachusetts 02118-2512, USA We have developed and genotyped >15,000 SNP assays by using a primer extension genotyping assay with fluorescence polarization (FP) detection. Although the 80% success rate of this assay is similar to those of other SNP genotyping assays, we wanted to determine the reasons for the failures and find ways to improve the assay. We observed that the failed assays fell into three general patterns: PCR failure, excess of heterozygous genotypes, and loss of FP signal for one of the dye labels. After analyzing several hundred failed assays, we concluded that 5% of the assays had PCR failure and had to be redesigned. We also discovered that the other two categories of failures were due to misincorporation of one of the dye-terminators during the primer extension reaction as a result of primer shortening with a reverse reaction involving inorganic pyrophosphate, and to the quenching of R110-terminator after its incorporation onto the SNP primer. The relatively slow incorporation of R110 acycloterminators by AcycloPol compounds the problem with the R110 label. In this report, we describe the source of the problems and simple ways to correct these problems by adding pyrophosphatase, using quenching as part of the analysis, and replacing R110 by Texas red as one of the dye labels. -
Neuropeptidergic Signaling Partitions Arousal Behaviors in Zebrafish
3142 • The Journal of Neuroscience, February 26, 2014 • 34(9):3142–3160 Behavioral/Cognitive Neuropeptidergic Signaling Partitions Arousal Behaviors in Zebrafish Ian G. Woods,1,2 David Schoppik,2 Veronica J. Shi,2 Steven Zimmerman,2 Haley A. Coleman,1 Joel Greenwood,3 Edward R. Soucy,3 and Alexander F. Schier2,3 1Department of Biology, Ithaca College, Ithaca, New York 14850, and 2Department of Molecular and Cellular Biology and 3Center for Brain Science, Harvard University, Cambridge, Massachusetts 02138 Animals modulate their arousal state to ensure that their sensory responsiveness and locomotor activity match environmental demands. Neuropeptides can regulate arousal, but studies of their roles in vertebrates have been constrained by the vast array of neuropeptides and their pleiotropic effects. To overcome these limitations, we systematically dissected the neuropeptidergic modulation of arousal in larval zebrafish. We quantified spontaneous locomotor activity and responsiveness to sensory stimuli after genetically induced expression of seven evolutionarily conserved neuropeptides, including adenylate cyclase activating polypeptide 1b (adcyap1b), cocaine-related and amphetamine-related transcript (cart), cholecystokinin (cck), calcitonin gene-related peptide (cgrp), galanin, hypocretin, and nocicep- tin. Our study reveals that arousal behaviors are dissociable: neuropeptide expression uncoupled spontaneous activity from sensory responsiveness, and uncovered modality-specific effects upon sensory responsiveness. Principal components analysis and phenotypic clustering revealed both shared and divergent features of neuropeptidergic functions: hypocretin and cgrp stimulated spontaneous locomotor activity, whereas galanin and nociceptin attenuated these behaviors. In contrast, cart and adcyap1b enhanced sensory respon- siveness yet had minimal impacts on spontaneous activity, and cck expression induced the opposite effects. Furthermore, hypocretin and nociceptin induced modality-specific differences in responsiveness to changes in illumination. -
Ras/Raf/MEK/ERK and PI3K/PTEN/Akt/Mtor Cascade Inhibitors: How Mutations Can Result in Therapy Resistance and How to Overcome Resistance
