Differential chromatin proteomics of the MMS-induced DNA damage response in yeast

Proteome Science

Brendan J McConkey¹  Bernard P Duncker¹  Brian P Ingalls²  Rohan D Gidvani¹  Dong Ryoung Kim¹ 

[1] Department of Biology, University of Waterloo, 200 University Avenue, Waterloo, ON, Canada;Department of Applied Mathematics, University of Waterloo, 200 University Avenue, Waterloo, ON, Canada

关键词: DNA damage;    MMS;    proteomics;    differential;    DIGE;    fractionation;    chromatin;   

Others  :  820246
DOI  :  10.1186/1477-5956-9-62

 received in 2011-05-05, accepted in 2011-10-04,  发布年份 2011

【 摘 要 】

Background

Protein enrichment by sub-cellular fractionation was combined with differential-in-gel-electrophoresis (DIGE) to address the detection of the low abundance chromatin proteins in the budding yeast proteome. Comparisons of whole-cell extracts and chromatin fractions were used to provide a measure of the degree of chromatin association for individual proteins, which could be compared across sample treatments. The method was applied to analyze the effect of the DNA damaging agent methyl methanesulfonate (MMS) on levels of chromatin-associated proteins.

Results

Up-regulation of several previously characterized DNA damage checkpoint-regulated proteins, such as Rnr4, Rpa1 and Rpa2, was observed. In addition, several novel DNA damage responsive proteins were identified and assessed for genotoxic sensitivity using either DAmP (decreased abundance by mRNA perturbation) or knockout strains, including Acf2, Arp3, Bmh1, Hsp31, Lsp1, Pst2, Rnr4, Rpa1, Rpa2, Ste4, Ycp4 and Yrb1. A strain in which the expression of the Ran-GTPase binding protein Yrb1 was reduced was found to be hypersensitive to genotoxic stress.

Conclusion

The described method was effective at unveiling chromatin-associated proteins that are less likely to be detected in the absence of fractionation. Several novel proteins with altered chromatin abundance were identified including Yrb1, pointing to a role for this nuclear import associated protein in DNA damage response.

