DOCSLIB.ORG

Identification of Human APC10/Doc1 As a Subunit of Anaphase Promoting

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Identification of Human APC10/Doc1 As a Subunit of Anaphase Promoting Oncogene (1999) 18, 5131 ± 5137 ã 1999 Stockton Press All rights reserved 0950 ± 9232/99 $15.00 http://www.stockton-press.co.uk/onc Identi®cation of human APC10/Doc1 as a subunit of anaphase promoting complex Yasuhiro Kurasawa1 and Kazuo Todokoro*,1 1Tsukuba Life Science Center, The Institute of Physical and Chemical Research (RIKEN), 3-1, Koyadai, Tsukuba, Ibaraki 305-0074, Japan Anaphase-promoting complex or cyclosome (APC) is a (He et al., 1997; Li et al., 1997; Gorbsky et al., 1998; ubiquitin ligase which speci®cally targets mitotic reg- Kim et al., 1998; Hwang et al., 1998; Fang et al., ulatory factors such as Pds1/Cut2 and cyclin B. 1998b; Kallio et al., 1998), and regulation by Bub2 and Identi®cation of the subunits of multiprotein complex the RENT complex system (Visintin et al., 1998, 1999; APC in several species revealed the highly conserved Fesquet et al., 1999; Fraschini et al., 1999; Alexandru composition of APC from yeast to human. It has been et al., 1999; Shou et al., 1999). reported, however, that vertebrate APC is composed of Identi®cation of the subunits of multiprotein at least eight subunits, APC1 to APC8, while budding complex APC in several species revealed its highly yeast APC is constituted of at least 12 components, conserved composition from yeast to human (Peters et Apc1 to Apc13. It has not yet been clearly understood al., 1996; Zachariae et al., 1998a; Yu et al., 1998). whether additional components found in budding yeast, Vertebrate APC is composed of at least eight subunits, Apc9 to Apc13, are actually composed of mammalian APC1/Tsg24, APC2, APC3/CDC27Hs, APC4, APC5, APC. Here we isolated and characterized human APC6/CDC16Hs, APC7 and APC8/CDC23Hs (Peters APC10/Doc1, and found that APC10/Doc1 binds to et al., 1996; Yu et al., 1998). The APC of budding APC core subunits throughout the cell cycle. Further, it yeast is constituted of at least 12 components, Apc1, was found that APC10/Doc1 is localized in centrosomes Rsi1/Apc2, Cdc27/Apc3, Apc4, Apc5, Cdc16/Apc6, and mitotic spindles throughout mitosis, while it is also Cdc23/Apc8, Apc9, Doc1/Apc10, Apc11, Cdc26/ localized in kinetochores from prophase to anaphase and Apc12 and Apc13 (Zachariae et al., 1996, 1998a). It in midbody in telophase and cytokinesis. These results has not yet been clearly established whether or not strongly support the notion that human APC10/Doc1 additional components found in budding yeast, Apc9 may be one of the APC core subunits rather than the to Apc13, are actually composed of mammalian APC. transiently associated regulatory factor. APC10/DOC1, which was originally isolated by the enrichment of a mitotic arrest mutant of budding yeast, Keywords: APC; APC10; centromere; Doc1; ubiquitin was identi®ed as a gene encoding a component of cyclin proteolysis machinery (Irniger et al., 1995; Hwang and Murray, 1997). The doc1 mutants were arrested as large budded cells at 378C with a single Introduction nucleus and a short spindle, which were typical phenotypes in mitotic cyclin proteolysis mutants and Anaphase-promoting complex or cyclosome (APC) is a showed a defect in the ubiquitination of mitotic cyclin ubiquitin ligase which speci®cally targets the mitotic in G1-arrested cells (Hwang and Murray, 1997). regulatory factors (King et al., 1995; Sudakin et al., Indeed, identi®cation of budding yeast APC subunits 1995; Funabiki et al., 1996; Peters et al., 1996; revealed that Doc1 is one of the APC components Zachariae et al., 1998a; Yu et al., 1998). The APC (Zachariae et al., 1998a). Fission yeast apc10 mutant activity was found to be regulated by at least four was isolated as a G1 arrest mutant which showed distinct mechanisms; activation and inactivation by sterility and temperature-sensitive growth with a defect phosphorylation and dephosphorylation (Hershko et in chromosome segregation (Kominami et al., 1998). al., 1994; Lahav-Baratz et al., 