DOCSLIB.ORG

The RNA-Binding Zinc-Finger Protein Tristetraprolin Regulates AU-Rich Mrnas Involved in Breast Cancer-Related Processes

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
The RNA-Binding Zinc-Finger Protein Tristetraprolin Regulates AU-Rich Mrnas Involved in Breast Cancer-Related Processes Oncogene (2010) 29, 4205–4215 & 2010 Macmillan Publishers Limited All rights reserved 0950-9232/10 www.nature.com/onc ORIGINAL ARTICLE The RNA-binding zinc-finger protein tristetraprolin regulates AU-rich mRNAs involved in breast cancer-related processes N Al-Souhibani1, W Al-Ahmadi1, JE Hesketh2, PJ Blackshear3 and KSA Khabar1 1Program in BioMolecular Research, King Faisal Specialist Hospital and Research Center, Riyadh, Saudi Arabia; 2Institute for Cell and Molecular Biosciences, Newcastle University, Newcastle upon Tyne, UK and 3Laboratory of Signal Transduction, NIEHS, National Institutes of Health, Research Triangle, NC, USA Tristetraprolin (TTP or ZFP36) is a tandem CCCH zinc- Introduction finger RNA-binding protein that regulates the stability of certain AU-rich element (ARE) mRNAs. Recent work Breast cancer is the most common type of malignant suggests that TTP is deficient in cancer cells when cancer among women, with a high incidence and compared with normal cell types. In this study we found mortality rate, and comprises almost a fifth of all female that TTP expression was lower in invasive breast cancer cancers (McPherson et al., 2000). Cancer metastasis is cells (MDAMB231) compared with normal breast cell dependent on the tumor’s ability to degrade components lines MCF12A and MCF-10. TTP targets were probed of the extracellular matrix by different proteolytic using a novel approach by expressing the C124R zinc- enzymes (Liotta et al., 1980; Liotta, 1986; Bacac and finger TTP mutant that functions as dominant negative Stamenkovic, 2008). Alterations in the expression of and increases target mRNA expression. In contrast to many genes have been implicated in the invasiveness and wild-type TTP, C124R TTP was able to increase certain metastatic potential of malignant breast cancers (Liotta ARE-mRNA expressions in serum-stimulated breast et al., 1980; Liotta, 1986). Several gene products of the cancer cells. Using an ARE-gene microarray, novel AU-rich element mRNA (ARE-mRNA) family are targets of TTP regulation were identified, namely, overexpressed in cancer cells and such increased expres- urokinase plasminogen activator (uPA), uPA receptor sion correlates with increased invasion and metastasis. and matrix metalloproteinase-1, all known to have These include cyclooxygenase 2 (Soslow et al., 2000), prominent roles in breast cancer invasion and metastasis. urokinase plasminogen activator (uPA) (Nanbu et al., Expression of these targets was upregulated in tumori- 1994), uPA receptor (uPAR) (Roldan et al., 1990) and genic types, particularly in highly invasive MDAMB231. matrix metalloproteinase 1 (MMP1) (Fini et al., 1987). The mRNA half-lives of these TTP-regulated genes were The expression of ARE-mRNA gene products is increased in TTP-knockout embryonic mouse fibroblasts, regulated by RNA-binding proteins, which are trans- as assessed using real-time polymerase chain reaction, acting factors that bind to AREs and regulate the whereas forced restoration of TTP by transfection led to a stability of ARE-mRNAs. Among those is tristetrapro- reduction in their mRNA levels. RNA immunoprecipita- lin (TTP), an RNA-binding protein that is rapidly and tion confirmed an association of TTP, but not C124R, transiently expressed in response to extracellular stimuli with these target transcripts. Moreover, TTP reduced, such as serum, growth factors, phorbol esters and whereas the mutant C124R TTP increased, the activity of insulin (DuBois et al., 1990; Lai et al., 1990) and binds reporter constructs fused to target ARE. As a result of to and destabilizes mRNAs that contain AREs in their TTP regulation, invasiveness of