The Dm-Myb Oncoprotein Coordinates Higher-Order Chromatin Structure and Stabilizes Facultative Heterochromatin

Abstract

Mutation of the c-Myb proto-oncogene causes leukemias and lymphomas in birds and mammals. Vertebrates contain three representatives of the Myb gene family consisting of A-, B- and c-Myb, all of which encode DNA-binding factors that bind to promoter regions and are important for the proper expression of target genes. In flies, which have a single Myb gene, Dm-Myb is required for epigenetic maintenance of the ON state of target genes subsequent to their activation, and plays a key role in regulation of mitotic phase genes. Here, we identify additional mitotic gene targets, a nearly three-fold increase compared to known targets, and further refine the role of Dm-Myb as an activator of gene expression that is required to 1) promote the proper recruitment of RNA pol II at promoters, 2) promote efficient H3K4 methylation along target promoter regions, and 3) potentiate the transcriptional response. In contrast to its role in gene activation, Myb is also required for repression of a multitude of genes, although no specific mechanism has yet been proposed for how Myb might be functioning in this context. We now reveal a critical role for Myb in promoting the binding of insulator proteins BEAF-32 and CP190, and show that Myb is required genome-wide for insulator function. Targeting of the Myb complex to insulator sites is primarily through Myb rather than E2F2, and its insulator function is independent of E2F2/Rbf. Notably, Myb is required to stabilize facultative heterochromatin domains by binding to the boundaries that demarcate the domains; in the absence of Myb, the repressing mark H3K27me3 is markedly reduced throughout the genome, leading to encroachment of the activating mark H3K4me3 and gene derepression within the domains. Finally, Myb binding strongly correlates with topologically associating domain (TAD) boundaries, which play a key role in chromatin looping. These results reveal that Myb plays a significantly larger role in insulator and chromatin organization than previously appreciated.