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

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.

Methylation of the T-DNA inAgrobacterium tumefaciensand In several crown gall tumors

“…Many genes, including those coding for ovalbumin (7), globin (8,9,10), metallothionein (11), and certain viral DNAs (12) are undermethylated in cells in which they are expressed relative to cells in which they are inactive. Furthermore, inhibitors of DNA methylation, such as 5-azacytidine, can activate endogenous viruses.…”

“…E. coZi K12 strains harboring recombinant plasmids were grown at 37°C in M9 + 1.0% casamino acids with 100 jig/ml ampicillin (27). Recombinant plasmids containing Bam Hi fragments 8,30, and 30' (previously denoted pNW31C-8,29-1 [16,19,21]) and Bam Hi fragment 19 were derived from pTiA6 (16). Recombinant plasmids containing Bam Hi fragment 2 and subfragments of Bam Hi fragment 2 were derived from pTiB6806 and constructed using standard procedures (28).…”

Altered globin gene transcription pattern and the presence of a 7-8 kb alpha A globin gene transcript in avian erythroblastosis virus-transformed cells.

“…The possibility therefore arises that transcription of the aA gene is norinal in the AEV cell, and that the viral transformation has perturbed only processing. In this context we may recall that the known a and ( globin gene domains in AEV-transformed cell DNA were found to be normal, except for their methylation pattern (Marcaud et al, 1981).…”

Fetal Globin Gene Repressors as Drug Targets for Molecular Therapies To Treat the β-Globinopathies

“…Perhaps, intriguingly, DNMTs, HDACs, and the LSD1 enzymes are all implicated in fetal globin gene silencing by both TR2/TR4 and BCL11A (39, 86). DNA methylation within the locus has for many years been one of the hallmarks of globin gene inactivity through the strong correlation of methylation with repression (104)(105)(106)(107)(108). Histone acetylation of globin gene promoters also correlates with the expression levels of these genes (109), and HDAC inhibitors such as butyrate have been shown to induce HbF (110).…”