Portal de Periódicos da CAPES

Nuclear localization of the human mutY homologue hMYH

2000; Wiley; Volume: 77; Issue: 4 Linguagem: Inglês

10.1002/(sici)1097-4644(20000615)77

1097-4644

Jyy-Jih Tsai-Wu, Ho‐Ting Su, Yalei Wu, Su-Ming Hsu, Chih‐Hsiung Wu,

DNA Repair Mechanisms

Journal of Cellular BiochemistryVolume 77, Issue 4 p. 666-677 Article Nuclear localization of the human mutY homologue hMYH Jyy-Jih Tsai-Wu, Jyy-Jih Tsai-Wu Department of Clinical Research, National Taiwan University Hospital, National Taiwan University College of Medicine, Taipei, Taiwan, Republic of China Jyy-Jih Tsai-Wu and Ho-Ting Su contributed equally to this work.Search for more papers by this authorHo-Ting Su, Ho-Ting Su Institute of Molecular Medicine, National Taiwan University College of Medicine, Taipei, Taiwan, Republic of China Jyy-Jih Tsai-Wu and Ho-Ting Su contributed equally to this work.Search for more papers by this authorYa-Lei Wu, Ya-Lei Wu Department of Pathology, National Taiwan University College of Medicine, Taipei, Taiwan, Republic of ChinaSearch for more papers by this authorSu-Ming Hsu, Su-Ming Hsu Department of Pathology, National Taiwan University College of Medicine, Taipei, Taiwan, Republic of ChinaSearch for more papers by this authorC.H. Herbert Wu, Corresponding Author C.H. Herbert Wu [email protected] Institute of Molecular Medicine, National Taiwan University College of Medicine, Taipei, Taiwan, Republic of ChinaInstitute of Molecular Medicine, National Taiwan University College of Medicine, 7 Chung-Shan S. Road, Taipei, Taiwan 100, Republic of ChinaSearch for more papers by this author Jyy-Jih Tsai-Wu, Jyy-Jih Tsai-Wu Department of Clinical Research, National Taiwan University Hospital, National Taiwan University College of Medicine, Taipei, Taiwan, Republic of China Jyy-Jih Tsai-Wu and Ho-Ting Su contributed equally to this work.Search for more papers by this authorHo-Ting Su, Ho-Ting Su Institute of Molecular Medicine, National Taiwan University College of Medicine, Taipei, Taiwan, Republic of China Jyy-Jih Tsai-Wu and Ho-Ting Su contributed equally to this work.Search for more papers by this authorYa-Lei Wu, Ya-Lei Wu Department of Pathology, National Taiwan University College of Medicine, Taipei, Taiwan, Republic of ChinaSearch for more papers by this authorSu-Ming Hsu, Su-Ming Hsu Department of Pathology, National Taiwan University College of Medicine, Taipei, Taiwan, Republic of ChinaSearch for more papers by this authorC.H. Herbert Wu, Corresponding Author C.H. Herbert Wu [email protected] Institute of Molecular Medicine, National Taiwan University College of Medicine, Taipei, Taiwan, Republic of ChinaInstitute of Molecular Medicine, National Taiwan University College of Medicine, 7 Chung-Shan S. Road, Taipei, Taiwan 100, Republic of ChinaSearch for more papers by this author First published: 14 April 2000 https://doi.org/10.1002/(SICI)1097-4644(20000615)77:4 3.0.CO;2-XCitations: 26 AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onEmailFacebookTwitterLinkedInRedditWechat Abstract The cDNA of the human mutY homologue (hMYH) was cloned from the total RNA of the tumor cell line SU-DHL-1 by reverse transcription-polymerase chain reaction (RT-PCR). Expression of hMYH in a plasmid can partially revert the mutator phenotype of the Escherichia coli mutY mutant MK609(DE3). The majority of the recombinant hMYH protein in E. coli was precipitated in the inclusion bodies. A minor fraction of the soluble recombinant protein was concentrated as the source of the protein in the activity assay. Recombinant hMYH displayed both glycosylase and AP lyase activity. Three independent rabbit antisera against an N-terminal peptide, HY90, a recombinant C-terminal fragment, and the full-length hMYH recombinant protein were prepared and affinity-purified, and these antisera recognized the 59 kDa endogenous hMYH protein in HeLa cells. Immunofluorescent staining