www.impactjournals.com/oncotarget/ Oncotarget, October, Vol.3, No 10 Ras/Raf/MEK/ERK and PI3K/PTEN/Akt/mTOR Cascade Inhibitors: How Mutations Can Result in Therapy Resistance and How to Overcome Resistance James A. McCubrey1, Linda S. Steelman1, William H. Chappell1, Stephen L. Abrams1, Richard A. Franklin1, Giuseppe Montalto2, Melchiorre Cervello3, Massimo Libra4, Saverio Candido4, Grazia Malaponte4, Maria C. Mazzarino4, Paolo Fagone4, Ferdinando Nicoletti4, Jörg Bäsecke5, Sanja Mijatovic6, Danijela Maksimovic- Ivanic6, Michele Milella7, Agostino Tafuri8, Francesca Chiarini9, Camilla Evangelisti9, Lucio Cocco10, Alberto M. Martelli9,10 1 Department of Microbiology and Immunology, Brody School of Medicine at East Carolina University, Greenville, NC, USA 2 Department of Internal Medicine and Specialties, University of Palermo, Palermo, Italy 3 Consiglio Nazionale delle Ricerche, Istituto di Biomedicina e Immunologia Molecolare “Alberto Monroy”, Palermo, Italy 4 Department of Bio-Medical Sciences, University of Catania, Catania, Italy 5 Department of Medicine, University of Göttingen, Göttingen, Germany 6 Department of Immunology, Instititue for Biological Research “Sinisa Stankovic”, University of Belgrade, Belgrade, Serbia 7 Regina Elena National Cancer Institute, Rome, Italy 8 Sapienza, University of Rome, Department of Cellular Biotechnology and Hematology, Rome, Italy 9 Institute of Molecular Genetics, National Research Council-Rizzoli Orthopedic Institute, Bologna, Italy. 10 Department of Biomedical and Neuromotor Sciences, University of Bologna, Bologna, Italy Correspondence to: James A. McCubrey, email: [email protected] Keywords: Targeted Therapy, Therapy Resistance, Cancer Stem Cells, Raf, Akt, PI3K, mTOR Received: September 12, 2012, Accepted: October 18, 2012, Published: October 20, 2012 Copyright: © McCubrey et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. -
Discovery of the Essential Roles of CRK9 in Gene Expression and Of
University of Connecticut OpenCommons@UConn Doctoral Dissertations University of Connecticut Graduate School 5-27-2015 Lessons from Trypanosome TFIIH: Discovery of the Essential Roles of CRK9 in Gene Expression and of the Divergent Helicase Paralog XPB-R in Nucleotide Excision Repair Nitika Badjatia University of Connecticut - Storrs, [email protected] Follow this and additional works at: https://opencommons.uconn.edu/dissertations Recommended Citation Badjatia, Nitika, "Lessons from Trypanosome TFIIH: Discovery of the Essential Roles of CRK9 in Gene Expression and of the Divergent Helicase Paralog XPB-R in Nucleotide Excision Repair" (2015). Doctoral Dissertations. 866. https://opencommons.uconn.edu/dissertations/866 Lessons from Trypanosome TFIIH: Discovery of the Essential Roles of CRK9 in Gene Expression and of the Divergent Helicase Paralog XPB-R in Nucleotide Excision Repair Nitika Badjatia University of Connecticut, [2015] Eukaryotic TFIIH consists of a core of seven subunits, including the DNA helicase Xeroderma Pigmentosum B (XPB), and a cyclin-dependent kinase (CDK)-activating complex (CAK) that contains CDK7. XPB is crucial for DNA unwinding during transcription and nucleotide excision repair (NER) while the CAK complex phosphorylates RNA polymerase II (RNAPII) carboxy terminal domain (CTD), enabling the enzyme’s promoter clearance. Trypanosoma brucei, Trypanosoma cruzi and Leishmania spp. are lethal human parasites, belonging to the early-diverged order Kinetoplastida. They transcribe genes polycistronically and require spliced leader (SL) trans splicing for the maturation of mRNAs from polycistronic precursors. SL RNA gene (SLRNA) transcription by RNAPII depends on trypanosome TFIIH complex which contains orthologs of all core subunits but lacks the CAK complex. Despite this, the trypanosome CTD is phosphorylated and essential for SLRNA transcription. -
Melanomas Are Comprised of Multiple Biologically Distinct Categories