MDAMB231 cells was 30 untranslated regions (UTRs) (Carballo et al., 1998). reduced. The data suggest that TTP, in a 30 untranslated TTP is a tandem zinc-finger protein and consists of two region—and ARE-dependent manner, regulates an zinc-fingers that are necessary for TTP binding to important subset of cancer-related genes that are involved tumor necrosis factor (TNF)-a 30UTR, a major target in cellular growth, invasion and metastasis. for TTP (Lai et al., 2000). Mutation of either zinc-finger Oncogene (2010) 29, 4205–4215; doi:10.1038/onc.2010.168; by a single-point mutation (Cys to Arg), for example, published online 24 May 2010 C124R mutant, fails to bind TNF-a 30UTR and ARE (Lai et al., 1999). Well-established targets of TTP Keywords: posttranscriptional control; AU-rich elements; involved in cancer have been identified, namely, RNA-binding proteins; RNA stability; tristetraprolin cyclooxygenase 2 (Sawaoka et al., 2003; Young et al., 2009) and vascular endothelial growth factor (Essafi- Benkhadir et al., 2007). Recently, we have shown that a Correspondence: Dr KSA Khabar, Program in BioMolecular polyadenylation transcript variant of HuR is an ARE- Research, King Faisal Specialist Hospital and Research Center, mRNA to which wild-type TTP, but not the zinc-finger P3354, Riyadh 11211, Saudi Arabia. mutant (C124R), competes with HuR for binding to E-mail: [email protected] Received 19 November 2009; revised 3 April 2010; accepted 9 April 2010; HuR mRNA itself (Al-Ahmadi et al., 2009a). Further, published online 24 May 2010 TTP mRNA targets have been recently identified, TTP regulation of AU-rich mRNAs in breast cancer lines N Al-Souhibani et al 4206 including immediate early response 3 (Lai et al., 2006) and interleukin (IL)-10 (Stoecklin et al., 2008). MCF10A ER- TTP mRNA and protein levels were recently found to be significantly decreased in tumors of the thyroid, lung, ER+ ** ANOVA ovary, uterus and breast compared with nontransformed MCF12A tissue samples (Brennan et al., 2009). Furthermore, loss + of TTP expression was observed in several adenomas Cell line MCF7 ER and adenocarcinomas and correlates with increased expression of HuR and cyclooxygenase 2 (Young et al., Tumor Normal 2009). As a result, TTP dysfunction may lead to MDAMB231 ** ER- abnormalities that contribute to cancer processes. Our aim was to investigate the role of TTP in global 0 regulation of ARE-gene expression and identify novel 20,000 40,000 60,000 80,000 100,000120,000140,000 TTP-regulated genes in the context of cancer, particu- Relative TTP expression (arbitrary units) larly breast cancer. Observations that the TTP zinc- finger mutant C124R can lead to an increase in TNF-a 1,500,000 mRNA (Lai et al., 2002) and HuR mRNA expression Control (Al-Ahmadi et al., 2009a) in an ARE-dependent manner TTP 1,250,000 C124R promoted us to further explore this dominant-negative feature. Thus, we used C124R as a biological probe for 1,000,000 finding a functional ARE-mRNA repertoire using microarray experiments. We showed that TTP binds to and destabilizes important cancer-related transcripts, 750,000 namely, MMP1, uPA and uPAR transcripts, in a 30UTR and ARE-dependent manner. We confirmed these 500,000 targets using different experimental approaches. We also showed that TTP can suppress the invasive ARE-GFP Fluorescence 250,000 potential of breast cancer cells. 0 0 25 50 75 100 125 150 175 200 plasmid concentration (ng) Results HEK293 MDAMB231 TTP mRNA expression in normal and tumor breast cell TTP lines To compare the expression of endogenous TTP mRNA in normal versus tumor breast cell lines, four different β-actin mammary cell lines were used. Estrogen receptor (ER) status was considered when selecting cell lines: one normal CC TTP CC TTP C12R C12R and one tumor cell line were chosen that were ER positive Vector Vector (MCF12A and MCF-7, respectively) and similarly one Figure 1 (a) TTP expression profile in normal and tumor breast normal and one