DOCSLIB.ORG

Probing the Structural Topology of HIV-1 Virion Infectivity Factor (VIF): a Dissertation

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Probing the Structural Topology of HIV-1 Virion Infectivity Factor (VIF): a Dissertation University of Massachusetts Medical School eScholarship@UMMS GSBS Dissertations and Theses Graduate School of Biomedical Sciences 2007-12-14 Probing the Structural Topology of HIV-1 Virion Infectivity Factor (VIF): A Dissertation Jared R. Auclair University of Massachusetts Medical School Let us know how access to this document benefits ou.y Follow this and additional works at: https://escholarship.umassmed.edu/gsbs_diss Part of the Life Sciences Commons, and the Medicine and Health Sciences Commons Repository Citation Auclair JR. (2007). Probing the Structural Topology of HIV-1 Virion Infectivity Factor (VIF): A Dissertation. GSBS Dissertations and Theses. Retrieved from https://escholarship.umassmed.edu/gsbs_diss/359 This material is brought to you by eScholarship@UMMS. It has been accepted for inclusion in GSBS Dissertations and Theses by an authorized administrator of eScholarship@UMMS. For more information, please contact [email protected]. PROBING THE STRUCTURAL TOPOLOGY OF HIV-1 VIRION INFECTIVITY FACTOR (VIF) A Dissertation Presented By Jared R. Auclair Submitted to the Faculty of the University of Massachusetts Graduate School of Biomedical Sciences, Worcester In partial fulfillment of the requirement for the degree of DOCTOR OF PHILOSPOHY December 14, 2007 Biochemistry and Molecular Pharmacology PROBING THE STRUCTURAL TOPOLOGY OF HIV-1 VIRION INFECTIVITY FACTOR (VIF) A Dissertation Presented by Jared R. Auclair The signatures of the Dissertation Defense Committee signifies completion and approval as to the style and content of the Dissertation. ________________________________________ Celia A. Schiffer Ph.D., Thesis Advisor ________________________________________ Mohan Somasundaran Ph.D., Thesis Advisor ________________________________________ Paul Clapham Ph.D., Member of Committee ________________________________________ Reuben Harris Ph.D., Member of Committee ________________________________________ C. Robert Matthews Ph.D., Member of Committee ________________________________________ Mary Munson Ph.D., Member of Committee The signature of the Chair of the Committee signifies that the written dissertation meets the requirements of the Dissertation Committee ________________________________________ Phillip Zamore Ph.D., Chair of Committee The signature of the Dean of the Graduate School of Biomedical Sciences signifies that the student has met all graduation requirements of the school. ________________________________________ Anthony Carruthers, Ph.D., Dean of the Graduate School Biomedical Sciences Interdisciplinary Graduate Program December 14, 2007 iii ACKNOWLEDGEMENTS I would like to acknowledge all those who have helped me on my path to completing my graduate work. I would like to start by thanking my thesis research advisory committee: Dr. Phillip Zamore, Dr. Mary Munson, Dr. Mario Stevenson, and Dr. Mohan Somasundaran for keeping me on track and helpful discussions over the years. In particular, I would like to thank Phil and Mary for always challenging me to ask the right questions, critically think about my hypothesis, problem solve, and to design experiments that would address the question I was asking. In addition, to my thesis research advisory committee I would like to thank my dissertation examination committee: Dr. Phillip Zamore, Dr. Mary Munson, Dr. Paul Clapham, Dr. C. Robert Matthews, and Dr. Reuben Harris. I especially thank Dr. Harris for making the trip from Minnesota to serve as my external examiner. Finally, I thank Dr. Sean Ryder for his helpful discussions on peptide competition experiments and agreeing to serve as a back- up DEC committee member. The Schiffer lab has been instrumental in my success over the last 6 and half years. The supportive environment of the lab is second to none. I truly enjoyed coming to work every day, it has been a great place to work and I will miss coming in each day. When I first joined the lab I was the fifth person and we were still on the 7th floor of the medical school, now we have around 15 people in the lab which is a sign of the supportive environment created by Dr. Schiffer. I would like to thank both Alex and Van, two undergraduate students, for there help with my project and giving me the opportunity to teach them. Thank you to Dr. Moses Prabu for critically reading drafts of this thesis iv and for always making lab fun and interesting. Thank you Madhavi Nalam for helping me map my cross-linking data onto the theoretical HIV-1 Vif models in Chapter I. In addition, I would like to thank Claire Baldwin for critically reading this thesis. Finally, thanks to Shiven Shandilya for