Effects of N-methyl-N-nitrosourea on transcriptionally active and inactive nucleosomes: macromolecular damage and DNA repair.

Abstract

We previously reported a separation, on an organomercurial column, of transcriptionally inactive nucleosomes (class 1) from those containing active gene sequences (classes 2 and 3). In this paper, we analyzed nucleosomal damage caused by exposure of HeLa S3 cells in suspension culture to the directly alkylating carcinogen N-methyl-N-nitrosourea (MNU). The extent and site of methylation induced by the compound in nucleosomal DNA and RNA were determined by cell incubation in the presence of [3H]MNU. The highest amount of damage was detected in DNA of class 3 nucleosomes, while RNA alkylation was comparable in all nucleosomal classes. Cellular capacity for repair of MNU-induced DNA strand breaks (estimated after a short pulse with [3H]thymidine) was found to be higher in active nucleosomal fractions (classes 2 and 3) than in the inactive fraction (class 1). Our data support the postulate that chromatin primary structure plays a role in modulating carcinogen damage to chromosomal macromolecules and in DNA strand breakage and repair mechanisms. Some of these initial steps are believed to be critical in the process of carcinogenesis.