Melanomas are comprised of multiple biologically distinct categories, which differ in cell of origin, age of onset, clinical and histologic presentation, pattern of metastasis, ethnic distribution, causative role of UV radiation, predisposing germ line alterations, mutational processes, and patterns of somatic mutations. Neoplasms are initiated by gain of function mutations in one of several primary oncogenes, typically leading to benign melanocytic nevi with characteristic histologic features. The progression of nevi is restrained by multiple tumor suppressive mechanisms. Secondary genetic alterations override these barriers and promote intermediate or overtly malignant tumors along distinct progression trajectories. The current knowledge about pathogenesis, clinical, histological and genetic features of primary melanocytic neoplasms is reviewed and integrated into a taxonomic framework. THE MOLECULAR PATHOLOGY OF MELANOMA: AN INTEGRATED TAXONOMY OF MELANOCYTIC NEOPLASIA Boris C. Bastian Corresponding Author: Boris C. Bastian, M.D. Ph.D. Gerson & Barbara Bass Bakar Distinguished Professor of Cancer Biology Departments of Dermatology and Pathology University of California, San Francisco UCSF Cardiovascular Research Institute 555 Mission Bay Blvd South Box 3118, Room 252K San Francisco, CA 94158-9001 [email protected] Key words: Genetics Pathogenesis Classification Mutation Nevi Table of Contents Molecular pathogenesis of melanocytic neoplasia .................................................... 1 Classification of melanocytic neoplasms -
Figure S1. HAEC ROS Production and ML090 NOX5-Inhibition
Figure S1. HAEC ROS production and ML090 NOX5-inhibition. (a) Extracellular H2O2 production in HAEC treated with ML090 at different concentrations and 24 h after being infected with GFP and NOX5-β adenoviruses (MOI 100). **p< 0.01, and ****p< 0.0001 vs control NOX5-β-infected cells (ML090, 0 nM). Results expressed as mean ± SEM. Fold increase vs GFP-infected cells with 0 nM of ML090. n= 6. (b) NOX5-β overexpression and DHE oxidation in HAEC. Representative images from three experiments are shown. Intracellular superoxide anion production of HAEC 24 h after infection with GFP and NOX5-β adenoviruses at different MOIs treated or not with ML090 (10 nM). MOI: Multiplicity of infection. Figure S2. Ontology analysis of HAEC infected with NOX5-β. Ontology analysis shows that the response to unfolded protein is the most relevant. Figure S3. UPR mRNA expression in heart of infarcted transgenic mice. n= 12-13. Results expressed as mean ± SEM. Table S1: Altered gene expression due to NOX5-β expression at 12 h (bold, highlighted in yellow). N12hvsG12h N18hvsG18h N24hvsG24h GeneName GeneDescription TranscriptID logFC p-value logFC p-value logFC p-value family with sequence similarity NM_052966 1.45 1.20E-17 2.44 3.27E-19 2.96 6.24E-21 FAM129A 129. member A DnaJ (Hsp40) homolog. NM_001130182 2.19 9.83E-20 2.94 2.90E-19 3.01 1.68E-19 DNAJA4 subfamily A. member 4 phorbol-12-myristate-13-acetate- NM_021127 0.93 1.84E-12 2.41 1.32E-17 2.69 1.43E-18 PMAIP1 induced protein 1 E2F7 E2F transcription factor 7 NM_203394 0.71 8.35E-11 2.20 2.21E-17 2.48 1.84E-18 DnaJ (Hsp40) homolog. -
TBC1D8B Mutations Implicate RAB11-Dependent Vesicular Trafficking in the Pathogenesis of Nephrotic Syndrome
BASIC RESEARCH www.jasn.org TBC1D8B Mutations Implicate RAB11-Dependent Vesicular Trafficking in the Pathogenesis of Nephrotic Syndrome Lina L. Kampf,1 Ronen Schneider,2 Lea Gerstner,1 Roland Thünauer,3,4 Mengmeng Chen,1 Martin Helmstädter,1 Ali Amar,2 Ana C. Onuchic-Whitford,2,5 Reyner Loza Munarriz,6 Afig Berdeli,7 Dominik Müller,8 Eva Schrezenmeier,9 Klemens Budde,9 Shrikant Mane,10 Kristen M. Laricchia,11 Heidi L. Rehm ,11 Daniel G. MacArthur,11 Richard P. Lifton,10,12 Gerd Walz,1 Winfried Römer,3 Carsten Bergmann,13,14,15 Friedhelm Hildebrandt,2 and Tobias Hermle 1 Due to the number of contributing authors, the affiliations are listed at the end of this article. ABSTRACT Background Mutations in about 50 genes have been identified as monogenic causes of nephrotic syndrome, a frequent cause of CKD. These genes delineated the pathogenetic pathways and rendered significant insight into podocyte biology. Methods We used whole-exome sequencing to identify novel monogenic causes of steroid-resistant nephrotic syndrome (SRNS). We analyzed the functional significance of an SRNS-associated gene in vitro and in podocyte-like Drosophila nephrocytes. Results We identified hemizygous missense mutations in the gene TBC1D8B in five families with nephrotic syndrome. Coimmunoprecipitation assays indicated interactions between TBC1D8B and active forms of RAB11. Silencing TBC1D8B in HEK293T cells increased basal autophagy and exocytosis, two cellular functions that are independently regulated by RAB11. This suggests that TBC1D8B plays a regulatory role by inhibiting endogenous RAB11. Coimmunoprecipitation assays showed TBC1D8B also interacts with the slit diaphragm protein nephrin, and colocalizes with it in immortalized cell lines. -