1995; Sudakin et al., The apc10+ was essential for cell viability, and the 1995; Yamashita et al., 1996; Ishii et al., 1996; Peters et mutant showed two kinds of terminal morphology, al., 1996; Shirayama et al., 1998; Descombes and Nigg, elongated cells and cut phenotypes (Kominami et al., 1998; Charles et al., 1998; Kotani et al., 1998, 1999), 1998). It was found that Apc10 genetically and activation by the binding of substrate-speci®c WD- physically interacts with a universal subunit of the repeat proteins, Cdc20/p55CDC/Fizzy and Cdh1/Hct1/ APC, and is required for the ubiquitination of B type Fizzy-related (Visintin et al., 1997; Schwab et al., 1997; cyclin (Kominami et al., 1998). These results suggest Kallio et al., 1998; Fang et al., 1998a,b; Kramer et al., that Apc10/Doc1 is crucial for mitotic exit and for the 1998; Zachariae et al., 1998b; Lorca et al., 1998), degradation of mitotic cyclins; however, it was suppression of APC activity by the spindle assembly demonstrated that Apc10/Doc1 may not be a core checkpoint through the Mad family and Bub family component of APC. The majority but not all Doc1 population co-sedimented at 20S APC in a sucrose gradient in budding yeast (Hwang and Murray, 1997). In ®ssion yeast, myc-tagged Apc10 was not co- sedimented with APC, and the sedimentation pro®les *Correspondence: K Todokoro Received 26 June 1999; revised 9 August 1999; accepted 9 August of APC components appear normal in apc10 mutant 1999 (Kominami et al., 1998), whereas APC was physically Human APC10/Doc1 as a subunit of APC Y Kurasawa and K Todokoro 5132 disrupted in other mutations of APC components, such Expression of human APC10/Doc1 during cell cycle as cut4, cut9 and nuc2 (Yamashita et al., 1996; Yamada et al., 1997). In addition, APC10/Doc1 has not been A polyclonal rabbit antiserum was raised against identi®ed in vertebrate APC. human APC10/Doc1 fused to glutathione S-transfer- In spite of the rapid progress on the studies of ase, and puri®ed by anity chromatography of the regulation of APC activity, whole APC core compo- fusion protein. The puri®ed antibody detected APC10/ nents and their precise subcellular localization during Doc1 of 24 kDa in HeLa cell lysates in G1/S phase mitosis remains obscure. We therefore isolated and and M phase (Figure 2a). characterized human APC10/Doc1, and found that The expression level of APC10/Doc1 during the cell human APC10/Doc1 is one of the APC core cycle was examined by immunoblot analysis. HeLa components. cells were arrested at G1/S phase by aphidicolin, released, and then re-arrested by nocodazole at prometaphase. The cells were harvested at various time points after nocodazole release, and the cell Results extracts were probed with anti-APC10/Doc1 anti- body, anti-cyclin B1 antibody and anti-APC3/Cdc27 Isolation of human APC10/Doc1 cDNA antibody. The cyclin B1 was mostly degraded within A homology search of the human expressed sequence 60 min after nocodazole release (Figure 2b, upper tag (EST) data base identi®ed a cDNA fragment that is panel) as cells entered G1 phase. The supershift of signi®cantly homologous to APC10/Doc1 of budding APC3/Cdc27 by phosphorylation similarly diminished yeast and ®ssion yeast (Irniger et al., 1995; Hwang and within 60 min (Figure 2b, middle panel), while the level Murray, 1997; Kominami et al., 1998). Full-length of protein (sum of phosphorylated and unphosphory- human APC10/Doc1 cDNA (DDBJ/EMBL/GenBank lated forms) was unchanged. Figure 2b (lower panel) accession number AB012109) was isolated from a shows that the level of APC10/Doc1 was constant from cDNA library of human erythroleukemia K562 cells, prometaphase to G1/S phase, indicating that APC using an EST fragment ampli®ed by PCR as a probe. activity is not correlated with the protein level of Sequence analysis of seven independent clones revealed APC10/Doc1 during the cell cycle. that all cDNAs isolated encode the same open reading frame but have various amino-terminal sequences. APC10/Doc1 as a subunit of APC Thus the translational initiation site was con®rmed by cap site PCR. Figure 1 shows the con®rmed