tumor cell line that did not express ER at cell lines. Cells were serum starved (0.5% fetal bovine serum) detectable levels, namely, MCF10A and MDAMB231. overnight, then restimulated with 10% serum for 1 h. Total RNA was then extracted and reverse transcribed for quantitative PCR. The latter is well known for its increased invasiveness. Taqman expression assays specific for human TTP were performed To induce the expression of TTP mRNA, serum- and quantitation was normalized to human RPLP0 as the starved cell lines were stimulated with serum for 1 and endogenous control in the real-time PCR experiments. Values 2 h. TTP mRNA expression in all four cell lines showed shown are means±s.e.m. from three independent experiments, a pattern that is typical of TTP, in which mRNA **Po0.01 as determined by ANOVA. ER: estrogen receptor. (b) Response of ARE-GFP reporter in HEK293 cells transfected accumulation peaked at 1 h and rapidly decreased to with 25 ng ARE-GFP reporter plasmid and cotransfected with near-basal levels at 2 h (data not shown). A comparison various amounts of TTP, C124R (mutant TTP) or a vector plasmid of TTP mRNA expression levels at 1 h was carried out alone. GFP fluorescence was measured 24 h after transfection. (c) for the four cell lines (Figure 1a). Normal mammary cell Level of TTP protein in untransfected and transfected cells after serum induction. A total of 40 mg of total protein was loaded onto lines, particularly MCF10A, exhibited a high expression sodium dodecyl sulfate-PAGE gel and probed with an antibody of TTP mRNA, when compared with the highly invasive against TTP or b-actin as a loading control and transfection MDAMB231 cell line (Figure 1a). The ER-negative, efficiency was determined by measurement of cotransfected GFP. nontumorigenic MCF10A cell line showed the highest expression; it was 10-fold higher than MDAMB231 cells that exhibited the least TTP expression of all four cell varying mobilities were
Load more

Recommended publications
  • Analysis of Gene Expression Data for Gene Ontology
    ANALYSIS OF GENE EXPRESSION DATA FOR GENE ONTOLOGY BASED PROTEIN FUNCTION PREDICTION A Thesis Presented to The Graduate Faculty of The University of Akron In Partial Fulfillment of the Requirements for the Degree Master of Science Robert Daniel Macholan May 2011 ANALYSIS OF GENE EXPRESSION DATA FOR GENE ONTOLOGY BASED PROTEIN FUNCTION PREDICTION Robert Daniel Macholan Thesis Approved: Accepted: _______________________________ _______________________________ Advisor Department Chair Dr. Zhong-Hui Duan Dr. Chien-Chung Chan _______________________________ _______________________________ Committee Member Dean of the College Dr. Chien-Chung Chan Dr. Chand K. Midha _______________________________ _______________________________ Committee Member Dean of the Graduate School Dr. Yingcai Xiao Dr. George R. Newkome _______________________________ Date ii ABSTRACT A tremendous increase in genomic data has encouraged biologists to turn to bioinformatics in order to assist in its interpretation and processing. One of the present challenges that need to be overcome in order to understand this data more completely is the development of a reliable method to accurately predict the function of a protein from its genomic information. This study focuses on developing an effective algorithm for protein function prediction. The algorithm is based on proteins that have similar expression patterns. The similarity of the expression data is determined using a novel measure, the slope matrix. The slope matrix introduces a normalized method for the comparison of expression levels throughout a proteome. The algorithm is tested using real microarray gene expression data. Their functions are characterized using gene ontology annotations. The results of the case study indicate the protein function prediction algorithm developed is comparable to the prediction algorithms that are based on the annotations of homologous proteins.
