Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: DrugBank:EXPT01586 (G418)
 2,237 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Chinese hamster ovary cells with no detectable (less than 200 molecules/cell) O6-methylguanine-DNA methyltransferase (EC 2.1.1.63) were transfected with human cell DNA and pSV2neo plasmid by electroporation. Two stable transformant clones, GC-1 and GC-2, containing 4 X 10(4) and 4-6 X 10(3) methyltransferase molecules/cell respectively were isolated by successive screening in the presence of G418 and 2-chloroethyl-N-nitrosourea (CNU). Only three or four copies of pSV2neo DNA and no repetitive human DNA sequence were detected in these isolates. Secondary transfection of parent cells with GC-1 DNA yielded several clones containing 2-10 X 10(3) methyltransferase molecules/cell. The rate of removal of O6-methylguanine in GC-1, GC-2 and parent cells in vivo reflected their methyltransferase levels, while the N-methylpurines were removed at similar rates in all three cell lines. The differential sensitivity of these cells to several alkylating agents, namely CNU, N-methyl-N-nitrosourea, N-methyl-N'-nitro-N-nitrosoguanidine and methyl-methane sulfonate (MMS), known to yield different proportions of O6-alkylguanine among the alkyl adducts in DNA, varied widely. The largest and smallest differences in toxic response were observed with CNU and MMS respectively. These cell lines showed no difference in sensitivity to the DNA cross-linking agent psoralen. These data strongly suggest that alkylating agents produce two classes of lethal lesions, one of which is O6-alkylguanine. Induction of mutations at the hypoxanthine-phosphoribosyltransferase locus in these cells lines suggests that, regardless of its relative yield, O6-methylguanine is the major mutagenic lesion for all alkylating agents.
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PMID:The role of O6-alkylguanine in cell killing and mutagenesis in Chinese hamster ovary cells. 198 86

AtT-20/IDG8 cells contain the stably transfected, selectable gene, neomycin phosphotransferase, under negative glucocorticoid regulation. Thus, when cultured in the simultaneous presence of the neomycin analogue, G418, and dexamethasone, AtT-20/IDG8 cells fail to grow. Our hypothesis was that mutated AtT-20/IDG8 cells capable of growth in such medium would have a defect in the glucocorticoid-mediated regulation of the neor gene. AtT-20/IDG8 cells were chemically mutagenized using ethyl-methane sulfonate and cloned in the presence of G418 and dexamethasone. Fourteen clones were obtained and loss of glucocorticoid control of neor expression was confirmed in them all. The naturally occurring gene, pro-opiomelanocortin, which is down-regulated by glucocorticoids in parent AtT-20/IDG8 cells, was down-regulated by dexamethasone in ten of the mutant lines, indicating that in those cells the receptor was functional in spite of aberrant regulation of neor. In the other four lines, pro-opiomelanocortin regulation was lost, also suggesting that a general transcription factor, such as the receptor, had been altered. These results indicate that multiple factors are involved in glucocorticoid-mediated gene regulation and that new, informative mutations can be produced after insertion of a regulated, selectable gene into a previously non-selectable cell line.
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PMID:Selection of glucocorticoid-resistant mutations from an AtT-20 cell line containing a glucocorticoid-regulated selectable transgene. 772 33