Mutations In DNA Methyltransferase DNMT3B In ICF Syndrome Affect Its Regulation By DNMT3L

ICF (immunodeficiency, centromeric instability involving the pericentromericregions of chromosomes 1, 9 and 16 and facial anomalies) syndrome is a rareautosomal recessive disease with defects in genomic methylation in the classicalsatellite 2 and 3. The ICF locus was mapped on chromosome 20q11.2 byhomozygosity mapping, where the DNMT3B gene is localized. Mutation analysis ofthe DNMT3B gene revealed the presence of nonsence and missence mutations in itsexons. The majority of ICF mutations are single amino acid substitutions in theconserved catalytic domain of DNMT3B which may impair the enzymatic activitydirectly.The establishment of intact genomic methylation patterns in mammaliandevelopment requires a fine control of the de novo methylation activity of the tworelated methyltransferases Dnmt3a and Dnmt3b by regulatory factors includingDnmt3L. Dnmt3L is a new member of Dnmt3 family. Although lackingmethyltransferase activity, Dnmt3L has been shown to stimulate the emzymaticactivity of Dnmt3a during the establishment of genomic imprints. During ICF fetaldevelopment, the establishment of DNA methylation is defective and patients'genomic DNA displays extraembryonic-like methylation profile in their constitutiveheterochromatin. The co-expression of DNMT3B and DNMT3L mRNA during earlyembryonic development suggests their collaboration in this de novo methylationprocess.In this work, we showed that Dnmt3b interacts with Dnmt3L in endogenous EScells and further mapped the interaction regions to their respective C-terminal region.We found that two Dnmt3b mutant proteins with ICF-related substitutions (A772Pand R846Q) in the catalytic domain displayed a methylation activity similar to thewild-type enzyme both in vitro and in vivo, but their stimulation by Dnmt3L wasseverely compromised due to deficient protein interaction. We also detected theco-expression of Dnmt3b and Dnmt3L proteins in the pre-gastrulation embryos,which indicated that they might interact to establish the DNA methylation patternduring this stage.We suggest that DNA methylation defects in ICF syndrome may also result fromimpaired stimulation of DNMT3B activity by DNMT3L or other unknown regulatoryfactors apart from a weakened basal catalytic activity of the mutant DNMT3B proteinper se. This study provides evidence for a novel view of the pathogenesis of the ICFsyndrome and further suggests a role of DNMT3L in the stimulation of DNMT3Bactivity in early embryonic development.