Major and Essential Role for the DNA Methylation Mark in Mouse Embryogenesis and Stable Association of DNMT1 with Newly Replicated Regions

Abstract

ABSTRACT DNA methyltransferase 1 (DNMT1) plays an important role in the inheritance of genomic DNA methylation, which is coupled to the DNA replication process. Early embryonic lethality in DNMT1-null mutant ( Dnmt1 c ) mice indicates that DNA methylation is essential for mammalian development. DNMT1, however, interacts with a number of transcriptional regulators and has a transcriptional repressor activity independent of its catalytic activity. To examine the roles of the catalytic activity of DNMT1 in vivo, we generated a Dnmt1 ps allele that expresses a point-mutated protein that lacks catalytic activity (DNMT1-C1229S). Dnmt1 ps mutant mice showed developmental arrest shortly after gastrulation, near-complete loss of DNA methylation, and an altered distribution of repressive chromatin markers in the nuclei; these phenotypes are quite similar to those of the Dnmt1 c mutant. The mutant DNMT1 protein failed to associate with replication foci in Dnmt1 ps cells. Reconstitution experiments and replication labeling in Dnmt1 −/− Dnmt3a −/− Dnmt3b −/− (i.e., unmethylated) embryonic stem cells revealed that preexisting DNA methylation is a major determinant for the cell cycle-dependent localization of DNMT1. The C-terminal catalytic domain of DNMT1 inhibited its stable association with unmethylated chromatin. Our results reveal essential roles for the DNA methylation mark in mammalian development and in DNMT1 localization.