CHROMATIN STRUCTURE AND METHYLATION OF RAT RIBOSOMAL-RNA GENES STUDIED BY FORMALDEHYDE FIXATION AND PSORALEN CROSS-LINKING

By using formaldehyde cross-linking of histones to DNA and gel retarda tion assays we show that formaldehyde fixation, similar to previously established psoralen photocross-linking, discriminates between nucleos ome- packed (inactive) and nucleosome-free (active) fractions of ribos omal RNA genes, By both cross-linking techniques we were able to purif y fragments from agarose gels, corresponding to coding, enhancer and p romoter sequences of rRNA genes, which were further investigated with respect to DNA methylation. This approach allows us to analyse indepen dently and in detail methylation patterns of active and inactive rRNA gene copies by the combination of HpaII and MspI restriction enzymes. We found CpG methylation mainly present in enhancer and promoter regio ns of inactive rRNA gene copies. The methylation of one single HpaII s ite, located in the promoter region, showed particularly strong correl ation with the transcriptional activity.