Mouse Piwi Interactome Identifies Binding Mechanism of Tdrkh Tudor Domain to Arginine Methylated Miwi

Total Page:16

File Type:pdf, Size:1020Kb

Mouse Piwi Interactome Identifies Binding Mechanism of Tdrkh Tudor Domain to Arginine Methylated Miwi Mouse Piwi interactome identifies binding mechanism of Tdrkh Tudor domain to arginine methylated Miwi Chen Chena, Jing Jina, D. Andrew Jamesa, Melanie A. Adams-Cioabab, Jin Gyoon Parka, Yahong Guob, Enrico Tenagliaa,c, Chao Xub, Gerald Gisha, Jinrong Minb,d, and Tony Pawsona,c,1 aSamuel Lunenfeld Research Institute, Mount Sinai Hospital, Toronto, ON, Canada M5G 1X5; bStructural Genomics Consortium, University of Toronto, Toronto, ON, Canada M5G 1L6; and cDepartment of Molecular Genetics and dDepartment of Physiology, University of Toronto, Toronto, ON, Canada M5S 1A8 Contributed by Tony Pawson, October 8, 2009 (sent for review September 15, 2009) Tudor domains are protein modules that mediate protein–protein spermatogenesis, and small RNA pathways (7–9). However, the interactions, potentially by binding to methylated ligands. A group of binding properties of these germline Tudor proteins are poorly germline specific single and multiTudor domain containing proteins understood. (TDRDs) represented by drosophila Tudor and its mammalian or- Piwi proteins are conserved germline-specific Argonaute fam- thologs Tdrd1, Tdrd4/RNF17, and Tdrd6 play evolutionarily conserved ily members that are associated with Piwi-interacting RNAs roles in germinal granule/nuage formation and germ cell specification (piRNAs), and thereby function in piRNA-mediated posttranscrip- and differentiation. However, their physiological ligands, and the tional silencing (10). Three murine Piwi paralogs, Miwi, Mili, and biochemical and structural basis for ligand recognition, are largely Miwi2, play pivotal roles in germ cell development, transposon unclear. Here, by immunoprecipitation of endogenous murine Piwi silencing and spermatogenesis (11–13). The presence of multiple proteins (Miwi and Mili) and proteomic analysis of complexes related arginine-glycine and arginine-alanine (RG/RA)-rich clusters at the to the piRNA pathway, we show that the TDRD group of Tudor N-termini of these proteins prompted us to question whether these proteins are physiological binding partners of Piwi family proteins. In RG/RA motifs can be methylated in vivo and thereby serve as addition, mass spectrometry indicates that arginine residues in RG docking sites for the binding of various germline Tudor proteins. repeats at the N-termini of Miwi and Mili are methylated in vivo. To test this hypothesis, we performed a comprehensive pro- Notably, we found that Tdrkh/Tdrd2, a novel single Tudor domain teomic analysis of Miwi and Mili complexes in adult male germ cells CELL BIOLOGY containing protein identified in the Miwi complex, is expressed in the and determined the methylation status of these Piwi proteins. We cytoplasm of male germ cells and directly associates with Miwi. show that several germline Tudor proteins are physiological binding Mutagenesis studies mapped the Miwi–Tdrkh interaction to the very partners of the Piwi family. In particular, we identify Tdrkh as a N-terminal RG/RA repeats of Miwi and showed that the Tdrkh Tudor novel Miwi-interacting protein that binds Miwi through its