Repair of O6-methylguanine in rat pancreatic beta-cells after exposure to N-methyl-N-nitrosourea.
Because of the possible involvement of O6-methyldeoxyguanosine as a cytotoxic and carcinogenic lesion in pancreatic beta-cells, studies were undertaken to assess the ability of rat beta-cells to repair this DNA lesion. Primary cultures of neonatal rat beta-cells were shown to contain very low levels of O6-methylguanine-DNA-methyltransferase activity, the predominant mechanism for repairing O6-methyldeoxyguanosine in mammalian cells. However, using a 32P-endlabeling assay to measure O6-methyldeoxyguanosine in cells after exposure to N-methyl-N-nitrosourea, it was determined that rat beta-cells repaired O6-methyldeoxyguanosine to a substantial extent over a 24-h period. To elucidate the mechanism of O6-methyldeoxyguanosine repair in the virtual absence of constitutive O6-methylguanine-DNA-methyltransferase expression, studies were performed to determine if O6-methylguanine-DNA-methyltransferase expression was enhanced in N-methyl-N-nitrosourea-treated beta-cells. No increase in O6-methylguanine-DNA-methyltransferase activity was detected 24 or 48 h after exposure. However, Northern blot analysis showed a two- to threefold elevation in O6-methylguanine-DNA-methyltransferase messenger RNA levels in beta-cells 12 and 24 h after N-methyl-N-nitrosourea treatment. This finding is the first demonstration of a change in O6-methylguanine-DNA-methyltransferase messenger RNA levels in a cell type with low constitutive activity.
Nelson
J W
JW
Department of Structural and Cellular Biology, University of South Alabama, Mobile 36688.
Ledoux
S P
SP
Wilson
G L
GL
D000818:Animals; D015152:Blotting, Northern; D016466:CHO Cells; D002478:Cells, Cultured; D006224:Cricetinae; D006147:Guanine / Q000031:analogs & derivatives* / Q000378:metabolism; D007515:Islets of Langerhans / Q000187:drug effects / Q000378:metabolism*; D008770:Methylnitrosourea / Q000494:pharmacology*; D008780:Methyltransferases / Q000235:genetics / Q000378:metabolism*; D019853:O(6)-Methylguanine-DNA Methyltransferase; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D014407:Tumor Cells, Cultured
D012333:RNA, Messenger; D006147:Guanine; D008770:Methylnitrosourea; C008449:O-(6)-methylguanine; D008780:Methyltransferases; D019853:O(6)-Methylguanine-DNA Methyltransferase