Abstract
DNA methyltransferase 1 (DNMT1) is a multidomain protein believed to be involved only in the passive transmission of genomic methylation patterns via maintenance methylation. The mechanisms that regulate DNMT1 activity and targeting are complex and poorly understood. We used embryonic stem (ES) cells to investigate the function of the uncharacterized bromo-adjacent homology (BAH) domains and the glycine-lysine (GK) repeats that join the regulatory and catalytic domains of DNMT1. We removed the BAH domains by means of a CRISPR/Cas9-mediated deletion within the endogenous Dnmt1 locus. The internally deleted protein failed to associate with replication foci during S phase in vivo and lost the ability to mediate maintenance methylation. The data indicate that ablation of the BAH domains causes DNMT1 to be excluded from replication foci even in the presence of the replication focus-targeting sequence (RFTS). The GK repeats resemble the N-terminal tails of histones H2A and H4 and are normally acetylated. Substitution of lysines within the GK repeats with arginines to prevent acetylation did not alter the maintenance activity of DNMT1 but unexpectedly activated de novo methylation of paternal imprinting control regions (ICRs) in mouse ES cells; maternal ICRs remained unmethylated. We propose a model under which DNMT1 deposits paternal imprints in male germ cells in an acetylation-dependent manner. These data reveal that DNMT1 responds to multiple regulatory inputs that control its localization as well as its activity and is not purely a maintenance methyltransferase but can participate in the de novo methylation of a small but essential compartment of the genome.
Highlights
DNA methyltransferase 1 (DNMT1) is a multidomain protein believed to be involved only in the passive transmission of genomic methylation patterns via maintenance methylation
In the crystal structure of DNMT1 bound to unmethylated DNA [5] (Fig. 1B), the bromo-adjacent homology (BAH) domains are remote from the active site of the catalytic domain, but an acidic autoinhibitory linker extends from BAH1 to the CXXC domain, which binds to unmethylated CpG dinucleotides and prevents their entry into the active site of the catalytic domain [5]
The function of this interaction is unknown. Both the BAH1 autoinhibitory linker and the BAH2-target recognition domain (TRD) loops bind to the BAH-binding pockets (Fig. 1C) that in other proteins interact with histone tails in a manner that depends on the modification status of lysine residues
Summary
There has been a focus on the mechanism of interaction of UHRF1 and DNMT1, but little attention has been directed to the function of well-conserved domains located within the N-terminal regulatory region of DNMT1 that are not known to interact with UHRF1. Ognition domain; IAP, intracisternal A-particle; PCNA, proliferating cell nuclear antigen. It was very surprising to find that the GR substitution caused de novo methylation of imprinting control regions (ICRs) in mouse embryonic stem (ES) cells that are normally methylated in male germ cells; maternal ICRs remained unmethylated. These and other data suggest that DNMT1 is likely to be involved in the de novo methylation of paternal ICRs in male germ cells
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