discussions on HIV-1 Vif and his help in advancing the HIV-1 Vif project as a whole. A few people in the lab and Biochemistry department I would like to thank specifically. First, I would like to thank Tiffanie Covello for being a great friend, helping keep me organized, and just making my job easier in general. Karen Welch has been an amazing friend and extremely supportive over the last six plus years. She has always been there for me professional and personal; going to work without her around will not be the same. Along with many other things, I will miss Karen, Ellen, and I’s Weenie Wednesdays. I also cannot even begin to express my thanks to Ellen Nalivaika. She has always been one of my biggest supporters, and over the years we have become really great friends. There are few people in the world who understand and know me as well as she does and I’m truly grateful for that because without her, getting through graduate school would not have been as fun and it would have been much more difficult. I’ll be sending many emails with my brain-dead calculation questions! Of course, I have to thank all my great friends for their support and putting up with me and my “moods” over the last six years. Jenn Murzycki, a former labmate, has been a great friend and without her I could not have done this. Coffee breaks with Dianne Schwarz helped keep me sane over the years and she is one of my closest friends from graduate school. Thanks to Melonnie Furgason for listening to me complain, we v share a common bond of torturous projects, and for critical reading my thesis. Thanks to my bowling friends (Kapil, Evan, Dan, Mike, Jay, and Steve) without our weekly bowling league graduate school just would not have been the same. In addition to weekly bowling, Wednesday night get togethers with Jes, Chris, Crytal and Dave also helped keep me sane through this whole process. They are some of my oldest friends and I appreciate all the support they have given me; I would truly do anything for them. I do not even know where to begin in thanking both Dr. Celia Schiffer and Dr. Mohan Somasundaran. Celia has been an amazing mentor who has given me every opportunity to succeed. I’m most grateful for her support both scientifically and personally and faith in me. She has always given me all the tools I need to be successful and has taught and allowed me to guide my own research. She not only has been a great mentor but colleague and friend. Mohan is the person I have known the longest at UMass, and he is responsible for sparking an excitement in me for research during my senior year at WPI that persists to this day. Without his support and guidance I could not have done this! I not only consider him an amazing mentor, but an amazing person and friend! Finally, I would like to thank my family for without there love and support I could have never done this. My Mom and Dad have always been supportive of everything I do; even this crazy thing called graduate school. They have given me the tools and drive to be successful and I’m proud to be their son. I thank my sister, Heather, and my brother, Tyler, for all their support as well. I know that I have not always been the easiest person to get along with but regardless they have always been there for me. I looking forward to vi sharing many more of life’s adventures with them; and I’m always here for them no matter what. I also would like to thank my cousin Madeleine for all her support over the years; I could not have done this without her support as well. I thank my grandparent’s for their support as well; just knowing they are thinking of me helped me through some tough times. I also thank Mr., Mrs., and Adam Grabowski for their support in my quest to get a Ph.d.; and for raising an amazing woman, Melissa. I thank my wife, Melissa, for all her love and support over the years. You are my best friend and I am blessed to have you in my life. You know me better than anyone else, and I am so happy we were able to share this experience together. There is no doubt in my mind that I could not have done this without you. You are an amazing person and I love you with all my heart. I look forward to the next chapters in our life together and the experiences that lay ahead including the birth of our daughter, Madison. I cannot wait to meet you Madison and love you. Finally, as I finish these acknowledgements I remember all those family and friends who have died and take a moment to pause and remember them and thank them for helping make me the person I am today! vii ABSTRACT Human Immunodeficiency Virus Type 1 (HIV-1), the virus that causes Acquired Immunodeficiency Syndrome (AIDS), attacks the immune system leaving patients susceptible to opportunistic infections that eventually cause death. Highly Active Antiretroviral Therapy, HAART, is the current drug strategy used to combat HIV.