full- To examine whether or not mammalian APC10/Doc1 length human APC10/Doc1 sequence aligned with is one of the APC core subunits, the binding of those of budding yeast and ®ssion yeast. Human APC10/Doc1 with other APC components during the APC10/Doc1 cDNA encodes a peptide of 185 amino cell cycle was examined in HeLa cells by immunopre- acids with a predicted molecular mass of 21 252 cipitation followed by immunoblot analysis. The daltons and a predicted isoelectric point of 9.71. APC10/Doc1 was clearly detected in the immunopre- Human APC10/Doc1 shares 28.5% and 37.8% amino cipitates with anti-APC3/Cdc27 antibody in aphidico- acid sequence identity and 40.0% and 48.2% sequence lin-treated cells (G1/S phase) (Figure 3a, lane 2), similarity with those of budding yeast and ®ssion yeast, nocodazole-treated cells (prometaphase) (Figure 3a, respectively. lane 3), and cells within 45 min after nocodazole Figure 1 Sequence alignments of human APC10/Doc1 with corresponding yeast proteins. Identical amino acid residues are shown on a black background. Conservative amino acid substitutions are shown on a gray background Human APC10/Doc1 as a subunit of APC Y Kurasawa and K Todokoro 5133 Figure 2 Expression level of human APC10/Doc1 during cell cycle. (a) Cell lysates of aphidicolin-treated (G1/S phase) (lane 1) or nocodazole-treated (prometaphase) HeLa cells were immunoblotted with anti-human APC10/Doc1 antibody. Arrow indicates APC10/Doc1. (b) HeLa cells were arrested at G1/S phase by aphidicolin, released, and then re-arrested by nocodazole at prometaphase. The cells were harvested at the indicated time points after nocodazole release, and equal amounts of the cell extracts were probed with anti-cyclin B1 antibody (upper panel), anti-APC3/Cdc27 antibody (middle panel) and anti-APC10/Doc1 antibody (lower panel) release (late mitosis) (Figure 3a, lane 4), but not in the dots on the chromosomes were kinetochores of the immunoprecipitate with non-immune serum (Figure 3a, sister chromatids (Figure 5, lane 1).
Load more

Recommended publications
  • Genomic Correlates of Relationship QTL Involved in Fore- Versus Hind Limb Divergence in Mice
    Loyola University Chicago Loyola eCommons Biology: Faculty Publications and Other Works Faculty Publications 2013 Genomic Correlates of Relationship QTL Involved in Fore- Versus Hind Limb Divergence in Mice Mihaela Palicev Gunter P. Wagner James P. Noonan Benedikt Hallgrimsson James M. Cheverud Loyola University Chicago, [email protected] Follow this and additional works at: https://ecommons.luc.edu/biology_facpubs Part of the Biology Commons Recommended Citation Palicev, M, GP Wagner, JP Noonan, B Hallgrimsson, and JM Cheverud. "Genomic Correlates of Relationship QTL Involved in Fore- Versus Hind Limb Divergence in Mice." Genome Biology and Evolution 5(10), 2013. This Article is brought to you for free and open access by the Faculty Publications at Loyola eCommons. It has been accepted for inclusion in Biology: Faculty Publications and Other Works by an authorized administrator of Loyola eCommons. For more information, please contact [email protected]. This work is licensed under a Creative Commons Attribution-Noncommercial-No Derivative Works 3.0 License. © Palicev et al., 2013. GBE Genomic Correlates of Relationship QTL Involved in Fore- versus Hind Limb Divergence in Mice Mihaela Pavlicev1,2,*, Gu¨ nter P. Wagner3, James P. Noonan4, Benedikt Hallgrı´msson5,and James M. Cheverud6 1Konrad Lorenz Institute for Evolution and Cognition Research, Altenberg, Austria 2Department of Pediatrics, Cincinnati Children‘s Hospital Medical Center, Cincinnati, Ohio 3Yale Systems Biology Institute and Department of Ecology and Evolutionary Biology, Yale University 4Department of Genetics, Yale University School of Medicine 5Department of Cell Biology and Anatomy, The McCaig Institute for Bone and Joint Health and the Alberta Children’s Hospital Research Institute for Child and Maternal Health, University of Calgary, Calgary, Canada 6Department of Anatomy and Neurobiology, Washington University *Corresponding author: E-mail: [email protected].