University of Florida Thesis Or Dissertation Formatting
AAA ATPASE CDC48A AND ITS ASSOCIATION WITH UBIQUITIN-LIKE SAMP1 AND DNA REPAIR IN ARCHAEA By SWATHI DANTULURI A DISSERTATION PRESENTED TO THE GRADUATE SCHOOL OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY UNIVERSITY OF FLORIDA 2018 © 2018 Swathi Dantuluri I dedicate this to my family and friends for their endless love, support and company.Everybody who has taken the time and energy to teach me everything small or big. ACKNOWLEDGMENTS During my graduate career at the University of Florida, there are many people to whom I will be eternally indebted to for supporting and advising me along the way. Firstly, I would like to thank my advisor, Dr. Julie A. Maupin-Furlow for providing me the opportunity to work under her guidance as well as for all her help, advice, support, and encouragement throughout my time in her lab. I would also like to thank my committee members Dr. Sixue Chen, Dr.Tony Romeo and Dr. Nemat.O.Keyhani for their constructive advice and guidance. I must also express my gratitude to all the past and current Maupin lab members with whom I developed a warm and friendly working relationship over time, especially Dr. Nathaniel Hepowit who patiently taught me all the lab techniques, Dr. Xian Fu, Dr. Lana J. McMillan, Dr. Shiyun Cao, Dr. Sungmin Hwang, Zachary Adams, Cuper Ramirez, Paula Mondragon, and Jou Chin Chan. I also would like to thank all the undergraduate researchers in our lab for their dedication: Shae Marguiles, Sonam Parag, Gayathri Srinivasan, and Whinkie Leung. -
Small Gtpase Ran and Ran-Binding Proteins
BioMol Concepts, Vol. 3 (2012), pp. 307–318 • Copyright © by Walter de Gruyter • Berlin • Boston. DOI 10.1515/bmc-2011-0068 Review Small GTPase Ran and Ran-binding proteins Masahiro Nagai 1 and Yoshihiro Yoneda 1 – 3, * highly abundant and strongly conserved GTPase encoding ∼ 1 Biomolecular Dynamics Laboratory , Department a 25 kDa protein primarily located in the nucleus (2) . On of Frontier Biosciences, Graduate School of Frontier the one hand, as revealed by a substantial body of work, Biosciences, Osaka University, 1-3 Yamada-oka, Suita, Ran has been found to have widespread functions since Osaka 565-0871 , Japan its initial discovery. Like other small GTPases, Ran func- 2 Department of Biochemistry , Graduate School of Medicine, tions as a molecular switch by binding to either GTP or Osaka University, 2-2 Yamada-oka, Suita, Osaka 565-0871 , GDP. However, Ran possesses only weak GTPase activ- Japan ity, and several well-known ‘ Ran-binding proteins ’ aid in 3 Japan Science and Technology Agency , Core Research for the regulation of the GTPase cycle. Among such partner Evolutional Science and Technology, Osaka University, 1-3 molecules, RCC1 was originally identifi ed as a regulator of Yamada-oka, Suita, Osaka 565-0871 , Japan mitosis in tsBN2, a temperature-sensitive hamster cell line (3) ; RCC1 mediates the conversion of RanGDP to RanGTP * Corresponding author in the nucleus and is mainly associated with chromatin (4) e-mail: [email protected] through its interactions with histones H2A and H2B (5) . On the other hand, the GTP hydrolysis of Ran is stimulated by the Ran GTPase-activating protein (RanGAP) (6) , in con- Abstract junction with Ran-binding protein 1 (RanBP1) and/or the large nucleoporin Ran-binding protein 2 (RanBP2, also Like many other small GTPases, Ran functions in eukaryotic known as Nup358).