Modification of the methylation pattern in the vicinity of the chicken globin genes in avian erythroblastosis virus transformed cells

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Abstract
Having previously found a reduced transcription of globin genes and an abortive processing of the already transcribed globin pre-mRNA in Avian Erythroblastosis Virus (AEV) transformed cells (1), we compared the genomic DNA of these cells with that of normal chicken erythroblasts, using 32-P-labelled cDNA probes specific for the beta, alpha A and alpha D globin sequences. Restriction endonuclease digestion, electrophoresis of digests in agarose gels, Southern blotting and hybridization were carried out. Our results show that the overall genome organization is not disturbed in the immediate neighbourhood of the adult globin genes; the observed restriction fragments are identical for both DNAs after EcoRI, HindIII, BamHI and XbaI digestion, using the beta, alpha A and alpha D globin cDNA probes. However, we observe specific modifications at some methylation sites in the beta, beta-like and alpha D regions: after HpaII or MspI digestion in the alpha D region and after HhaI digestion in the beta and beta-like region, heavier bands appear in the transformed cell DNA in addition to the ones observed in normal DNA. This implies that, at some specific sites, the transformed cell DNA is more methylated than the normal erythroblast DNA. The possible significance of this observation is discussed.