Influence of hydrocortisone on the binding of nitrosoureas to nuclear chromatin subfractions.
The effects of steroid-induced modifications of chromatin structure on the extent and sites of chloroethylnitrosourea binding to chromatin were studied using log-phase HeLa cells. The cells were exposed to 0.1 to 2.0 microM hydrocortisone for 22 hr; this resulted in depressed DNA synthesis while transcriptional activity was stimulated. Hydrocortisone had no effect upon cellular or nuclear uptake of the two nitrosoureas under study, 0.6 mM chlorozotocin or 1-(2-chloroethyl-3-cyclohexyl-1-nitrosourea). Both drugs were found to alkylate transcriptional chromatin preferentially, as demonstrated by DNase II and DNase I digestion. This alkylation was stimulated 2-fold by the same micromolar concentrations of hydrocortisone, 0.1 to 2.0 microM, which stimulated transcription. The extent of nuclear RNA alkylation, determined using RNase T2 as a probe, was found to contribute less than 20% of total chromatin alkylation and was unaffected by steroid pretreatment. Instead, the increased alkylation within these chromatin subfractions was attributed to a steroid-induced alteration of chromatin structure. Electron microscopic examination of HeLa nuclear morphology revealed a hydrocortisone-induced disaggregation of nuclear membrane-associated heterochromatin resulting in a more heterogeneous, less condensed distribution of chromatin. Such data are consistent with a relaxation of the supercoiled chromatin structure, resulting in increased transcription and increased accessibility of potential target sites for nitrosourea alkylation.
D000478:Alkylation; D001665:Binding Sites; D002467:Cell Nucleus / Q000378:metabolism / Q000648:ultrastructure; D002843:Chromatin / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D004247:DNA / Q000096:biosynthesis; D003851:Deoxyribonucleases / Q000378:metabolism; D006367:HeLa Cells; D006651:Histocytochemistry; D006801:Humans; D006854:Hydrocortisone / Q000494:pharmacology*; D008274:Magnesium / Q000494:pharmacology; D009607:Nitrosourea Compounds / Q000378:metabolism*; D012260:Ribonucleases / Q000378:metabolism
D002843:Chromatin; D009607:Nitrosourea Compounds; D004247:DNA; D003851:Deoxyribonucleases; D012260:Ribonucleases; D008274:Magnesium; D006854:Hydrocortisone