Progressive aberrant methylation of the RASSF1A gene in simian virus 40 infected human mesothelial cells.
Mesotheliomas are tumors arising from mesothelial cells and are associated with asbestos exposure and approximately 50% contain simian virus 40 (SV40) DNA sequences. SV40 infection of human mesothelial cells (HM) causes early cellular immortalization and late transformation. Aberrant methylation is a major mech-anism for loss of function of tumor suppressor genes (TSGs). We recently reported that of seven genes frequently methylated in epithelial tumors, only RASSF1A gene was frequently methylated in mesotheliomas, and its methylation was correlated with loss of RASSF1A expression and the presence of SV40. We studied whether SV40 infection of normal HM induces aberrant methylation of the genes previously studied in mesotheliomas. Of six infected foci examined at early passages (passages 8-30) there was no methylation of the seven genes examined. Of two foci examined at late passages (passages 51-86) after the appearance of morphological changes suggestive of transformation, methylation and loss of expression of RASSF1A was detected. Sequencing of the CpG dense region around the transcription start site and semi-quantitative real-time methylation specific PCR (MSP) assay for RASSF1A methylation demonstrated progressive methylation during late passages. Exposure to the demethylating agent 5-aza-2'-deoxycytidine restored RASSF1A expression, while exposure to the histone deacetylation inhibitor trichostatin A had no effect. These data, together with our previous findings, support a causal relationship between SV40 infection, progressive RASSF1A methylation and its silencing, and the pathogenesis of mesothelioma.
Toyooka
Shinichi
S
Hamon Center for Therapeutic Oncology Research, University of Texas Southwestern Medical Center, Dallas, Texas, TX 75390, USA.
Carbone
Michele
M
Toyooka
Kiyomi O
KO
Bocchetta
Maurizio
M
Shivapurkar
Narayan
N
Minna
John D
JD
Gazdar
Adi F
AF
D000483:Alleles; D001374:Azacitidine / Q000031:analogs & derivatives* / Q000494:pharmacology; D002461:Cell Line, Transformed; D002472:Cell Transformation, Viral / Q000235:genetics*; D003043:Cocarcinogenesis; D018899:CpG Islands; D019175:DNA Methylation / Q000187:drug effects; D004273:DNA, Neoplasm / Q000737:chemistry / Q000235:genetics; D000077209:Decitabine; D005786:Gene Expression Regulation / Q000187:drug effects; D020868:Gene Silencing; D016147:Genes, Tumor Suppressor; D006801:Humans; D006877:Hydroxamic Acids / Q000494:pharmacology; D008654:Mesothelioma / Q000235:genetics* / Q000821:virology; D009363:Neoplasm Proteins / Q000235:genetics* / Q000502:physiology; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D013539:Simian virus 40 / Q000472:pathogenicity / Q000502:physiology*; D025521:Tumor Suppressor Proteins
D004273:DNA, Neoplasm; D006877:Hydroxamic Acids; D009363:Neoplasm Proteins; C411762:RASSF1 protein, human; D025521:Tumor Suppressor Proteins; C012589:trichostatin A; D000077209:Decitabine; D001374:Azacitidine