single domain is critical for binding. Furthermore, we have solved the crystal Tudor domain, likely via arginine methylation, as suggested by a structure of the Tdrkh Tudor domain, which revealed an aromatic combination of mass spectrometry, mutagenesis, and structural binding pocket and negatively charged binding surface appropriate analysis. for accommodating methylated arginine. Our findings identify a methylation-directed protein interaction mechanism in germ cells Results mediated by germline Tudor domains and methylated Piwi family Tudor Domain-Containing Proteins Are Major Physiological Binding proteins, and suggest a complex mode of regulating the organization Partners of Piwi Family Proteins. To test whether Tudor domain and function of Piwi proteins in piRNA silencing pathways. family proteins comprise the in vivo binding partners of the Piwi proteins, we immunoprecipitated endogenous Miwi and Mili from udor domains, together with Chromo, MBT, PWWP, and lysates of adult testes and purified the complexes by acid elution. To TAgenet-like domains, comprise the ‘‘Tudor Royal Family’’ of obtain a comprehensive survey of the components of the Piwi domains (1). The core structure of this protein domain superfamily complexes, we used a gel-free liquid chromatography coupled is characterized by an antiparallel ␤-barrel-like topology and me- tandem mass spectrometry (LC-MS/MS) approach employing solid diates protein–protein interactions, in some cases by recognizing phase tryptic digestion. This technique allowed us to unambigu- methylated lysine/arginine-containing ligands with a binding site ously identify an extensive list of candidate proteins that specifically composed of aromatic residues (2, 3). Their methylated target associated with Piwi proteins (Fig. 1). Hierarchical clustering of 2 proteins are implicated in diverse biological processes such as independent repeats of Miwi and Mili immunoprecipitations (IP) chromatin remodeling and RNA splicing. For example, the Tudor with their respective IgG control IPs reproducibly revealed distinct domain of Smn binds to methylated arginine-glycine (RG) motifs protein complex profiles for Miwi and Mili (Fig. 1). We found on Sm proteins essential for spliceosome assembly (4), while the proteins that were specifically associated with either Miwi (Fig. 1A, Tudor domains of Jmjd2a bind to methylated lysines in histone blue box and Fig. S2) or Mili (Fig. 1A, red box), and proteins shared H4K20 (5). by both Miwi and Mili complexes (Fig. 1A, cyan box). All of these Drosophila Tudor, the founding member of the Tudor domain proteins were absent from the IgG control IPs, which contain family, is a germ cell-specific protein with multiple Tudor domains nonspecific binding proteins (Fig. S2). In this analysis, we observed and is involved in germ plasm formation and germ cell specification several previously known Piwi-interacting proteins or piRNA path- (6). By analyzing the expression pattern of mammalian genes encoding Tudor domain proteins, we identified a group whose Author contributions: C.C., J.J., and T.P. designed research; C.C., J.J., D.A.J., M.A.A.