    [Show full text]
  • A Yeast Phenomic Model for the Gene Interaction Network Modulating
    Louie et al. Genome Medicine 2012, 4:103 http://genomemedicine.com/content/4/12/103 RESEARCH Open Access A yeast phenomic model for the gene interaction network modulating CFTR-ΔF508 protein biogenesis Raymond J Louie3†, Jingyu Guo1,2†, John W Rodgers1, Rick White4, Najaf A Shah1, Silvere Pagant3, Peter Kim3, Michael Livstone5, Kara Dolinski5, Brett A McKinney6, Jeong Hong2, Eric J Sorscher2, Jennifer Bryan4, Elizabeth A Miller3* and John L Hartman IV1,2* Abstract Background: The overall influence of gene interaction in human disease is unknown. In cystic fibrosis (CF) a single allele of the cystic fibrosis transmembrane conductance regulator (CFTR-ΔF508) accounts for most of the disease. In cell models, CFTR-ΔF508 exhibits defective protein biogenesis and degradation rather than proper trafficking to the plasma membrane where CFTR normally functions. Numerous genes function in the biogenesis of CFTR and influence the fate of CFTR-ΔF508. However it is not known whether genetic variation in such genes contributes to disease severity in patients. Nor is there an easy way to study how numerous gene interactions involving CFTR-ΔF would manifest phenotypically. Methods: To gain insight into the function and evolutionary conservation of a gene interaction network that regulates biogenesis of a misfolded ABC transporter, we employed yeast genetics to develop a ‘phenomic’ model, in which the CFTR-ΔF508-equivalent residue of a yeast homolog is mutated (Yor1-ΔF670), and where the genome is scanned quantitatively for interaction. We first confirmed that Yor1-ΔF undergoes protein misfolding and has reduced half-life, analogous to CFTR-ΔF. Gene interaction was then assessed quantitatively by growth curves for approximately 5,000 double mutants, based on alteration in the dose response to growth inhibition by oligomycin, a toxin extruded from the cell at the plasma membrane by Yor1.
    [Show full text]
  • WO 2012/174282 A2 20 December 2012 (20.12.2012) P O P C T
    (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 WO 2012/174282 A2 20 December 2012 (20.12.2012) P O P C T (51) International Patent Classification: David [US/US]; 13539 N . 95th Way, Scottsdale, AZ C12Q 1/68 (2006.01) 85260 (US). (21) International Application Number: (74) Agent: AKHAVAN, Ramin; Caris Science, Inc., 6655 N . PCT/US20 12/0425 19 Macarthur Blvd., Irving, TX 75039 (US). (22) International Filing Date: (81) Designated States (unless otherwise indicated, for every 14 June 2012 (14.06.2012) kind of national protection available): AE, AG, AL, AM, AO, AT, AU, AZ, BA, BB, BG, BH, BR, BW, BY, BZ, English (25) Filing Language: CA, CH, CL, CN, CO, CR, CU, CZ, DE, DK, DM, DO, Publication Language: English DZ, EC, EE, EG, ES, FI, GB, GD, GE, GH, GM, GT, HN, HR, HU, ID, IL, IN, IS, JP, KE, KG, KM, KN, KP, KR, (30) Priority Data: KZ, LA, LC, LK, LR, LS, LT, LU, LY, MA, MD, ME, 61/497,895 16 June 201 1 (16.06.201 1) US MG, MK, MN, MW, MX, MY, MZ, NA, NG, NI, NO, NZ, 61/499,138 20 June 201 1 (20.06.201 1) US OM, PE, PG, PH, PL, PT, QA, RO, RS, RU, RW, SC, SD, 61/501,680 27 June 201 1 (27.06.201 1) u s SE, SG, SK, SL, SM, ST, SV, SY, TH, TJ, TM, TN, TR, 61/506,019 8 July 201 1(08.07.201 1) u s TT, TZ, UA, UG, US, UZ, VC, VN, ZA, ZM, ZW.