Load more

Recommended publications
  • RNA-Dependent RNA Polymerase Consensus Sequence of the L-A Double-Stranded RNA Virus: Definition of Essential Domains
    Proc. Nati. Acad. Sci. USA Vol. 89, pp. 2185-2189, March 1992 Biochemistry RNA-dependent RNA polymerase consensus sequence of the L-A double-stranded RNA virus: Definition of essential domains JUAN CARLOS RIBAS AND REED B. WICKNER Section on the Genetics of Simple Eukaryotes, Laboratory of Biochemical Pharmacology, National Institute of Diabetes and Digestive and Kidney Diseases, Building 8, Room 207, National Institutes of Health, Bethesda, MD 20892 Communicated by Herbert Tabor, November 27, 1991 (received for review October 2, 1991) ABSTRACT The L-A double-stranded RNA virus of Sac- lacking M1 (reviewed in refs. 10 and 18). M1 depends on L-A charomyces cerevisiac makes a gag-pol fusion protein by a -1 for its coat and replication proteins (19). MAK10 is one of ribosomal frameshift. The pol amino acid sequence includes three chromosomal genes needed for L-A virus propagation consensus patterns typical of the RNA-dependent RNA poly- within yeast cells (20). In a maklO host, L-A proteins merases (EC 2.7.7.48) of (+) strand and double-stranded RNA expressed from a cDNA clone of L-A support the replication viruses of animals and plants. We have carried out "alanine- of the M1 satellite virus but (for unknown reasons) do not scanning mutagenesis" of the region of L-A including the two support propagation of the L-A virus itself (21). Thus, while most conserved polymerase motifs, SG...T...NT..N (. = any L-A requires the MAK10 product itself, M1 requires MAK10 amino acid) and GDD. By constructing and analyzing 46 only because it requires the L-A-encoded proteins.
    [Show full text]
  • UC Berkeley UC Berkeley Electronic Theses and Dissertations
    UC Berkeley UC Berkeley Electronic Theses and Dissertations Title Mechanistic insights into the roles of P-TEFb and its novel cofactors in tumorigenesis and HIV transcription Permalink https://escholarship.org/uc/item/54n5v31q Author He, Nanhai Publication Date 2012 Peer reviewed|Thesis/dissertation eScholarship.org Powered by the California Digital Library University of California Mechanistic insights into the roles of P-TEFb and its novel cofactors in tumorigenesis and HIV transcription By Nanhai He A dissertation submitted in partial satisfaction of the requirements for the degree of Doctor of Philosophy in Molecular and Cell Biology in the Graduate Division of the University of California, Berkeley Committee in charge: Professor Qiang Zhou, Chair Professor Mike Botchan Professor Tom Abler Professor Fenyong Liu Spring 2012 1 Abstract Mechanistic insights into the roles of P-TEFb and its novel cofactors in tumorigenesis and HIV transcription by Nanhai He Doctor of Philosophy in Molecular and Cell Biology University of California, Berkeley Professor Qiang Zhou, Chair Ongoing research in the field of transcription has given rise to the unappreciated role of elongation control as a rate limiting step for transcription, and the general transcription elongation factor P-TEFb therefore has taken the central stages. P-TEFb is composed of cyclin T1 and the cyclin-dependent kinase Cdk9. It stimulates transcription elongation by releasing the paused RNA Polymerase II (Pol II) through phosphorylating Pol II at Ser2 and antagonizing the effects of negative elongation factors. P-TEFb is not only essential for the transcription of the vast majority of cellular genes, but also critical for the expression of HIV genome.