【 授权许可】

   
2011 Kim et al; licensee BioMed Central Ltd.

【 图 表 】

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【 参考文献 】

• [1]Tonge R, Shaw J, Middleton B, Rowlinson R, Rayner S, Young J, Pognan F, Hawkins E, Currie I, Davison M: Validation and development of fluorescence two-dimensional differential gel electrophoresis proteomics technology. Proteomics 2001, 1:377-396.
• [2]Aerbersold R, Mann M: Mass spectrometry-based proteomics. Nature 2003, 422:198-207.
• [3]Ghaemmaghami S, Huh W-K, Bower K, Howson RW, Belle A, Dephoure N, O'shea EK, Weissman JS: Global analysis of protein expression in yeast. Nature 2003, 425:737-741.
• [4]Gavin AC, Bösche M, Krause R, Grandi P, Marzioch M, Bauer A, Schultz J, Rick JM, Michon AM, Cruciat CM, Remor M, Höfert C, Schelder M, Brajenovic M, Ruffner H, Merino A, Klein K, Hudak M, Dickson D, Rudi T, Gnau V, Bauch A, Bastuck S, Huhse B, Leutwein C, Heurtier MA, Copley RR, Edelmann A, Querfurth E, Rybin V, Drewes G, Raida M, Bouwmeester T, Bork P, Seraphin B, Kuster B, Neubauer G, Superti-Furga G: Functional organization of yeast proteome by systematic analysis of protein complexes. Nature 2002, 415:141-147.
• [5]Smolka MB, Albuquerque CP, Chen S-H, Zhou H: Proteome-wide identification of in vivo targets of DNA damage checkpoint kinases. Proc Natl Acad Sci USA 2007, 104:10364-10369.
• [6]Yates JR III, Gilchrist A, Howell KE, Bergeron JJ: Proteomics of organelles and large cellular structure. Nat Rev Mol Cell Biol 2005, 6:702-714.
• [7]Forner F, Foster LJ, Campanaro S, Valle G, Mann M: Quantitative proteomic comparison of rat mitochondria from muscle, heart, and liver. Mol Cell Proteomics 2006, 5:608-619.
• [8]Andersen J, Mann M: Organellar proteomics: turning inventories into insights. EMBO rep 2006, 9:874-879.
• [9]Rattner JB, Saunders C, Davie JR, Hamkalo BA: Ultrastructural organization of yeast chromatin. J Cell Biol 1982, 92:217-222.
• [10]Szent-Gyorgyi C, Isenberg I: The organization of oligonucleosome in yeast. Nucleic Acids Res 1983, 11:3717-3736.
• [11]Liang C, Stillman B: Persistent inhibition of DNA replication and chromatin-bound MCM proteins during the cell cycle in cdc6 mutants. Genes Dev 1997, 11:3375-3386.
• [12]Khoudoli GA, Gillespie PJ, Stewart G, Andersen JS, Swedlow JR, Blow JJ: Temporal profiling of the chromatin proteome reveals system-wide responses to replication inhibition. Curr Biol 2008, 18:838-843.
• [13]Chu DS, Liu H, Nix P, Wu TF, Ralston EJ, Yates JR, Meyer BJ: Sperm chromatin proteomics identifies evolutionarily conserved fertility factors. Nature 2006, 443:101-105.
• [14]Tang W, Deng Z, Oses-Prieto JA, Suzuki N, Zhu S, Zhang X, Burlingame AL, Wang ZY: Proteomics studies of brassinosteroid signal transduction using prefractionation and two-dimensional DIGE. Mol Cell Proteomics 2008, 7:728-738.
• [15]Bernstein BE, Schreiber SL: Global approaches to chromatin. Chem Biol 2002, 9:1167-1173.
• [16]Sclafani RA, Holzen TM: Cell cycle regulation of DNA replication. Annu Rev Genet 2007, 41:237-280.
• [17]Morgan DO, Loog M: Cyclin specificity in the phosphorylation of cyclin-dependent kinase substrates. Nature 2005, 434:104-108.
• [18]Koundrioukoff S, Polo S, Almouzni G: Interplay between chromatin and cell cycle checkpoints in the context of ATR/ATM-dependent checkpoints. DNA repair 2004, 3:969-978.
• [19]Harrison JC, Haber JE: Surviving the breakup: the DNA damage checkpoint. Annu Rev Genet 2006, 40:209-235.
• [20]Jiang Y-H, Bressler J, Beaudet AL: Epigenetics and human disease. Annu Rev Genomics Hum Genet 2004, 5:479-510.