    [Show full text]
  • Supplementary Material Contents
    Supplementary Material Contents Immune modulating proteins identified from exosomal samples.....................................................................2 Figure S1: Overlap between exosomal and soluble proteomes.................................................................................... 4 Bacterial strains:..............................................................................................................................................4 Figure S2: Variability between subjects of effects of exosomes on BL21-lux growth.................................................... 5 Figure S3: Early effects of exosomes on growth of BL21 E. coli .................................................................................... 5 Figure S4: Exosomal Lysis............................................................................................................................................ 6 Figure S5: Effect of pH on exosomal action.................................................................................................................. 7 Figure S6: Effect of exosomes on growth of UPEC (pH = 6.5) suspended in exosome-depleted urine supernatant ....... 8 Effective exosomal concentration....................................................................................................................8 Figure S7: Sample constitution for luminometry experiments..................................................................................... 8 Figure S8: Determining effective concentration .........................................................................................................
    [Show full text]
  • Genetic Identification of Brain Cell Types Underlying Schizophrenia
    bioRxiv preprint doi: https://doi.org/10.1101/145466; this version posted June 2, 2017. 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-NC-ND 4.0 International license. Genetic identification of brain cell types underlying schizophrenia Nathan G. Skene 1 †, Julien Bryois 2 †, Trygve E. Bakken3, Gerome Breen 4,5, James J Crowley 6, Héléna A Gaspar 4,5, Paola Giusti-Rodriguez 6, Rebecca D Hodge3, Jeremy A. Miller 3, Ana Muñoz-Manchado 1, Michael C O’Donovan 7, Michael J Owen 7, Antonio F Pardiñas 7, Jesper Ryge 8, James T R Walters 8, Sten Linnarsson 1, Ed S. Lein 3, Major Depressive Disorder Working Group of the Psychiatric Genomics Consortium, Patrick F Sullivan 2,6 *, Jens Hjerling- Leffler 1 * Affiliations: 1 Laboratory of Molecular Neurobiology, Department of Medical Biochemistry and Biophysics, Karolinska Institutet, SE-17177 Stockholm, Sweden. 2 Department of Medical Epidemiology and Biostatistics, Karolinska Institutet, SE-17177 Stockholm, Sweden. 3 Allen Institute for Brain Science, Seattle, Washington 98109, USA. 4 King’s College London, Institute of Psychiatry, Psychology and Neuroscience, MRC Social, Genetic and Developmental Psychiatry (SGDP) Centre, London, UK. 5 National Institute for Health Research Biomedical Research Centre, South London and Maudsley National Health Service Trust, London, UK. 6 Departments of Genetics, University of North Carolina, Chapel Hill, NC, 27599-7264, USA. 7 MRC Centre for Neuropsychiatric Genetics and Genomics, Institute of Psychological Medicine and Clinical Neurosciences, School of Medicine, Cardiff University, Cardiff, UK.
    [Show full text]
  • Genome-Wide Association Study Identifies Genetic Susceptibility Loci
    Yang et al. J Transl Med (2020) 18:224 https://doi.org/10.1186/s12967-020-02390-0 Journal of Translational Medicine RESEARCH Open Access Genome-wide association study identifes genetic susceptibility loci and pathways of radiation-induced acute oral mucositis Da‑Wei Yang1,2†, Tong‑Min Wang1†, Jiang‑Bo Zhang1, Xi‑Zhao Li1, Yong‑Qiao He1, Ruowen Xiao1, Wen‑Qiong Xue1, Xiao‑Hui Zheng1, Pei‑Fen Zhang1, Shao‑Dan Zhang1, Ye‑Zhu Hu1, Guo‑Ping Shen3, Mingyuan Chen1,4, Ying Sun1,5 and Wei‑Hua Jia1,2,6* Abstract Background: Radiation‑induced oral mucositis (OM) is one of the most common acute complications for head and neck cancer. Severe OM is associated with radiation treatment breaks, which harms successful tumor management. Radiogenomics studies have indicated that genetic variants are associated with adverse efects of radiotherapy. Methods: A large‑scale genome‑wide scan was performed in 1467 nasopharyngeal carcinoma patients, including 753 treated with 2D‑CRT from Genetic Architecture of the Radiotherapy Toxicity and Prognosis (GARTP) cohort and 714 treated with IMRT (192 from the GARTP and 522 newly recruited). Subgroup analysis by radiotherapy technique was further performed in the top associations. We also performed physical and regulatory mapping of the risk loci and gene set enrichment analysis of the candidate target genes. Results: We identifed 50 associated genomic loci and 64 genes via positional mapping, expression quantitative trait locus (eQTL) mapping, chromatin interaction mapping and gene‑based analysis, and 36 of these loci were replicated in subgroup analysis. Interestingly, one of the top loci located in TNKS, a gene relevant to radiation toxicity, was associ‑ 6 ated with increased OM risk with OR 3.72 of the lead SNP rs117157809 (95% CI 2.10–6.57; P 6.33 10− ).