    [Show full text]
  • UC Santa Cruz UC Santa Cruz Electronic Theses and Dissertations
    UC Santa Cruz UC Santa Cruz Electronic Theses and Dissertations Title Structural Studies Of Tetrahymena Thermophila Telomerase Permalink https://escholarship.org/uc/item/06s5j7vf Author Loper, John Anderson Publication Date 2013 Peer reviewed|Thesis/dissertation eScholarship.org Powered by the California Digital Library University of California UNIVERSITY OF CALIFORNIA SANTA CRUZ STRUCTURAL STUDIES OF TETRAHYMENA THERMOPHILA TELOMERASE A thesis submitted in partial satisfaction of the requirements for the degree of MASTER OF SCIENCE in CHEMISTRY by John A. Loper June 2013 The Thesis of John A. Loper is approved: Assistant Professor Michael D. Stone, Chair Professor Harry F. Noller Associate Professor Seth M. Rubin Tyrus Miller Vice Provost and Dean of Graduate Studies Table of Contents Chapter 1. Introduction …………...……………………………………………..... 1 1.0 Telomeres ……………………………………………………………….. 2 1.1 Telomerase ………………………………………....…………………… 3 1.1.1 Telomerase RNA (TER) ……………………………...……… 4 1.1.2 Telomerase Reverse Transcriptase (TERT) ……...………… 5 1.2 Telomerase and Human Disease ………………………………….....… 6 1.3 Tetrahymena thermophila Telomerase ………………………................ 7 1.4 References ………………………………………….......................…… 11 Chapter 2. The C-terminal Domain of p65 is Required and Sufficient for TER Rearrangement and TERT Recruitment …………………………………....… 16 2.0 Introduction …………………………………………………..……… 17 2.0.1 p65 …………………………………...................…………… 17 2.0.2 FRET ………………………………................……………… 18 2.1 Results ………………………………………………………………… 20 iii 2.1.1 Role
    [Show full text]
  • The Yeast La Related Protein Slf1p Is a Key Activator of Translation During the Oxidative Stress Response
    The Yeast La Related Protein Slf1p Is a Key Activator of Translation during the Oxidative Stress Response Christopher J. Kershaw1, Joseph L. Costello1¤, Lydia M. Castelli1, David Talavera1, William Rowe1, Paul F. G. Sims2, Mark P. Ashe1, Simon J. Hubbard1, Graham D. Pavitt1, Chris M. Grant1* 1 Faculty of Life Sciences, The University of Manchester, Manchester, United Kingdom, 2 Faculty of Life Sciences, Manchester Institute of Biotechnology (MIB), University of Manchester, Manchester, United Kingdom Abstract The mechanisms by which RNA-binding proteins control the translation of subsets of mRNAs are not yet clear. Slf1p and Sro9p are atypical-La motif containing proteins which are members of a superfamily of RNA-binding proteins conserved in eukaryotes. RIP-Seq analysis of these two yeast proteins identified overlapping and distinct sets of mRNA targets, including highly translated mRNAs such as those encoding ribosomal proteins. In paralell, transcriptome analysis of slf1D and sro9D mutant strains indicated altered gene expression in similar functional classes of mRNAs following loss of each factor. The loss of SLF1 had a greater impact on the transcriptome, and in particular, revealed changes in genes involved in the oxidative stress response. slf1D cells are more sensitive to oxidants and RIP-Seq analysis of oxidatively stressed cells enriched Slf1p targets encoding antioxidants and other proteins required for oxidant tolerance. To quantify these effects at the protein level, we used label-free mass spectrometry to compare the proteomes of wild-type and slf1D strains following oxidative stress. This analysis identified several proteins which are normally induced in response to hydrogen peroxide, but where this increase is attenuated in the slf1D mutant.