experiments with these three antisera showed a consistent nuclear distribution of hMYH, excluding the nucleoli. This nuclear staining pattern was abolished if the antisera were incubated with specific peptide/protein competitors, whereas the staining pattern was unaffected if the antisera were incubated with nonspecific peptide competitors. Consistent with the immunofluorescent staining results, a flag-tagged transfected hMYH also showed a nuclear staining pattern excluding the nucleoli. These results suggest that hMYH is indeed a functional homologue of E. coli MutY and is localized in the nuclei of mammalian cells. J. Cell. Biochem. 77:666–677, 2000. © 2000 Wiley-Liss, Inc. REFERENCES Aburatani H, Hippo Y, Ishida T, Takashima R, Matsuba C, Kodama T, Takao M, Yasui A, Yamamoto K, Asano M. 1997. Cloning and characterization of mammalian 8-hydroxyguanine-specific DNA glycosylase/apurinic, apyrimidinic lyase, a functional mutM homologue. Cancer Res 57: 2151–2156. CASPubMedWeb of Science®Google Scholar Ames BN. 1989. Endogenous oxidative DNA damage, aging, and cancer. Free Radic Res Commun 7: 121–128. 10.3109/10715768909087933 CASPubMedWeb of Science®Google Scholar Ames BN, Gold LS. 1991. Endogenous mutagens and the causes of aging and cancer. Mutat Res 250: 3–16. 10.1016/0027-5107(91)90157-J CASPubMedWeb of Science®Google Scholar Boiteux S, O'Connor TR, Lederer F, Gouyette A, Laval J. 1990. Homogeneous Escherichia coli FPG protein. A DNA glycosylase which excises imidazole ring-opened purines and nicks DNA at apurinic/apyrimidinic sites. J Biol Chem 265: 3916–3922. CASPubMedWeb of Science®Google Scholar Cabrera M, Nghiem Y, Miller JH. 1988. mutM, a second mutator locus in Escherichia coli that generates G.C—T.A transversions. J Bacteriol 170: 5405–5407. 10.1128/jb.170.11.5405-5407.1988 CASPubMedWeb of Science®Google Scholar Cheng KC, Cahill DS, Kasai H, Nishimura S, Loeb LA. 1992. 8-Hydroxyguanine, an abundant form of oxidative DNA damage, causes G—T and A—C substitutions. J Biol Chem 267: 166–172. 10.1016/S0021-9258(18)48474-8 CASPubMedWeb of Science®Google Scholar Chetsanga CJ, Lindahl T. 1979. Release of 7-methylguanine residues whose imidazole rings have been opened from damaged DNA by a DNA glycosylase from Escherichia coli. Nucleic Acids Res 6: 3673–3684. 10.1093/nar/6.11.3673 CASPubMedWeb of Science®Google Scholar Feng XH, Filvaroff EH, Derynck R. 1995. Transforming growth factor-beta (TGF-beta)-induced down-regulation of cyclin A expression requires a functional TGF-beta receptor complex. Characterization of chimeric and truncated type I and type II receptors. J Biol Chem 270: 24237–24245. 10.1074/jbc.270.41.24237 CASPubMedWeb of Science®Google Scholar Fraga CG, Shigenaga MK, Park JW, Degan P, Ames BN. 1990. Oxidative damage to DNA during aging: 8-hydroxy-2′-deoxyguanosine in rat organ DNA and urine. Proc Natl Acad Sci USA 87: 4533–4537. 10.1073/pnas.87.12.4533 CASPubMedWeb of Science®Google Scholar Gogos A, Cillo J, Clarke ND, Lu AL. 1996. Specific recognition of A/G and A/7,8-dihydro-8-oxoguanine (8-oxoG) mismatches by Escherichia coli MutY: removal of the C-terminal domain preferentially affects A/8-oxoG recognition. Biochemistry 35: 16665–16671. 10.1021/bi960843w CASPubMedWeb of Science®Google Scholar Halliwell B, Gutteridge JMC. 1999. Free radicals in biology and medicine. New York: Oxford University Press. Google Scholar Harlow E, Lane D. 1988. Antibodies: a laboratory manual. Cold Spring Harbor, New York: Cold Spring Harbor Laboratory Press. CASPubMedWeb of Science®Google Scholar Kuo CF, McRee DE, Fisher CL, O'Handley SF, Cunningham RP, Tainer JA. 1992. Atomic structure of the DNA repair [4Fe-4S] enzyme endonuclease III. Science 258: 434–440. 10.1126/science.1411536 CASPubMedWeb of Science®Google Scholar Labahn J, Scharer OD, Long A, Ezaz-Nikpay K, Verdine GL, Ellenberger TE. 1996. Structural basis for the excision repair of alkylation-damaged DNA. Cell 86: 321–329. 