-C., Y.G., expression is highly enriched in germ cells, which we therefore term E.T., C.X., and G.G. performed research; C.C., J.G.P., and J.M. analyzed data; and C.C. and germline Tudor proteins (Tdrd1, Tdrkh/Tdrd2, RNF17/Tdrd4, T.P. wrote the paper. Tdrd5, Tdrd6, Tdrd7, Stk31/Tdrd8, Tdrd9, Tdrd10, Akap1) (sup- The authors declare no conflict of interest. porting information (SI) Fig. S1). While the physiological functions Data deposition: The atomic coordinates have been deposited in the Protein Data Bank, of germline proteins with a single Tudor domain (Tdrkh, Tdrd5, www.pdb.org (PDB ID code 3fdr). Stk31, and Tdrd9) are largely unknown, mouse knockout studies of 1To whom correspondence should be addressed. E-mail: [email protected]. Tdrd1, Tdrd4, and Tdrd6 have revealed crucial roles for these This article contains supporting information online at www.pnas.org/cgi/content/full/ multiTudor domain proteins in nuage/chromatoid body formation, 0911640106/DCSupplemental. www.pnas.org͞cgi͞doi͞10.1073͞pnas.0911640106 PNAS Early Edition ͉ 1of6 Downloaded by guest on September 29, 2021 AB Ddx1 Kctd3 Pacsin1 1700020L24Rik Igf2r Golga5 Sucla2 Miwi_IP_2 Mili_IP_2 IgG_IP_(Mili1) IgG_IP_(Mili2) Mili_IP_1 Miwi_IP_1 IgG_IP_(Miwi 1) IgG_IP_(Miwi 2) Trpd52l3 1110012J17Rik 100046745 Kng1 Cep72 1700020L24Rik Ppp2r1a Pacsin2 Cul9 Pacsin2 Ddx1 Ilf3 Psmd2 Sccpdh Tdrd7 OTTMUSG00000023442 8030462N17Rik Stk22s1 Rai14 Rnf213 C330043M08Rik Cep72 Tdrd6 Git1 Stk31 ENSMUSG00000057808 Atp9a ENSMUSG00000076684 Hnrnpl Spata20 8030462N17Rik Stk22s1 Tpr Hdac6 Rai14 Sept2 Rab3ip B230339M05Rik Hsp90ab1 Tjp1 Birc6 Sept9 ENSMUSG00000076684 Ccdc6 Tdrkh Stk31 Kif17 Tjp1 Ccdc6 Golga5 Birc6 Kif17 Pgp Hsp90b1 Ssb Rnf213 Rpl5 1110012J17Rik Acta1 Tpr Rps4y2 Isyna1 Eef1g Hnrnpf Rps9 Hsp90ab1 Lta4h Igf2r Ilf3 Ppp2r1a Akap4 Tdrd6 Amotl1 Atp9a Phldb1 Spata20 Kctd3 Myst3 Pacsin1 Trpd52l3 Sept9 Phldb1 Pkd1l1 Hnrnpl OTTMUSG00000023442 Vps13d Miwi Ybx2 Ahsg ENSMUSG00000076502 C330043M08Rik Fcgr4 Krt5 Psmd2 Hdac6 Rps9 Sept7 Eef1a1 Tdrkh C3 Igh-VX24 Pcca LOC100046359 ENSMUSG00000076718 Hsp90aa1 Igh-6 Dbt Hpx Amotl1 ENSMUSG00000076683 LOC675759 Git1 ENSMUSG00000076939 Igh-VJ558 Fgb Sept2 Hrg Gc Igk-C ENSMUSG00000076709 Pkd1l1 Pzp Ddx4 Aldh1a1 Fgg Sccpdh Mug1 Myst3 Tubb3 Eef1g OTTMUSG00000015054 Eef2 ENSMUSG00000076665 Rpl5 Atp5b ENSMUSP00000112082 Keap1 Ldhc Ttn Dhx9 B230339M05Rik Khsrp Akap4 Trim21 Myo18a Rassf2 Ryr1 Acaa2 Srrm2 Nasp Wdr37 Psmc3 Tns1 Fxr1 ENSMUSG00000057808 Golga3 Nucb1 Ddx5 Tln1 Vim Rps7 1700012A16Rik Sucla2 Ruvbl2 Pbxip1 Hnrnpf Cct8 Hspa8 Tdrd7 Cct2 Dync1h1 Acta1 Rpl23 Cct5 Crybg3 Rab3ip Ddx4 Cct6a Cul9 Trim28 Dnpep Fth1 Hnrnpc Mtap4 Pcbp1 Hspa4l 4932438A13Rik Suclg1 Snrpd2 Tdrd1 Rps13 Rpl28 Rps14 ENSMUSG00000074479 Prdx1 C1qa Cct4 Slc25a31 Tcp1 Hnrnpu ENSMUSG00000076557 Mov10l1 ENSMUSG00000076652 LOC100041230 Blzf1 ENSMUSG00000076692 ENSMUSG00000076691 Rpl8 ENSMUSG00000073028 ENSMUSG00000076649 ENSMUSG00000076695 ENSMUSG00000076736 LOC630322 Fip1l1 ENSMUSG00000076740 Hnrnpm Matr3 Hspa9 Vcp Fga Rps5 Hnrnpa1 Hadha Mocs1 Trf Ywhaz Rps3 Vasp Cct3 Rps20 Rps25 Rpl18 Rps18 Napa Rps26 Mili 1700019E19Rik Zfp219 Fip1l1 Fyttd1 Enthd1 Gramd3 Gbf1 Snrpn Atp5a1 4932438A13Rik Fyttd1 Caprin1 G3bp1 Vasp Nucb1 Tns1 Sorbs2 Blzf1 Cul3 Eml4 Txndc2 Sssca1 Mov10l1 Txndc2 Sssca1 Enthd1 Mocs1 Snrpd2 Crybg3 G3bp1 Pbxip1 Napa 1700019E19Rik Wdhd1 Gramd1a Gas2l1
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]
  • Protein Kinase A-Mediated Septin7 Phosphorylation Disrupts Septin Filaments and Ciliogenesis