    [Show full text]
  • Protein Interaction Network of Alternatively Spliced Isoforms from Brain Links Genetic Risk Factors for Autism
    ARTICLE Received 24 Aug 2013 | Accepted 14 Mar 2014 | Published 11 Apr 2014 DOI: 10.1038/ncomms4650 OPEN Protein interaction network of alternatively spliced isoforms from brain links genetic risk factors for autism Roser Corominas1,*, Xinping Yang2,3,*, Guan Ning Lin1,*, Shuli Kang1,*, Yun Shen2,3, Lila Ghamsari2,3,w, Martin Broly2,3, Maria Rodriguez2,3, Stanley Tam2,3, Shelly A. Trigg2,3,w, Changyu Fan2,3, Song Yi2,3, Murat Tasan4, Irma Lemmens5, Xingyan Kuang6, Nan Zhao6, Dheeraj Malhotra7, Jacob J. Michaelson7,w, Vladimir Vacic8, Michael A. Calderwood2,3, Frederick P. Roth2,3,4, Jan Tavernier5, Steve Horvath9, Kourosh Salehi-Ashtiani2,3,w, Dmitry Korkin6, Jonathan Sebat7, David E. Hill2,3, Tong Hao2,3, Marc Vidal2,3 & Lilia M. Iakoucheva1 Increased risk for autism spectrum disorders (ASD) is attributed to hundreds of genetic loci. The convergence of ASD variants have been investigated using various approaches, including protein interactions extracted from the published literature. However, these datasets are frequently incomplete, carry biases and are limited to interactions of a single splicing isoform, which may not be expressed in the disease-relevant tissue. Here we introduce a new interactome mapping approach by experimentally identifying interactions between brain-expressed alternatively spliced variants of ASD risk factors. The Autism Spliceform Interaction Network reveals that almost half of the detected interactions and about 30% of the newly identified interacting partners represent contribution from splicing variants, emphasizing the importance of isoform networks. Isoform interactions greatly contribute to establishing direct physical connections between proteins from the de novo autism CNVs. Our findings demonstrate the critical role of spliceform networks for translating genetic knowledge into a better understanding of human diseases.
    [Show full text]
  • Supplementary Information Integrative Analyses of Splicing in the Aging Brain: Role in Susceptibility to Alzheimer’S Disease
    Supplementary Information Integrative analyses of splicing in the aging brain: role in susceptibility to Alzheimer’s Disease Contents 1. Supplementary Notes 1.1. Religious Orders Study and Memory and Aging Project 1.2. Mount Sinai Brain Bank Alzheimer’s Disease 1.3. CommonMind Consortium 1.4. Data Availability 2. Supplementary Tables 3. Supplementary Figures Note: Supplementary Tables are provided as separate Excel files. 1. Supplementary Notes 1.1. Religious Orders Study and Memory and Aging Project Gene expression data1. Gene expression data were generated using RNA- sequencing from Dorsolateral Prefrontal Cortex (DLPFC) of 540 individuals, at an average sequence depth of 90M reads. Detailed description of data generation and processing was previously described2 (Mostafavi, Gaiteri et al., under review). Samples were submitted to the Broad Institute’s Genomics Platform for transcriptome analysis following the dUTP protocol with Poly(A) selection developed by Levin and colleagues3. All samples were chosen to pass two initial quality filters: RNA integrity (RIN) score >5 and quantity threshold of 5 ug (and were selected from a larger set of 724 samples). Sequencing was performed on the Illumina HiSeq with 101bp paired-end reads and achieved coverage of 150M reads of the first 12 samples. These 12 samples will serve as a deep coverage reference and included 2 males and 2 females of nonimpaired, mild cognitive impaired, and Alzheimer's cases. The remaining samples were sequenced with target coverage of 50M reads; the mean coverage for the samples passing QC is 95 million reads (median 90 million reads). The libraries were constructed and pooled according to the RIN scores such that similar RIN scores would be pooled together.