    [Show full text]
  • Genomic Characterization of the Adolescent Idiopathic Scoliosis Associated Transcriptome and Regulome
    bioRxiv preprint doi: https://doi.org/10.1101/2020.03.02.973735; this version posted March 4, 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. Genomic characterization of the adolescent idiopathic scoliosis associated transcriptome and regulome Nadja Makki1*, Jingjing Zhao2,3, Zhaoyang Liu4, Walter L. Eckalbar2,3, Aki Ushiki2,3, Anas M. Khanshour5, Zhuoxi Wu1, Jonathan Rios5,6, Ryan S. Gray4, Carol A. Wise5,6, Nadav Ahituv2,3* 1Department of Anatomy and Cell Biology, University of Florida, College of Medicine, Gainesville, Florida, USA. 2Department of Bioengineering and Therapeutic Sciences, University of California San Francisco, San Francisco, California, USA. 3Institute for Human Genetics, University of California San Francisco, San Francisco, California, USA. 4Department of Pediatrics and Nutritional Sciences, Dell Pediatric Research Institute, University of Texas at Austin, Austin, Texas, USA. 5 Center for Pediatric Bone Biology and Translational Research, Texas Scottish Rite Hospital for Children, Dallas, Texas, USA. 6Departments of Orthopaedic Surgery, Pediatrics, and McDermott Center for Human Growth and Development, University of Texas Southwestern Medical Center, Dallas, USA. *Corresponding authors: [email protected]; [email protected] Key words: scoliosis, gene networks, gene regulation, RNA-seq, ChIP-seq Running Title: Regulatory networks underlying adolescent idiopathic scoliosis bioRxiv preprint doi: https://doi.org/10.1101/2020.03.02.973735; this version posted March 4, 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. Abstract Adolescent idiopathic scoliosis (AIS), a sideways curvature of the spine, is the most common pediatric musculoskeletal disorder, affecting ~3% of the population worldwide.
    [Show full text]
  • Autocrine IFN Signaling Inducing Profibrotic Fibroblast Responses By
    Downloaded from http://www.jimmunol.org/ by guest on September 23, 2021 Inducing is online at: average * The Journal of Immunology , 11 of which you can access for free at: 2013; 191:2956-2966; Prepublished online 16 from submission to initial decision 4 weeks from acceptance to publication August 2013; doi: 10.4049/jimmunol.1300376 http://www.jimmunol.org/content/191/6/2956 A Synthetic TLR3 Ligand Mitigates Profibrotic Fibroblast Responses by Autocrine IFN Signaling Feng Fang, Kohtaro Ooka, Xiaoyong Sun, Ruchi Shah, Swati Bhattacharyya, Jun Wei and John Varga J Immunol cites 49 articles Submit online. Every submission reviewed by practicing scientists ? is published twice each month by Receive free email-alerts when new articles cite this article. Sign up at: http://jimmunol.org/alerts http://jimmunol.org/subscription Submit copyright permission requests at: http://www.aai.org/About/Publications/JI/copyright.html http://www.jimmunol.org/content/suppl/2013/08/20/jimmunol.130037 6.DC1 This article http://www.jimmunol.org/content/191/6/2956.full#ref-list-1 Information about subscribing to The JI No Triage! Fast Publication! Rapid Reviews! 30 days* Why • • • Material References Permissions Email Alerts Subscription Supplementary The Journal of Immunology The American Association of Immunologists, Inc., 1451 Rockville Pike, Suite 650, Rockville, MD 20852 Copyright © 2013 by The American Association of Immunologists, Inc. All rights reserved. Print ISSN: 0022-1767 Online ISSN: 1550-6606. This information is current as of September 23, 2021. The Journal of Immunology A Synthetic TLR3 Ligand Mitigates Profibrotic Fibroblast Responses by Inducing Autocrine IFN Signaling Feng Fang,* Kohtaro Ooka,* Xiaoyong Sun,† Ruchi Shah,* Swati Bhattacharyya,* Jun Wei,* and John Varga* Activation of TLR3 by exogenous microbial ligands or endogenous injury-associated ligands leads to production of type I IFN.