    cells Article Protein Kinase A-Mediated Septin7 Phosphorylation Disrupts Septin Filaments and Ciliogenesis Han-Yu Wang 1,2, Chun-Hsiang Lin 1, Yi-Ru Shen 1, Ting-Yu Chen 2,3, Chia-Yih Wang 2,3,* and Pao-Lin Kuo 1,2,4,* 1 Department of Obstetrics and Gynecology, College of Medicine, National Cheng Kung University, Tainan 701, Taiwan; [email protected] (H.-Y.W.); [email protected] (C.-H.L.); [email protected] (Y.-R.S.) 2 Institute of Basic Medical Sciences, College of Medicine, National Cheng Kung University, Tainan 701, Taiwan; [email protected] 3 Department of Cell Biology and Anatomy, College of Medicine, National Cheng Kung University, Tainan 701, Taiwan 4 Department of Obstetrics and Gynecology, National Cheng-Kung University Hospital, Tainan 704, Taiwan * Correspondence: [email protected] (C.-Y.W.); [email protected] (P.-L.K.); Tel.: +886-6-2353535 (ext. 5338); (C.-Y.W.)+886-6-2353535 (ext. 5262) (P.-L.K.) Abstract: Septins are GTP-binding proteins that form heteromeric filaments for proper cell growth and migration. Among the septins, septin7 (SEPT7) is an important component of all septin filaments. Here we show that protein kinase A (PKA) phosphorylates SEPT7 at Thr197, thus disrupting septin filament dynamics and ciliogenesis. The Thr197 residue of SEPT7, a PKA phosphorylating site, was conserved among different species. Treatment with cAMP or overexpression of PKA catalytic subunit (PKACA2) induced SEPT7 phosphorylation, followed by disruption of septin filament formation. Constitutive phosphorylation of SEPT7 at Thr197 reduced SEPT7-SEPT7 interaction, but did not affect SEPT7-SEPT6-SEPT2 or SEPT4 interaction.
    [Show full text]
  • Supplementary Table 3: Genes Only Influenced By
    Supplementary Table 3: Genes only influenced by X10 Illumina ID Gene ID Entrez Gene Name Fold change compared to vehicle 1810058M03RIK -1.104 2210008F06RIK 1.090 2310005E10RIK -1.175 2610016F04RIK 1.081 2610029K11RIK 1.130 381484 Gm5150 predicted gene 5150 -1.230 4833425P12RIK -1.127 4933412E12RIK -1.333 6030458P06RIK -1.131 6430550H21RIK 1.073 6530401D06RIK 1.229 9030607L17RIK -1.122 A330043C08RIK 1.113 A330043L12 1.054 A530092L01RIK -1.069 A630054D14 1.072 A630097D09RIK -1.102 AA409316 FAM83H family with sequence similarity 83, member H 1.142 AAAS AAAS achalasia, adrenocortical insufficiency, alacrimia 1.144 ACADL ACADL acyl-CoA dehydrogenase, long chain -1.135 ACOT1 ACOT1 acyl-CoA thioesterase 1 -1.191 ADAMTSL5 ADAMTSL5 ADAMTS-like 5 1.210 AFG3L2 AFG3L2 AFG3 ATPase family gene 3-like 2 (S. cerevisiae) 1.212 AI256775 RFESD Rieske (Fe-S) domain containing 1.134 Lipo1 (includes AI747699 others) lipase, member O2 -1.083 AKAP8L AKAP8L A kinase (PRKA) anchor protein 8-like -1.263 AKR7A5 -1.225 AMBP AMBP alpha-1-microglobulin/bikunin precursor 1.074 ANAPC2 ANAPC2 anaphase promoting complex subunit 2 -1.134 ANKRD1 ANKRD1 ankyrin repeat domain 1 (cardiac muscle) 1.314 APOA1 APOA1 apolipoprotein A-I -1.086 ARHGAP26 ARHGAP26 Rho GTPase activating protein 26 -1.083 ARL5A ARL5A ADP-ribosylation factor-like 5A -1.212 ARMC3 ARMC3 armadillo repeat containing 3 -1.077 ARPC5 ARPC5 actin related protein 2/3 complex, subunit 5, 16kDa -1.190 activating transcription factor 4 (tax-responsive enhancer element ATF4 ATF4 B67) 1.481 AU014645 NCBP1 nuclear cap
    [Show full text]