10.1016/S0092-8674(00)80103-8 CASPubMedWeb of Science®Google Scholar Lu AL, Fawcett WP. 1998. Characterization of the recombinant MutY homolog, an adenine DNA glycosylase, from yeast Schizosaccharomyces pombe. J Biol Chem 273: 25098–25105. 10.1074/jbc.273.39.25098 CASPubMedWeb of Science®Google Scholar Lu AL, Yuen DS, Cillo J. 1996. Catalytic mechanism and DNA substrate recognition of Escherichia coli MutY protein. J Biol Chem 271: 24138–24143. 10.1074/jbc.271.39.24138 CASPubMedWeb of Science®Google Scholar Lu R, Nash HM, Verdine GL. 1997. A mammalian DNA repair enzyme that excises oxidatively damaged guanines maps to a locus frequently lost in lung cancer. Curr Biol 7: 397–407. 10.1016/S0960-9822(06)00187-4 CASPubMedWeb of Science®Google Scholar Maki H, Sekiguchi M. 1992. MutT protein specifically hydrolyses a potent mutagenic substrate for DNA synthesis. Nature 355: 273–275. 10.1038/355273a0 CASPubMedWeb of Science®Google Scholar Manuel RC, Lloyd RS. 1997. Cloning, overexpression, and biochemical characterization of the catalytic domain of MutY. Biochemistry 36: 11140–11152. 10.1021/bi9709708 CASPubMedWeb of Science®Google Scholar McGoldrick JP, Yeh YC, Solomon M, Essigmann JM, Lu AL. 1995. Characterization of a mammalian homolog of the Escherichia coli MutY mismatch repair protein. Mol Cell Biol 15: 989–996. 10.1128/MCB.15.2.989 CASPubMedWeb of Science®Google Scholar Michaels ML, Miller JH. 1992. The GO system protects organisms from the mutagenic effect of the spontaneous lesion 8-hydroxyguanine (7,8-dihydro-8-oxoguanine). J Bacteriol 174: 6321–6325. 10.1128/jb.174.20.6321-6325.1992 CASPubMedWeb of Science®Google Scholar Michaels ML, Tchou J, Grollman AP, Miller JH. 1992. A repair system for 8-oxo-7,8-dihydrodeoxyguanine. Biochemistry 31: 10964–10968. 10.1021/bi00160a004 CASPubMedWeb of Science®Google Scholar Moriya M, Grollman AP. 1993. Mutations in the mutY gene of Escherichia coli enhance the frequency of targeted G:C→T:A transversions induced by a single 8-oxoguanine residue in single-stranded DNA. Mol Gen Genet 239: 72–76. CASPubMedWeb of Science®Google Scholar Moriya M, Ou C, Bodepudi V, Johnson F, Takeshita M, Grollman AP. 1991. Site-specific mutagenesis using a gapped duplex vector: a study of translesion synthesis past 8-oxodeoxyguanosine in E. coli. Mutat Res 254: 281–288. 10.1016/0921-8777(91)90067-Y CASPubMedWeb of Science®Google Scholar Nghiem Y, Cabrera M, Cupples CG, Miller JH. 1988. The mutY gene: a mutator locus in Escherichia coli that generates G.C—T.A transversions. Proc Natl Acad Sci USA 85: 2709–2713. 10.1073/pnas.85.8.2709 CASPubMedWeb of Science®Google Scholar Porello SL, Leyes AE, David SS. 1998. Single-turnover and pre-steady-state kinetics of the reaction of the adenine glycosylase MutY with mismatch-containing DNA substrates. Biochemistry 37: 14756–14764. 10.1021/bi981594+ CASPubMedWeb of Science®Google Scholar Prickett KS, Amberg DC, Hopp TP. 1989. A calcium-dependent antibody for identification and purification of recombinant proteins. Biotechniques 7: 580–589. CASPubMedWeb of Science®Google Scholar Radicella JP, Clark EA, Fox MS. 1988. Some mismatch repair activities in Escherichia coli. Proc Natl Acad Sci USA 85: 9674–9678. 10.1073/pnas.85.24.9674 CASPubMedWeb of Science®Google Scholar Radicella JP, Dherin C, Desmaze C, Fox MS, Boiteux S. 1997. Cloning and characterization of hOGG1, a human homolog of the OGG1 gene of Saccharomyces cerevisiae. Proc Natl Acad Sci USA 94: 8010–8015. 10.1073/pnas.94.15.8010 CASPubMedWeb of Science®Google Scholar Roldan-Arjona T, Wei YF, Carter KC, Klungland A, Anselmino C, Wang RP, Augustus M, Lindahl T. 1997. Molecular cloning and functional expression of a human cDNA encoding the antimutator enzyme 8-hydroxyguanine-DNA glycosylase. Proc Natl Acad Sci USA 94: 8016–8020. 