• [21]Pollard KJ, Peterson CL: Chromatin remodeling: a marriage between two families. Bioessay 1998, 20:771-780.
• [22]Rodriguez BAT, Huang TDM: Tilling the chromatin landscape: emerging methods for the discovery and profiling of protein DNA interactions. Biochem Cell Biol 2005, 83:525-534.
• [23]Saha A, Wittmeyer J, Cairns BR: Chromatin remodeling: the industrial revolution of DNA around histones. Nature Rev Mol Cell Biol 2006, 7:437-447.
• [24]Lalo D, Carles C, Sentenac A, Thuriaux P: Interactions between three common subunits of yeast RNA polymerase I and III. Proc Natl Acad Sci USA 1993, 90:5524-5528.
• [25]Jourdain S, Acker J, Ducrot C, Sentenac A, Lefebvre O: The tau95 subunit of yeast TFIIIC influences upstream and downstream functions of TFIIIC. DNA complexes. J Biol Chem 2003, 278:10450-10457.
• [26]Nelson LD, Musso M, Van Dyke MW: The yeast STM1 gene encodes a purine motif triple helical DNA-binding protein. J Biol Chem 2000, 275:5573-5581.
• [27]Downey M, Houlsworth R, Maringele L, Rollie A, Brehme M, Galicia S, Cuillard S, Partington M, Zubko ML, Krogan NJ, Emili A, Greenblatt JF, Harrington L, Lydall D, Durocher D: A genome-wide screen identifies the evolutionary conserved KEOPS complex as a telomere regulator. Cell 2006, 124:1155-1168.
• [28]Longhese MP, Neecke H, Paciotti V, Lucchini G, Plevani P: The 70 kDa subunit of replication protein A is required for the G1/S and intra-S DNA damage checkpoints in budding yeast. Nucleic Acids Res 1996, 24:3533-3537.
• [29]Umezu K, Sugawara N, Chen C, Haber JE, Kolodner RD: Genetic analysis of yeast RPA1 reveals its multiple functions in DNA metabolism. Genetics 1998, 148:989-1005.
• [30]Santocanale C, Neecke H, Longhese MP, Lucchini G, Plevani P: Mutations in the gene encoding the 34 kDa subunit of yeast replication protein A cause defective S phase progression. J Mol Biol 1995, 254:595-607.
• [31]Huang M, Elledge SJ: Identification of RNR4, encoding a second essential small subunit of ribonucleotide reductase in Saccharomyces cerevisiae. Mol Cell Biol 1997, 17:6105-6113.
• [32]Yao R, Zhang Z, An X, Bucci B, Perlstein DL, Stubble J, Huang M: Subcellular localization of yeast ribonucleotide reductase regulated by the DNA replication and damage checkpoint pathways. Proc Natl Acad Sci USA 2003, 100:6628-6633.
• [33]Usui T, Petrini JHJ: The Saccharomyces cerevisiae 14-3-3 proteins Bmh1 and Bmh2 directly influence the DNA damage-dependent functions of Rad53. Proc Natl Acad Sci USA 2003, 104:2797-2802.
• [34]Liao C, Hu B, Arno MJ, Panaretou B: Genomic screening in vivo reveals the role played by vacuolar H+ ATPase and cytosolic acidification in sensitivity to DNA-damaging agents such as cisplatin. Mol Pharmacol 2006, 71:416-425.
• [35]Hinnebusch AG, Natarajan K: Gcn4p, a master regulator of gene expression, is controlled at multiple levels by diverse signals of starvation and stress. Eukaryot Cell 2002, 1:22-32.
• [36]Norman JO, Joe CO, Busbee DL: Inhibition of DNA polymerase activity by methyl methanesulfonate. Mutat Res 1986, 165:71-79.
• [37]Shin I, Kam Y, Ha K, Kang K, Joe CO: Inhibition of the phosphorylation of a myristoylated alanine-rich C kinase substrate by methyl methanesulfonate in cultured NIH 3T3 cells. Mutat Res 1996, 351:163-171.
• [38]Birrell GW, Brown JA, Wu HI, Giaever G, Chu AM, Davis RW, Brown JM: Transcriptional response of Saccharomyces cerevisiae to DNA-damaging agents does not identify the genes that protect against these agents. Proc Natl Acad Sci USA 2002, 99:8778-8783.