    [Show full text]
  • Individual Functions and Substrate Specificities of Importin Alpha
    From the Max-Delbrück-Center for Molecular Medicine Berlin Head of the research group: Prof. Dr. M. Bader “Individual functions and substrate specificities of importin α subtypes” Dissertation for Fulfillment of Requirements for the Doctoral Degree of the University of Lübeck from the Department of Natural Sciences Submitted by Stefanie Hügel from Hennigsdorf Lübeck 2013 First referee: Prof. Dr. M. Bader Second referee: Prof. Dr. N. Tautz Date of oral examination: 21.02.13 Approved for printing. Lübeck, 22.02.2013 2 1 TABLE OF CONTENTS 1 TABLE OF CONTENTS _______________________________________________________ 3 2 ABSTRACT _______________________________________________________________ 6 3 ZUSAMMENFASSUNG ______________________________________________________ 7 4 INTRODUCTION ___________________________________________________________ 9 4.1 Nucleocytoplasmic protein transport ____________________________________________ 9 4.1.1 The nuclear pore complex – gateway to the nucleus ______________________________________ 10 4.1.1 Transport factors __________________________________________________________________ 11 4.1.2 Nothing happens without RanGTP _____________________________________________________ 12 4.2 The classical nuclear protein import pathway ____________________________________ 13 4.2.1 Structure and function of importin α ___________________________________________________ 13 4.2.2 Importin α:β mediated nuclear protein import___________________________________________ 15 4.2.3 The importin α protein family ________________________________________________________
    [Show full text]
  • Prioritizing Candidate Genes Post-GWAS Using Multiple Sources
    Cai et al. BMC Genomics (2018) 19:656 https://doi.org/10.1186/s12864-018-5050-x RESEARCHARTICLE Open Access Prioritizing candidate genes post-GWAS using multiple sources of data for mastitis resistance in dairy cattle Zexi Cai* , Bernt Guldbrandtsen, Mogens Sandø Lund and Goutam Sahana Abstract Background: Improving resistance to mastitis, one of the costliest diseases in dairy production, has become an important objective in dairy cattle breeding. However, mastitis resistance is influenced by many genes involved in multiple processes, including the response to infection, inflammation, and post-infection healing. Low genetic heritability, environmental variations, and farm management differences further complicate the identification of links between genetic variants and mastitis resistance. Consequently, studies of the genetics of variation in mastitis resistance in dairy cattle lack agreement about the responsible genes. Results: We associated 15,552,968 imputed whole-genome sequencing markers for 5147 Nordic Holstein cattle with mastitis resistance in a genome-wide association study (GWAS). Next, we augmented P-values for markers in genes in the associated regions using Gene Ontology terms, Kyoto Encyclopedia of Genes and Genomes pathway analysis, and mammalian phenotype database. To confirm results of gene-based analyses, we used gene expression data from E. coli-challenged cow udders. We identified 22 independent quantitative trait loci (QTL) that collectively explained 14% of the variance in breeding values for resistance to clinical mastitis (CM). Using association test statistics with multiple pieces of independent information on gene function and differential expression during bacterial infection, we suggested putative causal genes with biological relevance for 12 QTL affecting resistance to CM in dairy cattle.
