Identification of a region of the DNMT1 methyltransferase that regulates the maintenance of genomic imprints

Abstract

Reprogramming of DNA methylation patterns during mammalian preimplantation development involves the concurrent maintenance of methylation on differentially methylated domains (DMDs) of imprinted genes and a marked reduction of global (non-DMD) genomic methylation. In the developing mammalian embryo, one allele of a DMD is unmethylated, and the opposite parental allele is methylated, having inherited this methylation from the parental gamete. The maintenance of DMDs is important for monoallelic imprinted gene expression and normal development of the embryo. Because the DNMT1 cytosine methyltransferase governs maintenance methylation in mammals, rearrangements of non-DMD, but not DMD methylation in preimplantation embryos suggest that the preimplantation DNMT1-dependent maintenance mechanism specifically targets DMD sequences. We explored this possibility using an engineered mouse ES cell line to screen for mutant DNMT1 proteins that protect against the loss of DMD and/or global (non-DMD) methylation in the absence of the wild-type endogenous DNMT1 methyltransferase. We identified DNMT1 mutants that were defective in maintenance of either DMD and/or non-DMD methylation. Among these, one mutant maintained non-DMD methylation but not imprinted DMD methylation and another mutant maintained just DMD methylation. The mutated amino acids of these mutants reside in a mammal-specific, disordered region near the amino terminus of DNMT1. These findings suggest that DNMT1 participates in epigenetic reprogramming through its ability to distinguish different categories of methylated sequences.epigenetic ͉ imprinting ͉ methylase ͉ methylation ͉ reprogramming G enomic imprinting is a mammalian epigenetic process that distinguishes maternal and paternal alleles to ensure parent-specific (monoallelic) expression of Ϸ80 imprinted genes (1). The molecular basis of this process is de novo methylation during gametogenesis and maintenance methylation during embryogenesis; this sequence of activities leads to the generation of imprinted DMDs (2). Methylation is placed on DMD sequences in the maternal and paternal germ lines by the DNMT3a cytosine methyltransferase (3, 4). Following fertilization DMD methylation is maintained (inherited) during preimplantation development by the combined action of different isoforms of the DNMT1 cytosine methyltransferase (5-8). The M r 175,000 DNMT1o form is synthesized in the oocyte and maintains methylation during preimplantation development (5, 9), whereas the M r 190,000 DNMT1s form is synthesized both in the oocyte and in the early embryo and this form also functions in preimplantation embryos (6). Along with this maintenance of DMD methylation is a significant reduction in the level of global (non-DMD) methylation (10, 11). The concurrent inheritance of DMD methylation and reduction of global methylation rearranges the genome's methylation patterns just before the formation of pluripotent embryo stem cells (12).Maintenance of DMD methylation in the presence of a reduction in the average level of g...

Zinc Finger Protein ZFP57 Requires Its Co-factor to Recruit DNA Methyltransferases and Maintains DNA Methylation Imprint in Embryonic Stem Cells via Its Transcriptional Repression Domain

“…These results suggest that ZFP57 is continuously required for maintaining DNA methylation imprint in ES cells, and maintenance of genomic imprinting may be an active process. Similar findings have been discovered previously for the mechanisms of the maintenance DNA methyltransferase DNMT1 in DNA methylation imprint (45,59,60).…”

“…ES cells have been increasingly employed as a model system for studying genomic imprinting (9,25,(42)(43)(44)(45)(46). Indeed, we found that ZFP57 maintains DNA methylation imprint at a large subset of imprinted regions in ES cells.…”

The Dnmt1 Intrinsically Disordered Domain Regulates Genomic Methylation During Development

“…This suggests that DNMT1 activity is regulated and that noninactivating mutations in Dnmt1 that generate informative mutant methylomes can be found. Based on this notion, we studied the effects on genomic methylation of expressing mutant DNMT1 proteins in ES cells using a Dnmt1 tet/tet line in which the endogenous DNMT1 enzyme is extinguished by the administration of doxycycline (Borowczyk et al 2009). We found that methylome changes were induced by expressing DNMT1 proteins with mutations in its large 300-aa intrinsically disordered domain (IDD)-some IDD mutants produced methylomes specifically lacking gDMD methylation, whereas other mutants produced methylomes retaining normal gDMD methylation but with much reduced global methylation.…”

Genomic imprinting in mammals: its life cycle, molecular mechanisms and reprogramming

“…More recently, it was reported that the zygotically expressed, somatic form of DNMT1, called DNMT1s, maintains the methylation imprints at the other cell cycles of pre-implantation development [49,50]. At present we do not know how the DNMT1 isoforms specifically find the DMRs among many other DNA regions, but a recent study suggested that a mammalian-specific region near the amino terminus of DNMT1 is probably involved in the discrimination [51].…”