• [39]Hillenmeyer ME, Fung E, Wildenhain J, Pierce SE, Hoon S, Lee W, Proctor M, St Onge RP, Tyers M, Koller D, Altman RB, Davis RW, Nislow C, Giaever G: The chemical genomic portrait of yeast: uncovering a phenotype for all genes. Science 2008, 320:362-365.
• [40]Varrin AE, Prasad AA, Scholz R-P, Ramer MD, Duncker BP: A mutation in Dbf4 motif M impairs interactions with DNA replication factors and confers increased resistance to genotoxic agents. Mol Cell Biol 205, 25:7494-7504.
• [41]Hanway D, Chin JK, Xia G, Oshiro G, Winzeler EA, Romesberg FE: Previously uncharacterized genes in the UV- and MMS-induced DNA damage response in yeast. Proc Natl Acad Sci USA 2002, 99:10605-10610.
• [42]Santocanale C, Diffley JFX: A Mec1- and Rad53-dependent checkpoint controls late-firing origins of DNA replication. Nature 1998, 395:615-618.
• [43]Künzler M, Trueheart J, Sette C, Hurt E, Thorner J: Mutations in the YRB1 gene encoding yeast Ran-binding-protein-1 that impair nucelocytoplasmic transport and suppress yeast mating defects. Genetics 2001, 157:1089-1105.
• [44]Ong S-E, Mann M: Mass spectrometry-based proteomics turns quantitative. Nat Chem Biol 2005, 1:252-262.
• [45]Gasch AP, Huang M, Metzner S, Botstein D, Elledge SJ, Brown PO: Genomic expression response to DNA-damaging agents and the regulatory role of the yeast ATR homolog Mec1p. Mol Biol Cell 2001, 12:2987-3003.
• [46]Begley TJ, Rosenbach AS, Ideker T, Samson LD: Damage recovery pathways in Saccharomyces cerevisiae revealed by genomic phenotyping and interactome mapping. Mol Cancer Res 2002, 1:103-112.
• [47]Ghavidel A, Kislinger T, Pogoutse O, Sopko R, Jurisica I, Emili A: Impaired tRNA nuclear export links DNA damage and cell-cycle checkpoint. Cell 2007, 131:915-926.
• [48]Karp NA, McCormick PS, Russell MR, Lilley KS: Experimental and statistical considerations to avoid false conclusions in proteomics studies using differential in-gel electrophoresis. Mol Cell Proteomics 2007, 6:1354-1364.
• [49]Molloy MP, Brzezinski EE, Hang J, McDowell MT, VanBogelen RA: Overcoming technical variation and biological variation in quantitative proteomics. Proteomics 2003, 3:1912-1919.
• [50]Team RDC: R: A Language and Environment for Statistical Computing. R Foundation for Statistical Computing. [http://www.r-project.org] webcite
• [51]Candiano G, Bruschi M, Musante L, Ghiggeri GM, Carnemolla B, Orecchia P, Zardi L, Righetti PG: Blue silver: a very sensitive colloidal Coomassie G-250 staining for proteome analysis. Electrophoresis 2004, 25:1327-1333.
• [52]Granvogl B, Plöcher M, Eichacker LA: Sample preparation by in-gel digestion for mass spectrometry-based proteomics. Anal Bioanal Chem 2007, 389:991-1002.
• [53]Ma B, Zhang K, Liang C: An effective algorithm for peptide de novo sequencing from MS/MS spectra. J Comput System Sci 2005, 70:418-430.
• [54]Yoo YD, Xu X, Guan J-L: A novel role of the actin-nucleating Arp2/3 complex in the regulation of RNA polymerase II-dependent transcription. J Biol Chem 2006, 282:7616-7623.
• [55]Franke J, Reimann B, Hartmann E, Köhlerl M, Wiedmann B: Evidence for a nuclear passage of nascent polypeptide-associated complex subunits in yeast. J Cell Sci 2001, 114:2641-2648.
• [56]Valencia-Burton M, Oki M, Johnson J, Seier TA, Kamakaka R, Haber JE: Different mating-type-regulated genes affect the DNA repair defects of Saccharomyces Rad51, Rad52 and Rad55 mutants. Genetics 2006, 174:41-55.
• [57]Murray JA, Scarpa M, Rossi N, Cesareni G: Antagonistic controls regulate copy number of the yeast mu plasmid. EMBO J 1987, 6:4205-4212.
• [58]Huh WK, Falvo JV, Gerke LC, Carroll AS, Howson RW, Weissman JS, O'Shea EK: Global analysis of protein localization in budding yeast. Nature 2003, 425:686-691.