    [Show full text]
  • Comparative Genomics and Evolution of Proteins Involved in RNA Metabolism Vivek Anantharaman, Eugene V
    Nucleic Acids Research, 2002, Vol. 30, No. 7 1427–1464 SURVEY AND SUMMARY Comparative genomics and evolution of proteins involved in RNA metabolism Vivek Anantharaman, Eugene V. Koonin* and L. Aravind National Center for Biotechnology Information, National Library of Medicine, 8600 Rockville Pike, Building 389, National Institutes of Health, Bethesda, MD 20894, USA Received November 1, 2001; Revised December 20, 2001; Accepted January 2, 2002 ABSTRACT eukaryote-specific RNA-binding domains. Subsequent, vast expansions of these domains mark the origin of RNA metabolism, broadly defined as the compen- alternative splicing in animals and probably in dium of all processes that involve RNA, including plants. In addition to the reconstruction of the evolu- transcription, processing and modification of tran- tionary history of RNA metabolism, this analysis scripts, translation, RNA degradation and its regula- produced numerous functional predictions, e.g. of tion, is the central and most evolutionarily conserved previously undetected enzymes of RNA modification. part of cell physiology. A comprehensive, genome- wide census of all enzymatic and non-enzymatic protein domains involved in RNA metabolism was INTRODUCTION conducted by using sequence profile analysis and All cells synthesize a vast array of RNAs, using DNA or RNA structural comparisons. Proteins related to RNA templates, through a nucleoside polymerization reaction catalyzed metabolism comprise from 3 to 11% of the complete by RNA polymerases (1). The mRNAs are templates for the protein repertoire in bacteria, archaea and eukary- ribosomal synthesis of proteins. Ribosomal RNAs are central otes, with the greatest fraction seen in parasitic to the functions of the ribosome, such as recognition and posi- bacteria with small genomes.
    [Show full text]
  • BMC Evolutionary Biology Biomed Central
    BMC Evolutionary Biology BioMed Central Research article Open Access Molecular phylogenetics and comparative modeling of HEN1, a methyltransferase involved in plant microRNA biogenesis Karolina L Tkaczuk1,2, Agnieszka Obarska1 and Janusz M Bujnicki*1,3 Address: 1Laboratory of Bioinformatics and Protein Engineering, International Institute of Molecular and Cell Biology, Trojdena 4, 02-109 Warsaw, Poland, 2Institute of Technical Biochemistry, Technical University of Lodz, Stefanowskiego 4/10, 90-924 Lodz, Poland and 3Institute of Molecular Biology and Biotechnology, Adam Mickiewicz University, Umultowska 89, 61-614 Poznan, Poland Email: Karolina L Tkaczuk - [email protected]; Agnieszka Obarska - [email protected]; Janusz M Bujnicki* - [email protected] * Corresponding author Published: 24 January 2006 Received: 09 November 2005 Accepted: 24 January 2006 BMC Evolutionary Biology 2006, 6:6 doi:10.1186/1471-2148-6-6 This article is available from: http://www.biomedcentral.com/1471-2148/6/6 © 2006 Tkaczuk et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Abstract Background: Recently, HEN1 protein from Arabidopsis thaliana was discovered as an essential enzyme in plant microRNA (miRNA) biogenesis. HEN1 transfers a methyl group from S- adenosylmethionine to the 2'-OH or 3'-OH group of the last nucleotide of miRNA/miRNA* duplexes produced by the nuclease Dicer. Previously it was found that HEN1 possesses a Rossmann-fold methyltransferase (RFM) domain and a long N-terminal extension including a putative double-stranded RNA-binding motif (DSRM).