  • Structure and Expression Analyses of SVA Elements in Relation to Functional Genes
    pISSN 1598-866X eISSN 2234-0742 Genomics Inform 2013;11(3):142-148 G&I Genomics & Informatics http://dx.doi.org/10.5808/GI.2013.11.3.142 ORIGINAL ARTICLE Structure and Expression Analyses of SVA Elements in Relation to Functional Genes Yun-Jeong Kwon, Yuri Choi, Jungwoo Eo, Yu-Na Noh, Jeong-An Gim, Yi-Deun Jung, Ja-Rang Lee, Heui-Soo Kim* Department of Biological Sciences, College of Natural Sciences, Pusan National University, Busan 609-735, Korea SINE-VNTR-Alu (SVA) elements are present in hominoid primates and are divided into 6 subfamilies (SVA-A to SVA-F) and active in the human population. Using a bioinformatic tool, 22 SVA element-associated genes are identified in the human genome. In an analysis of genomic structure, SVA elements are detected in the 5′ untranslated region (UTR) of HGSNAT (SVA-B), MRGPRX3 (SVA-D), HYAL1 (SVA-F), TCHH (SVA-F), and ATXN2L (SVA-F) genes, while some elements are observed in the 3′UTR of SPICE1 (SVA-B), TDRKH (SVA-C), GOSR1 (SVA-D), BBS5 (SVA-D), NEK5 (SVA-D), ABHD2 (SVA-F), C1QTNF7 (SVA-F), ORC6L (SVA-F), TMEM69 (SVA-F), and CCDC137 (SVA-F) genes. They could contribute to exon extension or supplying poly A signals. LEPR (SVA-C), ALOX5 (SVA-D), PDS5B (SVA-D), and ABCA10 (SVA-F) genes also showed alternative transcripts by SVA exonization events. Dominant expression of HYAL1_SVA appeared in lung tissues, while HYAL1_noSVA showed ubiquitous expression in various human tissues. Expression of both transcripts (TDRKH_SVA and TDRKH_noSVA) of the TDRKH gene appeared to be ubiquitous.
    [Show full text]
  • Two DNA Binding Domains of Mga Act in Combination to Suppress Ectopic Activation
    bioRxiv preprint doi: https://doi.org/10.1101/2020.07.21.215079; this version posted July 22, 2020. 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. Two DNA binding domains of Mga act in combination to suppress ectopic activation of meiosis-related genes in mouse embryonic stem cells Kousuke Uranishi1, Masataka Hirasaki2, Yuka Kitamura1, Yosuke Mizuno3, Masazumi Nishimoto1,3, Ayumu Suzuki1*, Akihiko Okuda1* 1Division of Biomedical Sciences, Research Center for Genomic Medicine, 2Department of Clinical Cancer Genomics, International Medical Center, 3Biomedical Research Center, Saitama Medical University, 1397-1, Yamane Hidaka, Saitama 350-1241, Japan *Corresponding authors Ayumu Suzuki: [email protected] Akihiko Okuda: [email protected] Running title: Roles of two DNA binding domains of Mga in ESCs AUTHOR CONTRIBUTIONS K.U.: performed the experiments, data analysis, and manuscript writing; Y.K., M.H., and Y.M.: performed the experiments and data analysis; M.N.: performed the data analysis, and commented on the manuscript; A.S.: performed the experiments and data analysis, conception and design; A.O.: conception and design, financial support, manuscript writing, and final approval of manuscript. 1 bioRxiv preprint doi: https://doi.org/10.1101/2020.07.21.215079; this version posted July 22, 2020. 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. SUMMARY Mouse embryonic stem cells (ESCs) have high potential for meiotic entry, like germ cells. Although the physiological meaning of this potential is not known, it is certain that a rigid safeguarding system is required to prevent ectopic onset of meiosis.
