Loss of mismatch repair activity in simian virus 40 large T antigen-immortalized BPH-1 human prostatic epithelial cell line.
Simian virus 40 large T antigen (SVLTAg) has been used to immortalize cells; however, the mechanism leading to immortalization is still unclear. We hypothesize that DNA mismatch repair (MMR) activity is important during SVLTAg-induced immortalization. To test this hypothesis, we used the SVLTAg-immortalized cell line BPH-1 derived from human benign prostate epithelial cells to analyze MMR activity and the expression of MMR genes (hMLH1, hPMS1, hPMS2, hMSH2, hMSH3, and hMSH6). The results demonstrated that BPH-1 cells were deficient in repairing G:T, A:C, and G:G mispairs in bacteriophage M13mp2. Reverse-transcription polymerase chain reaction experiments indicated MMR genes (hMSH3, hMSH6, and hPMS1) were expressed at a low level in BPH-1 cells. In contrast, all six MMR genes were expressed in human benign prostate hyperplasia tissues. Downregulation of hMSH3, hMSH6, and hPMS1 genes is not a result of the hypermethylation mechanism because demethylation with 5-aza-2'-deoxycytidine did not restore expression of these genes. Although the hMLH1 gene is expressed in BPH-1 cells, western blotting and exon analyses demonstrated that hMLH1 was mutated and/or deleted in BPH-1 cells.
Yeh
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CC
Department of Urology, Veterans Affairs Medical Center and University of California, San Francisco, California, USA.
Lee
C
C
Huang
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MC
Dahiya
R
R
D048868:Adaptor Proteins, Signal Transducing; D000251:Adenosine Triphosphatases; D000952:Antigens, Polyomavirus Transforming / Q000378:metabolism*; D001374:Azacitidine / Q000031:analogs & derivatives / Q000494:pharmacology; D020137:Base Pair Mismatch; D015153:Blotting, Western; D002352:Carrier Proteins / Q000096:biosynthesis; D002460:Cell Line; D019175:DNA Methylation; D004260:DNA Repair; D045643:DNA Repair Enzymes; D004268:DNA-Binding Proteins / Q000096:biosynthesis; D000077209:Decitabine; D015536:Down-Regulation; D004847:Epithelial Cells / Q000166:cytology*; D005091:Exons; D005656:Fungal Proteins / Q000096:biosynthesis; D017353:Gene Deletion; D006801:Humans; D008297:Male; D000070976:Mismatch Repair Endonuclease PMS2; D008954:Models, Biological; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D000070957:MutL Protein Homolog 1; D000070956:MutL Proteins; D051718:MutS Homolog 2 Protein; D000074081:MutS Homolog 3 Protein; D009154:Mutation; D009363:Neoplasm Proteins / Q000096:biosynthesis; D009687:Nuclear Proteins; D009692:Nucleic Acid Heteroduplexes; D011467:Prostate / Q000166:cytology*; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D029701:Saccharomyces cerevisiae Proteins
D048868:Adaptor Proteins, Signal Transducing; D000251:Adenosine Triphosphatases; D000952:Antigens, Polyomavirus Transforming; D001374:Azacitidine; D002352:Carrier Proteins; D045643:DNA Repair Enzymes; D004268:DNA-Binding Proteins; D000077209:Decitabine; D005656:Fungal Proteins; D000070976:Mismatch Repair Endonuclease PMS2; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D000070957:MutL Protein Homolog 1; D000070956:MutL Proteins; D051718:MutS Homolog 2 Protein; D000074081:MutS Homolog 3 Protein; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D009692:Nucleic Acid Heteroduplexes; D011518:Proto-Oncogene Proteins; D029701:Saccharomyces cerevisiae Proteins; C501486:MLH1 protein, human; C078444:MSH3 protein, human; C098790:MSH6 protein, S cerevisiae; C088996:PMS1 protein, human; C094934:PMS2 protein, human; C497172:MSH2 protein, human; C421109:multidrug resistance-associated protein 1