    [Show full text]
  • Title: Therapeutic Potential of HSP90 Inhibition for Neurofibromatosis Type 2
    Author Manuscript Published OnlineFirst on May 28, 2013; DOI: 10.1158/1078-0432.CCR-12-3167 Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited. Title: Therapeutic Potential of HSP90 Inhibition for Neurofibromatosis type 2 Karo Tanaka1, Ascia Eskin3, Fabrice Chareyre1, Walter J. Jessen4, Jan Manent5, Michiko Niwa-Kawakita6, Ruihong Chen7, Cory H. White2, Jeremie Vitte1, Zahara M. Jaffer1, Stanley F. Nelson3, Allan E. Rubenstein8, Marco Giovannini1,9§. Authors’ affiliations: House Research Institute, 1Center for Neural Tumor Research and 2Section on Genetics of Hereditary Ear Disorders, Los Angeles, CA; 3Department of Human Genetics, University of California, Los Angeles, CA; 4Informatics, Covance Inc., Princeton, NJ; 5Peter MacCallum Cancer Institute, Melbourne, Australia; 6Inserm U944, CNRS U7212, Université Paris, Institut Universitaire d'Hématologie, Paris, France; 7NexGenix Pharmaceuticals, Burlingame, CA; and 8New York University Langone Medical Center, New York, NY; and Department of Cell and Neurobiology, University of Southern California, Keck School of Medicine, Los Angeles, CA Running title: HSP90 Inhibition for NF2 Keywords: NF2, HSP90 inhibitors, Transcriptome Financial support: This work was supported by a Drug Discovery Initiative Award, Children’s Tumor Foundation, to M.G., and by the House Research Institute. Corresponding author: Marco Giovannini, House Research Institute, Center for Neural Tumor Research, 2100 West 3rd street, Los Angeles, CA90057. Phone: +1-213-989-6708; Fax: +1-213-989-6778; E-mail: [email protected] 1 Downloaded from clincancerres.aacrjournals.org on September 30, 2021. © 2013 American Association for Cancer Research. Author Manuscript Published OnlineFirst on May 28, 2013; DOI: 10.1158/1078-0432.CCR-12-3167 Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited.
    [Show full text]
  • CDC23 (NM 004661) Human Tagged ORF Clone Product Data
    OriGene Technologies, Inc. 9620 Medical Center Drive, Ste 200 Rockville, MD 20850, US Phone: +1-888-267-4436 [email protected] EU: [email protected] CN: [email protected] Product datasheet for RC224973L3 CDC23 (NM_004661) Human Tagged ORF Clone Product data: Product Type: Expression Plasmids Product Name: CDC23 (NM_004661) Human Tagged ORF Clone Tag: Myc-DDK Symbol: CDC23 Synonyms: ANAPC8; APC8; CUT23 Vector: pLenti-C-Myc-DDK-P2A-Puro (PS100092) E. coli Selection: Chloramphenicol (34 ug/mL) Cell Selection: Puromycin ORF Nucleotide The ORF insert of this clone is exactly the same as(RC224973). Sequence: Restriction Sites: SgfI-MluI Cloning Scheme: ACCN: NM_004661 ORF Size: 1791 bp This product is to be used for laboratory only. Not for diagnostic or therapeutic use. View online » ©2021 OriGene Technologies, Inc., 9620 Medical Center Drive, Ste 200, Rockville, MD 20850, US 1 / 2 CDC23 (NM_004661) Human Tagged ORF Clone – RC224973L3 OTI Disclaimer: The molecular sequence of this clone aligns with the gene accession number as a point of reference only. However, individual transcript sequences of the same gene can differ through naturally occurring variations (e.g. polymorphisms), each with its own valid existence. This clone is substantially in agreement with the reference, but a complete review of all prevailing variants is recommended prior to use. More info OTI Annotation: This clone was engineered to express the complete ORF with an expression tag. Expression varies depending on the nature of the gene. RefSeq: NM_004661.3 RefSeq Size: 3169 bp RefSeq ORF: 1794 bp Locus ID: 8697 UniProt ID: Q9UJX2 Domains: TPR, APC8 Protein Families: Druggable Genome Protein Pathways: Cell cycle, Oocyte meiosis, Progesterone-mediated oocyte maturation, Ubiquitin mediated proteolysis MW: 68.7 kDa Gene Summary: The protein encoded by this gene shares strong similarity with Saccharomyces cerevisiae Cdc23, a protein essential for cell cycle progression through the G2/M transition.