    [Show full text]
  • Mouse Piwi Interactome Identifies Binding Mechanism of Tdrkh Tudor Domain to Arginine Methylated Miwi
    Mouse Piwi interactome identifies binding mechanism of Tdrkh Tudor domain to arginine methylated Miwi Chen Chena, Jing Jina, D. Andrew Jamesa, Melanie A. Adams-Cioabab, Jin Gyoon Parka, Yahong Guob, Enrico Tenagliaa,c, Chao Xub, Gerald Gisha, Jinrong Minb,d, and Tony Pawsona,c,1 aSamuel Lunenfeld Research Institute, Mount Sinai Hospital, Toronto, ON, Canada M5G 1X5; bStructural Genomics Consortium, University of Toronto, Toronto, ON, Canada M5G 1L6; and cDepartment of Molecular Genetics and dDepartment of Physiology, University of Toronto, Toronto, ON, Canada M5S 1A8 Contributed by Tony Pawson, October 8, 2009 (sent for review September 15, 2009) Tudor domains are protein modules that mediate protein–protein spermatogenesis, and small RNA pathways (7–9). However, the interactions, potentially by binding to methylated ligands. A group of binding properties of these germline Tudor proteins are poorly germline specific single and multiTudor domain containing proteins understood. (TDRDs) represented by drosophila Tudor and its mammalian or- Piwi proteins are conserved germline-specific Argonaute fam- thologs Tdrd1, Tdrd4/RNF17, and Tdrd6 play evolutionarily conserved ily members that are associated with Piwi-interacting RNAs roles in germinal granule/nuage formation and germ cell specification (piRNAs), and thereby function in piRNA-mediated posttranscrip- and differentiation. However, their physiological ligands, and the tional silencing (10). Three murine Piwi paralogs, Miwi, Mili, and biochemical and structural basis for ligand recognition, are largely Miwi2, play pivotal roles in germ cell development, transposon unclear. Here, by immunoprecipitation of endogenous murine Piwi silencing and spermatogenesis (11–13). The presence of multiple proteins (Miwi and Mili) and proteomic analysis of complexes related arginine-glycine and arginine-alanine (RG/RA)-rich clusters at the to the piRNA pathway, we show that the TDRD group of Tudor N-termini of these proteins prompted us to question whether these proteins are physiological binding partners of Piwi family proteins.
    [Show full text]
  • A Genome-Wide Association Study of Idiopathic Dilated Cardiomyopathy in African Americans
    Journal of Personalized Medicine Article A Genome-Wide Association Study of Idiopathic Dilated Cardiomyopathy in African Americans Huichun Xu 1,* ID , Gerald W. Dorn II 2, Amol Shetty 3, Ankita Parihar 1, Tushar Dave 1, Shawn W. Robinson 4, Stephen S. Gottlieb 4 ID , Mark P. Donahue 5, Gordon F. Tomaselli 6, William E. Kraus 5,7 ID , Braxton D. Mitchell 1,8 and Stephen B. Liggett 9,* 1 Division of Endocrinology, Diabetes and Nutrition, Department of Medicine, University of Maryland School of Medicine, Baltimore, MD 21201, USA; [email protected] (A.P.); [email protected] (T.D.); [email protected] (B.D.M.) 2 Center for Pharmacogenomics, Department of Internal Medicine, Washington University School of Medicine, St. Louis, MO 63110, USA; [email protected] 3 Institute for Genome Sciences, University of Maryland School of Medicine, Baltimore, MD 21201, USA; [email protected] 4 Division of Cardiovascular Medicine, University of Maryland School of Medicine, Baltimore, MD 21201, USA; [email protected] (S.W.R.); [email protected] (S.S.G.) 5 Division of Cardiology, Department of Medicine, Duke University Medical Center, Durham, NC 27708, USA; [email protected] (M.P.D.); [email protected] (W.E.K.) 6 Department of Medicine, Division of Cardiology, Johns Hopkins University, Baltimore, MD 21218, USA; [email protected] 7 Duke Molecular Physiology Institute, Duke University Medical Center, Durham, NC 27701, USA 8 Geriatrics Research and Education Clinical Center, Baltimore Veterans Administration
    [Show full text]
  • 1 DMA-Tudor Interaction Modules Control the Specificity of In
    bioRxiv preprint doi: https://doi.org/10.1101/2020.09.15.297994; this version posted September 16, 2020. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-ND 4.0 International license. DMA-tudor interaction modules control the specificity of in vivo condensates Edward M. Courchaine1, Andrew E.S. Barentine2,3, Korinna Straube1, Joerg Bewersdorf2,3, Karla M. Neugebauer*1,2 5 1Molecular Biophysics and Biochemistry, Yale University, New Haven, CT 2Cell Biology, Yale University, New Haven, CT 3Biomedical Engineering, Yale University, New Haven, CT 10 *Correspondence to: [email protected] 1 bioRxiv preprint doi: https://doi.org/10.1101/2020.09.15.297994; this version posted September 16, 2020. The copyright holder for this preprint (which was not certified by peer review) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under aCC-BY-ND 4.0 International license. Summary: Biomolecular condensation is a widespread mechanism of cellular compartmentalization. Because the ‘survival of motor neuron protein’ (SMN) is required for the formation of three different 15 membraneless organelles (MLOs), we hypothesized that at least one region of SMN employs a unifying mechanism of condensation. Unexpectedly, we show here that SMN’s globular tudor domain was sufficient for dimerization-induced condensation in vivo, while its two intrinsically disordered regions (IDRs) were not. The condensate-forming property of the SMN tudor domain required binding to its ligand, dimethylarginine (DMA), and was shared by at least seven 20 additional tudor domains in six different proteins.