    [Show full text]
  • Towards Deciphering the Principles Underlying an Mrna Recognition Code Alexander Serganov and Dinshaw J Patel
    Available online at www.sciencedirect.com Towards deciphering the principles underlying an mRNA recognition code Alexander Serganov and Dinshaw J Patel Messenger RNAs interact with a number of different molecules potential and opportunities for manipulation of gene that determine the fate of each transcript and contribute to the expression at the post-transcriptional level, many struc- overall pattern of gene expression. These interactions are tural biology groups have focused their ongoing research governed by specific mRNA signals, which in principle could efforts toward determination of structures that would represent a special mRNA recognition ‘code’. Both, small uncover the complex network of relationships between molecules and proteins demonstrate a diversity of mRNA mRNA and its partners, thereby contributing toward a binding modes often dependent on the structural context of the comprehensive understanding of the principles under- regions surrounding specific target sequences. In this review, lying a ‘mRNA recognition code’. we have highlighted recent structural studies that illustrate the diversity of recognition principles used by mRNA binders for Although the past decade of intensive research has pro- timely and specific targeting and processing of the message. vided us with molecular details of many interesting intermolecular interactions involving mRNA, the past Addresses two years have been especially informative, with over Structural Biology Program, Memorial Sloan-Kettering Cancer Center, 30 structures reported of complexes containing mRNA. New York, NY 10021, USA This review analyzes recently published structural data (spanning 2006–2007) on specific mRNA recognition Corresponding author: Serganov, Alexander ([email protected]) and Patel, Dinshaw J ([email protected]) events and complements excellent earlier reviews on protein–RNA [2–5] and metabolite–mRNA [6–8] recog- nition.
    [Show full text]
  • Ribonucleoprotein SS-B/La Belongs to a Protein Family with Consensus Sequences for RNA-Binding
    Volume 17 Number 6 1989 Nucleic Acids Research Voue1Iubr618 NcecAisRsac Ribonucleoprotein SS-B/La belongs to a protein family with consensus sequences for RNA-binding Edward K.L.Chan, Kevin F.Sullivan and Eng M.Tan W.M.Keck Autoimmune Disease Center, Department of Molecular and Experimental Medicine, Scripps Clinic and Research Foundation, 10666 North Torrey Pines Road, La Jolla, CA 92037, USA Received December 2, 1988; Revised and Accepted February 15, 1989 ABSTRACT Autoantibodies from systemic rheumatic disorders have become useful reagents in molecular biology. SS-B/La, a major target of autoantibodies in lupus and Sjogren's syndrome, has been identified as a 46 kDa protein component of a ribonucleoprotein (RNP) particle implicated in the maturation of RNA polymerase III transcripts. This report describes the complete sequences of human and bovine SS-B/La and the identification of RNA-binding protein consensus sequences RNP1 and RNP2 in the N-terminal region previously shown to be complexed with RNA in UV-crosslinking experiments. Segments of about 95 residues from the RNA-binding domain of SS-B/La and from 29 RNA-binding domains of several other proteins are analysed with respect to the frequency of amino acids and their hydrophobicity at each position. The data suggest that SS-B/La belongs to a large family of RNA-binding proteins which includes heterogeneous nuclear RNPs, nucleolin, mRNA polyadenylate binding protein, and small nuclear RNPs. INTRODUCTION SS-B/La is a ubiquitous protein transiently associated with RNA polymerase III transcripts (1-8). SS-B/La also binds to Ul RNA (9), an RNA polymerase II transcript, and to the vesicular stomatitis virus leader RNA (10).
