Dynamics of the ZFP57/KAP1 and associated factors network at DNA methylated targets in murine pluripotent ES cells

Zfp57 plays a pivotal role in the maintenance of parent-of-origin-dependent epigenetic state of the Imprinting Control Regions (ICRs) that regulate hundreds of imprinted loci. We report co-occupancy of the sequence-specific endogenous ZFP57 and associated chromatin modifiers-KAP1, Hp1? and H3K9me3 in murine embryonal stem cells (ESCs). It seems to preferentially occur at regions harboring certain extended versions of the ZFP57 core exameric binding site (BS). The majority of ZFP57 bound regions reside at non-ICRs, notably at additional germ-line DNA methylated CpG islands (CGIs) and certain repeats, predominantly at ERVK endogenous retroviruses, implying a wider role for this factor in shaping the pluripotent epigenome. Non-ICR regions and cognate BSs display a higher DNA methylation compared to the genome of pluripotent ES cells either grown in two kinase inhibitor (2i) or serum+LIF, conditions known to affect their interconvertible DNA methylomes. Further, upon global comparison, while ICRs appear stable, non-ICR bound regions in ES cells grown in serum+LIF have a higher state of methylation in naive ES cells grown in 2i than their genome-wide counterparts and present with a broad spectrum of individual CpGs methylation levels. In vitro analyses support preferential binding at specific extended BS and a diffused sensitivity to methylation. We show that in vivo the factor globally selects DNA methylated alleles both at monoallelic marked ICRs and at additional targets. This indicates that the in vivo methylation state of the excedingly large number of potential targets represents a selective factor for ZFP57 stable association along with certain BS versions, density and arrangement, in a context dependent manner, marking and contributing towards epigenetic/epigenomic heterogeneity of pluripotent ES cells.