    [Show full text]
  • A High-Throughput Approach to Uncover Novel Roles of APOBEC2, a Functional Orphan of the AID/APOBEC Family
    Rockefeller University Digital Commons @ RU Student Theses and Dissertations 2018 A High-Throughput Approach to Uncover Novel Roles of APOBEC2, a Functional Orphan of the AID/APOBEC Family Linda Molla Follow this and additional works at: https://digitalcommons.rockefeller.edu/ student_theses_and_dissertations Part of the Life Sciences Commons A HIGH-THROUGHPUT APPROACH TO UNCOVER NOVEL ROLES OF APOBEC2, A FUNCTIONAL ORPHAN OF THE AID/APOBEC FAMILY A Thesis Presented to the Faculty of The Rockefeller University in Partial Fulfillment of the Requirements for the degree of Doctor of Philosophy by Linda Molla June 2018 © Copyright by Linda Molla 2018 A HIGH-THROUGHPUT APPROACH TO UNCOVER NOVEL ROLES OF APOBEC2, A FUNCTIONAL ORPHAN OF THE AID/APOBEC FAMILY Linda Molla, Ph.D. The Rockefeller University 2018 APOBEC2 is a member of the AID/APOBEC cytidine deaminase family of proteins. Unlike most of AID/APOBEC, however, APOBEC2’s function remains elusive. Previous research has implicated APOBEC2 in diverse organisms and cellular processes such as muscle biology (in Mus musculus), regeneration (in Danio rerio), and development (in Xenopus laevis). APOBEC2 has also been implicated in cancer. However the enzymatic activity, substrate or physiological target(s) of APOBEC2 are unknown. For this thesis, I have combined Next Generation Sequencing (NGS) techniques with state-of-the-art molecular biology to determine the physiological targets of APOBEC2. Using a cell culture muscle differentiation system, and RNA sequencing (RNA-Seq) by polyA capture, I demonstrated that unlike the AID/APOBEC family member APOBEC1, APOBEC2 is not an RNA editor. Using the same system combined with enhanced Reduced Representation Bisulfite Sequencing (eRRBS) analyses I showed that, unlike the AID/APOBEC family member AID, APOBEC2 does not act as a 5-methyl-C deaminase.
    [Show full text]
  • Novel Targets of Apparently Idiopathic Male Infertility
    International Journal of Molecular Sciences Review Molecular Biology of Spermatogenesis: Novel Targets of Apparently Idiopathic Male Infertility Rossella Cannarella * , Rosita A. Condorelli , Laura M. Mongioì, Sandro La Vignera * and Aldo E. Calogero Department of Clinical and Experimental Medicine, University of Catania, 95123 Catania, Italy; [email protected] (R.A.C.); [email protected] (L.M.M.); [email protected] (A.E.C.) * Correspondence: [email protected] (R.C.); [email protected] (S.L.V.) Received: 8 February 2020; Accepted: 2 March 2020; Published: 3 March 2020 Abstract: Male infertility affects half of infertile couples and, currently, a relevant percentage of cases of male infertility is considered as idiopathic. Although the male contribution to human fertilization has traditionally been restricted to sperm DNA, current evidence suggest that a relevant number of sperm transcripts and proteins are involved in acrosome reactions, sperm-oocyte fusion and, once released into the oocyte, embryo growth and development. The aim of this review is to provide updated and comprehensive insight into the molecular biology of spermatogenesis, including evidence on spermatogenetic failure and underlining the role of the sperm-carried molecular factors involved in oocyte fertilization and embryo growth. This represents the first step in the identification of new possible diagnostic and, possibly, therapeutic markers in the field of apparently idiopathic male infertility. Keywords: spermatogenetic failure; embryo growth; male infertility; spermatogenesis; recurrent pregnancy loss; sperm proteome; DNA fragmentation; sperm transcriptome 1. Introduction Infertility is a widespread condition in industrialized countries, affecting up to 15% of couples of childbearing age [1]. It is defined as the inability to achieve conception after 1–2 years of unprotected sexual intercourse [2].