    [Show full text]
  • WO 2010/066849 Al
    (12) INTERNATIONAL APPLICATION PUBLISHED UNDER THE PATENT COOPERATION TREATY (PCT) (19) World Intellectual Property Organization International Bureau (10) International Publication Number (43) International Publication Date 17 June 2010 (17.06.2010) WO 2010/066849 Al (51) International Patent Classification: (74) Common Representative: VIB VZW; Rijvisschestraat C12N 15/82 (2006.01) C07K 14/415 (2006.01) 120, B-9052 Gent (BE). C12N 15/10 (2006.01) (81) Designated States (unless otherwise indicated, for every (21) International Application Number: kind of national protection available): AE, AG, AL, AM, PCT/EP2009/066856 AO, AT, AU, AZ, BA, BB, BG, BH, BR, BW, BY, BZ, CA, CH, CL, CN, CO, CR, CU, CZ, DE, DK, DM, DO, (22) International Filing Date: DZ, EC, EE, EG, ES, FI, GB, GD, GE, GH, GM, GT, 10 December 2009 (10.12.2009) HN, HR, HU, ID, IL, IN, IS, JP, KE, KG, KM, KN, KP, (25) Filing Language: English KR, KZ, LA, LC, LK, LR, LS, LT, LU, LY, MA, MD, ME, MG, MK, MN, MW, MX, MY, MZ, NA, NG, NI, (26) Publication Language: English NO, NZ, OM, PE, PG, PH, PL, PT, RO, RS, RU, SC, SD, (30) Priority Data: SE, SG, SK, SL, SM, ST, SV, SY, TJ, TM, TN, TR, TT, 0817 1187. 1 10 December 2008 (10.12.2008) EP TZ, UA, UG, US, UZ, VC, VN, ZA, ZM, ZW. (71) Applicants (for all designated States except US): VIB (84) Designated States (unless otherwise indicated, for every VZW [BE/BE]; Rijvisschestraat 120, B-9052 Gent (BE). kind of regional protection available): ARIPO (BW, GH, UNIVERSITEIT GENT [BE/BE]; Sint-Pietersnieuw- GM, KE, LS, MW, MZ, NA, SD, SL, SZ, TZ, UG, ZM, straat 25, B-9000 Gent (BE).
    [Show full text]
  • A Bona Fide La Protein Is Required for Embryogenesis in Arabidopsis
    3306–3321 Nucleic Acids Research, 2007, Vol. 35, No. 10 Published online 25 April 2007 doi:10.1093/nar/gkm200 A bona fide La protein is required for embryogenesis in Arabidopsis thaliana Sophie Fleurde´ pine1, Jean-Marc Deragon1,2, Martine Devic2, Jocelyne Guilleminot2 and Ce´ cile Bousquet-Antonelli1,* 1CNRS UMR6547 GEEM, Universite´ Blaise Pascal, 63177 Aubie` re, France and 2CNRS UMR5096 LGDP, Universite´ de Perpignan Via Domitia, 66860 Perpignan, France Received March 7, 2007; Revised March 21, 2007; Accepted March 21, 2007 ABSTRACT protein involved in many aspects of RNA metabolism (3–5) and is present in a wide range of eukaryotes Searches in the Arabidopsis thaliana genome using including budding and fission yeasts, vertebrates, insects, the La motif as query revealed the presence of worm (5) and trypanosome (6). The La protein is one of Downloaded from eight La or La-like proteins. Using structural and the first proteins to bind to primary polymerase III phylogenetic criteria, we identified two putative (pol III) transcripts due to the specific recognition of the genuine La proteins (At32 and At79) and showed 30-UUU-OH motif present in these precursors (7). The that both are expressed throughout plant develop- Saccharomyces cerevisiae La protein (named Lhp1p) also ment but at different levels and under different binds polymerase II (pol II) transcribed small RNAs that http://nar.oxfordjournals.org/ regulatory conditions. At32, but not At79, restores terminate in 30-UUU-OH such as precursors to the U3 Saccharomyces cerevisiae La nuclear functions in snoRNA (small nucleolar RNA) or U snRNAs (small non-coding RNAs biogenesis and is able to bind to nuclear RNA) (8–10).