    [Show full text]
  • The Genome of Schmidtea Mediterranea and the Evolution Of
    OPEN ArtICLE doi:10.1038/nature25473 The genome of Schmidtea mediterranea and the evolution of core cellular mechanisms Markus Alexander Grohme1*, Siegfried Schloissnig2*, Andrei Rozanski1, Martin Pippel2, George Robert Young3, Sylke Winkler1, Holger Brandl1, Ian Henry1, Andreas Dahl4, Sean Powell2, Michael Hiller1,5, Eugene Myers1 & Jochen Christian Rink1 The planarian Schmidtea mediterranea is an important model for stem cell research and regeneration, but adequate genome resources for this species have been lacking. Here we report a highly contiguous genome assembly of S. mediterranea, using long-read sequencing and a de novo assembler (MARVEL) enhanced for low-complexity reads. The S. mediterranea genome is highly polymorphic and repetitive, and harbours a novel class of giant retroelements. Furthermore, the genome assembly lacks a number of highly conserved genes, including critical components of the mitotic spindle assembly checkpoint, but planarians maintain checkpoint function. Our genome assembly provides a key model system resource that will be useful for studying regeneration and the evolutionary plasticity of core cell biological mechanisms. Rapid regeneration from tiny pieces of tissue makes planarians a prime De novo long read assembly of the planarian genome model system for regeneration. Abundant adult pluripotent stem cells, In preparation for genome sequencing, we inbred the sexual strain termed neoblasts, power regeneration and the continuous turnover of S. mediterranea (Fig. 1a) for more than 17 successive sib- mating of all cell types1–3, and transplantation of a single neoblast can rescue generations in the hope of decreasing heterozygosity. We also developed a lethally irradiated animal4. Planarians therefore also constitute a a new DNA isolation protocol that meets the purity and high molecular prime model system for stem cell pluripotency and its evolutionary weight requirements of PacBio long-read sequencing12 (Extended Data underpinnings5.
    [Show full text]
  • Origin of a Folded Repeat Protein from an Intrinsically Disordered Ancestor
    RESEARCH ARTICLE Origin of a folded repeat protein from an intrinsically disordered ancestor Hongbo Zhu, Edgardo Sepulveda, Marcus D Hartmann, Manjunatha Kogenaru†, Astrid Ursinus, Eva Sulz, Reinhard Albrecht, Murray Coles, Jo¨ rg Martin, Andrei N Lupas* Department of Protein Evolution, Max Planck Institute for Developmental Biology, Tu¨ bingen, Germany Abstract Repetitive proteins are thought to have arisen through the amplification of subdomain- sized peptides. Many of these originated in a non-repetitive context as cofactors of RNA-based replication and catalysis, and required the RNA to assume their active conformation. In search of the origins of one of the most widespread repeat protein families, the tetratricopeptide repeat (TPR), we identified several potential homologs of its repeated helical hairpin in non-repetitive proteins, including the putatively ancient ribosomal protein S20 (RPS20), which only becomes structured in the context of the ribosome. We evaluated the ability of the RPS20 hairpin to form a TPR fold by amplification and obtained structures identical to natural TPRs for variants with 2–5 point mutations per repeat. The mutations were neutral in the parent organism, suggesting that they could have been sampled in the course of evolution. TPRs could thus have plausibly arisen by amplification from an ancestral helical hairpin. DOI: 10.7554/eLife.16761.001 *For correspondence: andrei. [email protected] Introduction † Present address: Department Most present-day proteins arose through the combinatorial shuffling and differentiation of a set of of Life Sciences, Imperial College domain prototypes. In many cases, these prototypes can be traced back to the root of cellular life London, London, United and have since acted as the primary unit of protein evolution (Anantharaman et al., 2001; Kingdom Apic et al., 2001; Koonin, 2003; Kyrpides et al., 1999; Orengo and Thornton, 2005; Ponting and Competing interests: The Russell, 2002; Ranea et al., 2006).
    [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]
  • Genome-Wide Identification and Analysis of Mirnas Complementary to Upstream Sequences of Mrna Transcription Start Sites
    In: Gene Silencing: Theory, Techniques and Applications ISBN: 978-1-61728-276-8 Editor: Anthony J. Catalano © 2010 Nova Science Publishers, Inc. The exclusive license for this PDF is limited to personal printing only. No part of this digital document may be reproduced, stored in a retrieval system or transmitted commercially in any form or by any means. The publisher has taken reasonable care in the preparation of this digital document, but makes no expressed or implied warranty of any kind and assumes no responsibility for any errors or omissions. No liability is assumed for incidental or consequential damages in connection with or arising out of information contained herein. This digital document is sold with the clear understanding that the publisher is not engaged in rendering legal, medical or any other professional services. Chapter XIII Genome-wide Identification and Analysis of miRNAs Complementary to Upstream Sequences of mRNA Transcription Start Sites Kenta Narikawa1, Kenji Nishi2, Yuki Naito2, Minami Mazda2 and Kumiko Ui-Tei1,2,* 1Department of Computational Biology, Graduate School of Frontier Sciences, University of Tokyo, 5-1-5 Kashiwanoha, Kashiwa-shi, Chiba-ken 277-8561, Japan 2Department of Biophysics and Biochemistry, Graduate School of Science, University of Tokyo, 2-11-16 Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan Abstract Small regulatory RNAs including short interfering RNAs (siRNAs) and microRNAs (miRNAs) are crucial regulators of gene expression at the posttranscriptional level. Recently, additional roles for small RNAs in gene activation and suppression at the transcriptional level were reported; these RNAs were shown to have sequences that closely or completely match to their respective promoter regions.