    [Show full text]
  • Global Analysis of O-Glcnac Glycoproteins in Activated Human T Cells Peder J
    Global Analysis of O-GlcNAc Glycoproteins in Activated Human T Cells Peder J. Lund, Joshua E. Elias and Mark M. Davis This information is current as J Immunol 2016; 197:3086-3098; Prepublished online 21 of October 2, 2021. September 2016; doi: 10.4049/jimmunol.1502031 http://www.jimmunol.org/content/197/8/3086 Downloaded from Supplementary http://www.jimmunol.org/content/suppl/2016/09/20/jimmunol.150203 Material 1.DCSupplemental References This article cites 89 articles, 32 of which you can access for free at: http://www.jimmunol.org/content/197/8/3086.full#ref-list-1 http://www.jimmunol.org/ Why The JI? Submit online. • Rapid Reviews! 30 days* from submission to initial decision • No Triage! Every submission reviewed by practicing scientists • Fast Publication! 4 weeks from acceptance to publication by guest on October 2, 2021 *average Subscription Information about subscribing to The Journal of Immunology is online at: http://jimmunol.org/subscription Permissions Submit copyright permission requests at: http://www.aai.org/About/Publications/JI/copyright.html Author Choice Freely available online through The Journal of Immunology Author Choice option Email Alerts Receive free email-alerts when new articles cite this article. Sign up at: http://jimmunol.org/alerts The Journal of Immunology is published twice each month by The American Association of Immunologists, Inc., 1451 Rockville Pike, Suite 650, Rockville, MD 20852 Copyright © 2016 by The American Association of Immunologists, Inc. All rights reserved. Print ISSN: 0022-1767 Online ISSN: 1550-6606. The Journal of Immunology Global Analysis of O-GlcNAc Glycoproteins in Activated Human T Cells Peder J.
    [Show full text]
  • Protein Symbol Protein Name Rank Metric Score 4F2 4F2 Cell-Surface
    Supplementary Table 2 Supplementary Table 2. Ranked list of proteins present in anti-Sema4D treated macrophage conditioned media obtained in the GSEA analysis of the proteomic data. Proteins are listed according to their rank metric score, which is the score used to position the gene in the ranked list of genes of the GSEA. Values are obtained from comparing Sema4D treated RAW conditioned media versus REST, which includes untreated, IgG treated and anti-Sema4D added RAW conditioned media. GSEA analysis was performed under standard conditions in November 2015. Protein Rank metric symbol Protein name score 4F2 4F2 cell-surface antigen heavy chain 2.5000 PLOD3 Procollagen-lysine,2-oxoglutarate 5-dioxygenase 3 1.4815 ELOB Transcription elongation factor B polypeptide 2 1.4350 ARPC5 Actin-related protein 2/3 complex subunit 5 1.2603 OSTF1 teoclast-stimulating factor 1 1.2500 RL5 60S ribomal protein L5 1.2135 SYK Lysine--tRNA ligase 1.2135 RL10A 60S ribomal protein L10a 1.2135 TXNL1 Thioredoxin-like protein 1 1.1716 LIS1 Platelet-activating factor acetylhydrolase IB subunit alpha 1.1067 A4 Amyloid beta A4 protein 1.0911 H2B1M Histone H2B type 1-M 1.0514 UB2V2 Ubiquitin-conjugating enzyme E2 variant 2 1.0381 PDCD5 Programmed cell death protein 5 1.0373 UCHL3 Ubiquitin carboxyl-terminal hydrolase isozyme L3 1.0061 PLEC Plectin 1.0061 ITPA Inine triphphate pyrophphatase 0.9524 IF5A1 Eukaryotic translation initiation factor 5A-1 0.9314 ARP2 Actin-related protein 2 0.8618 HNRPL Heterogeneous nuclear ribonucleoprotein L 0.8576 DNJA3 DnaJ homolog subfamily
    [Show full text]