    [Show full text]
  • A CRISPR Screen to Identify Combination Therapies of Cytotoxic
    CRISPRi Screens to Identify Combination Therapies for the Improved Treatment of Ovarian Cancer By Erika Daphne Handly B.S. Chemical Engineering Brigham Young University, 2014 Submitted to the Department of Biological Engineering in partial fulfillment of the requirements for the degree of Doctor of Philosophy in Biological Engineering at the MASSACHUSETTS INSTITUTE OF TECHNOLOGY February 2021 © 2020 Massachusetts Institute of Technology. All rights reserved. Signature of author………………………………………………………………………………… Erika Handly Department of Biological Engineering February 2021 Certified by………………………………………………………………………………………… Michael Yaffe Director MIT Center for Precision Cancer Medicine Department of Biological Engineering and Biology Thesis Supervisor Accepted by………………………………………………………………………………………... Katharina Ribbeck Professor of Biological Engineering Chair of Graduate Program, Department of Biological Engineering Thesis Committee members Michael T. Hemann, Ph.D. Associate Professor of Biology Massachusetts Institute of Technology Douglas A. Lauffenburger, Ph.D. (Chair) Ford Professor of Biological Engineering, Chemical Engineering, and Biology Massachusetts Institute of Technology Michael B. Yaffe, M.D., Ph.D. (Thesis Supervisor) David H. Koch Professor of Science Prof. of Biology and Biological Engineering Massachusetts Institute of Technology 2 CRISPRi Screens to Identify Combination Therapies for the Improved Treatment of Ovarian Cancer By Erika Daphne Handly B.S. Chemical Engineering Brigham Young University, 2014 Submitted to the Department of Biological Engineering in partial fulfillment of the requirements for the degree of Doctor of Philosophy in Biological Engineering ABSTRACT Ovarian cancer is the fifth leading cause of cancer death for women in the United States, with only modest improvements in patient survival in the past few decades. Standard-of-care consists of surgical debulking followed by a combination of platinum and taxane agents, but relapse and resistance frequently occur.
    [Show full text]
  • A Snapshot of the Physical and Functional Wiring of the Eps15 Homology Domain Network in the Nematode
    View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by AIR Universita degli studi di Milano A Snapshot of the Physical and Functional Wiring of the Eps15 Homology Domain Network in the Nematode Hanako Tsushima1.¤a, Maria Grazia Malabarba1,2., Stefano Confalonieri1, Francesca Senic-Matuglia1, Lisette G. G. C. Verhoef1, Cristina Bartocci1¤b, Giovanni D’Ario1, Andrea Cocito1, Pier Paolo Di Fiore1,2,3*, Anna Elisabetta Salcini1,4* 1 IFOM, Fondazione Istituto FIRC di Oncologia Molecolare, Milan, Italy, 2 Dipartimento di Medicina, Chirurgia ed Odontoiatria, Universita` degli Studi di Milano, Milan, Italy, 3 Istituto Europeo di Oncologia, Milan, Italy, 4 Biotech Research and Innovation Centre (BRIC), University of Copenhagen, Copenhagen, Denmark Abstract Protein interaction modules coordinate the connections within and the activity of intracellular signaling networks. The Eps15 Homology (EH) module, a protein-protein interaction domain that is a key feature of the EH-network, was originally identified in a few proteins involved in endocytosis and vesicle trafficking, and has subsequently also been implicated in actin reorganization, nuclear shuttling, and DNA repair. Here we report an extensive characterization of the physical connections and of the functional wirings of the EH-network in the nematode. Our data show that one of the major physiological roles of the EH-network is in neurotransmission. In addition, we found that the proteins of the network intersect, and possibly coordinate, a number of ‘‘territories’’ of cellular activity including endocytosis/recycling/vesicle transport, actin dynamics, general metabolism and signal transduction, ubiquitination/degradation of proteins, DNA replication/repair, and miRNA biogenesis and processing.
    [Show full text]