    [Show full text]
  • An Architectural Map of the Anaphase-Promoting Complex
    Downloaded from genesdev.cshlp.org on September 28, 2021 - Published by Cold Spring Harbor Laboratory Press An architectural map of the anaphase-promoting complex Brian R. Thornton,1,3 Tessie M. Ng,1,3 Mary E. Matyskiela,2 Christopher W. Carroll,2 David O. Morgan,2 and David P. Toczyski1,4 1Cancer Research Institute, Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, California 94115, USA; 2Department of Physiology, University of California, San Francisco, California 94143-2200, USA The anaphase-promoting complex or cyclosome (APC) is an unusually complicated ubiquitin ligase, composed of 13 core subunits and either of two loosely associated regulatory subunits, Cdc20 and Cdh1. We analyzed the architecture of the APC using a recently constructed budding yeast strain that is viable in the absence of normally essential APC subunits. We found that the largest subunit, Apc1, serves as a scaffold that associates independently with two separable subcomplexes, one that contains Apc2 (Cullin), Apc11 (RING), and Doc1/Apc10, and another that contains the three TPR subunits (Cdc27, Cdc16, and Cdc23). We found that the three TPR subunits display a sequential binding dependency, with Cdc27 the most peripheral, Cdc23 the most internal, and Cdc16 between. Apc4, Apc5, Cdc23, and Apc1 associate interdependently, such that loss of any one subunit greatly reduces binding between the remaining three. Intriguingly, the cullin and TPR subunits both contribute to the binding of Cdh1 to the APC. Enzymatic assays performed with APC purified from strains lacking each of the essential subunits revealed that only cdc27⌬ complexes retain detectable activity in the presence of Cdh1.
    [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]
  • Disruption of the Anaphase-Promoting Complex Confers Resistance to TTK Inhibitors in Triple-Negative Breast Cancer
    Disruption of the anaphase-promoting complex confers resistance to TTK inhibitors in triple-negative breast cancer K. L. Thua,b, J. Silvestera,b, M. J. Elliotta,b, W. Ba-alawib,c, M. H. Duncana,b, A. C. Eliaa,b, A. S. Merb, P. Smirnovb,c, Z. Safikhanib, B. Haibe-Kainsb,c,d,e, T. W. Maka,b,c,1, and D. W. Cescona,b,f,1 aCampbell Family Institute for Breast Cancer Research, Princess Margaret Cancer Centre, University Health Network, Toronto, ON, Canada M5G 1L7; bPrincess Margaret Cancer Centre, University Health Network, Toronto, ON, Canada M5G 1L7; cDepartment of Medical Biophysics, University of Toronto, Toronto, ON, Canada M5G 1L7; dDepartment of Computer Science, University of Toronto, Toronto, ON, Canada M5G 1L7; eOntario Institute for Cancer Research, Toronto, ON, Canada M5G 0A3; and fDepartment of Medicine, University of Toronto, Toronto, ON, Canada M5G 1L7 Contributed by T. W. Mak, December 27, 2017 (sent for review November 9, 2017; reviewed by Mark E. Burkard and Sabine Elowe) TTK protein kinase (TTK), also known as Monopolar spindle 1 (MPS1), ator of the spindle assembly checkpoint (SAC), which delays is a key regulator of the spindle assembly checkpoint (SAC), which anaphase until all chromosomes are properly attached to the functions to maintain genomic integrity. TTK has emerged as a mitotic spindle, TTK has an integral role in maintaining genomic promising therapeutic target in human cancers, including triple- integrity (6). Because most cancer cells are aneuploid, they are negative breast cancer (TNBC). Several TTK inhibitors (TTKis) are heavily reliant on the SAC to adequately segregate their abnormal being evaluated in clinical trials, and an understanding of karyotypes during mitosis.
    [Show full text]