Phosphatidylinositol 3-kinase (PI3K) signaling via glycogen synthase kinase-3 (Gsk-3) regulates DNA methylation of imprinted loci.

Glycogen synthase kinase-3 (Gsk-3) isoforms, Gsk-3α and Gsk-3ß, are constitutively active, largely inhibitory kinases involved in signal transduction. Underscoring their biological significance, altered Gsk-3 activity has been implicated in diabetes, Alzheimer disease, schizophrenia, and bipolar disorder. Here, we demonstrate that deletion of both Gsk-3α and Gsk-3ß in mouse embryonic stem cells results in reduced expression of the de novo DNA methyltransferase Dnmt3a2, causing misexpression of the imprinted genes Igf2, H19, and Igf2r and hypomethylation of their corresponding imprinted control regions. Treatment of wild-type embryonic stem cells and neural stem cells with the Gsk-3 inhibitor, lithium, phenocopies the DNA hypomethylation at these imprinted loci. We show that inhibition of Gsk-3 by phosphatidylinositol 3-kinase (PI3K)-mediated activation of Akt also results in reduced DNA methylation at these imprinted loci. Finally, we find that N-Myc is a potent Gsk-3-dependent regulator of Dnmt3a2 expression. In summary, we have identified a signal transduction pathway that is capable of altering the DNA methylation of imprinted loci.

UV sensitive mutations in histone H3 in Saccharomyces cerevisiae that alter specific K79 methylation states genetically act through distinct DNA repair pathways.

Chromatin serves as a regulator of various nuclear processes, with post-translational modifications of histone proteins serving as modulators to influence chromatin function. We have previously shown that histone H3 K79 methylation is important for repair of UV-induced DNA damage in Saccharomyces cerevisiae, acting through multiple repair pathways. To evaluate the potential role of distinct K79 methylation states in DNA repair, we identified four mutations in histone H3 that confer sensitivity to UV, each of which also has a distinct effect on specific K79 methylation states. Epistasis analyses indicate that each mutation exerts its phenotypic effects through distinct subsets of the various DNA damage response pathways, suggesting the existence of discrete roles for histone H3 in DNA damage checkpoint and repair pathways. Furthermore, we find that the distribution of K79 methylation states is altered by mutation of the acetylatable N terminal lysines in histone H4. The combined results suggest that K79 methylation states may be modulated in response to UV damage via a trans-histone regulatory pathway, and that distinct methylation states may provide a means of coordinating specific DNA repair and damage checkpoint pathways.

Methylation of histone H3 lysine-79 by Dot1p plays multiple roles in the response to UV damage in Saccharomyces cerevisiae.

Various proteins have been found to play roles in both the repair of UV damaged DNA and heterochromatin-mediated silencing in the yeast Saccharomyces cerevisiae. In particular, factors that are involved in the methylation of lysine-79 of histone H3 by Dot1p have been implicated in both processes, suggesting a bipartite function for this modification. We find that a dot1 null mutation and a histone H3 point mutation at lysine-79 cause increased sensitivity to UV radiation, suggesting that lysine-79 methylation is important for efficient repair of UV damage. Epistasis analysis between dot1 and various UV repair genes indicates that lysine-79 methylation plays overlapping roles within the nucleotide excision, post-replication and recombination repair pathways, as well as RAD9-mediated checkpoint function. In contrast, epistasis analysis with the H3 lysine-79 point mutation indicates that the lysine-to-glutamic acid substitution exerts specific effects within the nucleotide excision repair and post-replication repair pathways, suggesting that this allele only disrupts a subset of the functions of lysine-79 methylation. The overall results indicate the existence of distinct and separable roles of histone H3 lysine-79 methylation in the response to UV damage, potentially serving to coordinate the various repair processes.