    [Show full text]
  • An Intrinsically Disordered C Terminus Allows the La Protein to Assist the Biogenesis of Diverse Noncoding RNA Precursors
    An intrinsically disordered C terminus allows the La protein to assist the biogenesis of diverse noncoding RNA precursors Nathan J. Kuceraa, Michael E. Hodsdonb, and Sandra L. Wolina,c,1 aDepartment of Cell Biology, Yale University School of Medicine, New Haven, CT 06510; bDepartment of Laboratory Medicine, Yale University School of Medicine, New Haven, CT 06510; and cDepartment of Molecular Biophysics and Biochemistry, Yale University School of Medicine, New Haven, CT 06510 Edited by Jennifer A. Doudna, University of California, Berkeley, CA, and approved December 6, 2010 (received for review November 15, 2010) The La protein binds the 3′ ends of many newly synthesized non- and fission yeast, La becomes essential in the presence of muta- coding RNAs, protecting these RNAs from nucleases and influencing tions that compromise the structure of essential pre-tRNAs or folding, maturation, and ribonucleoprotein assembly. Although 3′ impair snRNP assembly (4, 5). While many of the mutations cause end binding by La involves the N-terminal La domain and adjacent the affected RNA to be degraded without La, Saccharomyces RNA recognition motif (RRM), the mechanisms by which La stabi- cerevisiae Arg La is required for correct folding of pre-tRNACCG when lizes diverse RNAs from nucleases and assists subsequent events the tRNA carries a mutation that weakens the anticodon stem, in their biogenesis are unknown. Here we report that a conserved allowing formation of an incorrect helix. In the absence of La, feature of La proteins, an intrinsically disordered C terminus, is re- the mature tRNA accumulates but is not aminoacylated (11). quired for the accumulation of certain noncoding RNA precursors The role of La in assisting pre-tRNA folding may be general, as and for the role of the Saccharomyces cerevisiae La protein Lhp1p La is required for efficient charging of two wild-type tRNAs in assisting formation of correctly folded pre-tRNA anticodon stems when yeast are grown at low temperature (11).
    [Show full text]
  • Binding Specificities of Human RNA-Binding Proteins Toward Structured and Linear RNA Sequences
    Downloaded from genome.cshlp.org on September 29, 2021 - Published by Cold Spring Harbor Laboratory Press Research Binding specificities of human RNA-binding proteins toward structured and linear RNA sequences Arttu Jolma,1,11 Jilin Zhang,1,11 Estefania Mondragón,2,11 Ekaterina Morgunova,1 Teemu Kivioja,3 Kaitlin U. Laverty,4 Yimeng Yin,1 Fangjie Zhu,1 Gleb Bourenkov,5 Quaid Morris,4,6,7,8,9 Timothy R. Hughes,4,6 Louis James Maher III,2 and Jussi Taipale1,3,10 1Department of Medical Biochemistry and Biophysics, Karolinska Institutet, SE-171 77, Solna, Sweden; 2Department of Biochemistry and Molecular Biology, Mayo Clinic Graduate School of Biomedical Sciences, Mayo Clinic College of Medicine and Science, Rochester, Minnesota 55905, USA; 3Genome-Scale Biology Program, University of Helsinki, FI-00014, Helsinki, Finland; 4Department of Molecular Genetics, University of Toronto, M5S 1A8, Toronto, Canada; 5European Molecular Biology Laboratory (EMBL), Hamburg Unit c/o DESY, D-22603 Hamburg, Germany; 6Donnelly Centre, University of Toronto, M5S 3E1, Toronto, Canada; 7Edward S. Rogers Sr. Department of Electrical and Computer Engineering, University of Toronto, M5S 3G4, Toronto, Canada; 8Department of Computer Science, University of Toronto, M5S 2E4, Toronto, Canada; 9Memorial Sloan Kettering Cancer Center, New York, New York 10065, USA; 10Department of Biochemistry, University of Cambridge, CB2 1QW, Cambridge, United Kingdom RNA-binding proteins (RBPs) regulate RNA metabolism at multiple levels by affecting splicing of nascent transcripts, RNA folding, base modification, transport, localization, translation, and stability. Despite their central role in RNA function, the RNA-binding specificities of most RBPs remain unknown or incompletely defined. To address this, we have assembled a genome-scale collection of RBPs and their RNA-binding domains (RBDs) and assessed their specificities using high-through- put RNA-SELEX (HTR-SELEX).
    [Show full text]