10.1073/pnas.94.15.8016 CASPubMedWeb of Science®Google Scholar Sambrook J, Fritsch EF, Maniatis T. 1989. Molecular cloning: a laboratory manual, 2nd edition. Cold Spring Harbor, New York: Cold Spring Harbor Laboratory Press. 10.1111/j.1095-8312.1996.tb01434.x Google Scholar Shibutani S, Takeshita M, Grollman AP. 1991. Insertion of specific bases during DNA synthesis past the oxidation-damaged base 8-oxodG. Nature 349: 431–434. 10.1038/349431a0 CASPubMedWeb of Science®Google Scholar Slupska MM, Baikalov C, Luther WM, Chiang JH, Wei YF, Miller JH. 1996. Cloning and sequencing a human homolog (hMYH) of the Escherichia coli mutY gene whose function is required for the repair of oxidative DNA damage. J Bacteriol 178: 3885–3892. 10.1128/jb.178.13.3885-3892.1996 CASPubMedWeb of Science®Google Scholar Tajiri T, Maki H, Sekiguchi M. 1995. Functional cooperation of MutT, MutM and MutY proteins in preventing mutations caused by spontaneous oxidation of guanine nucleotide in Escherichia coli. Mutat Res 336: 257–267. 10.1016/0921-8777(94)00062-B CASPubMedWeb of Science®Google Scholar Takao M, Aburatani H, Kobayashi K, Yasui A. 1998. Mitochondrial targeting of human DNA glycosylases for repair of oxidative DNA damage. Nucleic Acids Res 26: 2917–2922. 10.1093/nar/26.12.2917 CASPubMedWeb of Science®Google Scholar Takao M, Zhang QM, Yonei S, Yasui A. 1999. Differential subcellular localization of human MutY homolog (hMYH) and the functional activity of adenine:8-oxoguanine DNA glycosylase. Nucleic Acids Res 27: 3638–3644. 10.1093/nar/27.18.3638 CASPubMedWeb of Science®Google Scholar Tchou J, Grollman AP. 1993. Repair of DNA containing the oxidatively-damaged base, 8-oxoguanine. Mutat Res 299: 277–287. 10.1016/0165-1218(93)90104-L CASPubMedWeb of Science®Google Scholar Tchou J, Kasai H, Shibutani S, Chung MH, Laval J, Grollman AP, Nishimura S. 1991. 8-oxoguanine (8-hydroxyguanine) DNA glycosylase and its substrate specificity. Proc Natl Acad Sci USA 88: 4690–4694. 10.1073/pnas.88.11.4690 CASPubMedWeb of Science®Google Scholar Thayer MM, Ahern H, Xing D, Cunningham RP, Tainer JA. 1995. Novel DNA binding motifs in the DNA repair enzyme endonuclease III crystal structure. EMBO J 14: 4108–4120. 10.1002/j.1460-2075.1995.tb00083.x CASPubMedWeb of Science®Google Scholar Tsai-Wu JJ, Liu HF, Lu AL. 1992. Escherichia coli MutY protein has both N-glycosylase and apurinic/apyrimidinic endonuclease activities on A.C and A.G mispairs. Proc Natl Acad Sci USA 89: 8779–8783. 10.1073/pnas.89.18.8779 CASPubMedWeb of Science®Google Scholar Tsai-Wu JJ, Su MJ, Fang W, Wu CH. 1999. Preparation of heteroduplex DNA containing a mismatch base pair with magnetic beads. Anal Biochem 275: 127–129. 10.1006/abio.1999.4296 CASPubMedWeb of Science®Google Scholar Wood ML, Dizdaroglu M, Gajewski E, Essigmann JM. 1990. Mechanistic studies of ionizing radiation and oxidative mutagenesis: genetic effects of a single 8-hydroxyguanine (7-hydro-8-oxoguanine) residue inserted at a unique site in a viral genome. Biochemistry 29: 7024–7032. 10.1021/bi00482a011 CASPubMedWeb of Science®Google Scholar Wu CH, Tsai-Wu JJ, Huang YT, Lin CY, Lioua GG, Lee FJ. 1998. Identification and subcellular localization of a novel Cu,Zn superoxide dismutase of Mycobacterium tuberculosis. FEBS Letters 439: 192–196. 10.1016/S0014-5793(98)01373-8 CASPubMedWeb of Science®Google Scholar Yamagata Y, Kato M, Odawara K, Tokuno Y, Nakashima Y, Matsushima N, Yasumura K, Tomita K, Ihara K, Fujii Y, Nakabeppu Y, Sekiguchi M, Fujii S. 1996. Three-dimensional structure of a DNA repair enzyme, 3-methyladenine DNA glycosylase II, from Escherichia coli. Cell 86: 311–319. 10.1016/S0092-8674(00)80102-6 CASPubMedWeb of Science®Google Scholar Yeh YC, Chang DY, Masin J, Lu AL. 1991. Two nicking enzyme systems specific for mismatch-containing DNA in nuclear extracts from human cells. J Biol Chem 266: 6480–6484. 10.1016/S0021-9258(18)38143-2 CASPubMedWeb of Science®Google Scholar Citing Literature Volume77, Issue415 June 2000Pages 666-677 ReferencesRelatedInformation