DNMT1 mutations found in HSANIE patients affect interaction with UHRF1 and neuronal differentiation

Abstract

DNMT1 is recruited to substrate sites by PCNA and UHRF1 to maintain DNA methylation after replication. The cell cycle dependent recruitment of DNMT1 is mediated by the PCNA-binding domain (PBD) and the targeting sequence (TS) within the N-terminal regulatory domain. The TS domain was found to be mutated in patients suffering from hereditary sensory and autonomic neuropathies with dementia and hearing loss (HSANIE) and autosomal dominant cerebellar ataxia deafness and narcolepsy (ADCA-DN) and is associated with global hypomethylation and site specific hypermethylation. With functional complementation assays in mouse embryonic stem cells, we showed that DNMT1 mutations P496Y and Y500C identified in HSANIE patients not only impair DNMT1 heterochromatin association, but also UHRF1 interaction resulting in hypomethylation. Similar DNA methylation defects were observed when DNMT1 interacting domains in UHRF1, the UBL and the SRA domain, were deleted. With cell-based assays, we could show that HSANIE associated mutations perturb DNMT1 heterochromatin association and catalytic complex formation at methylation sites and decrease protein stability in late S and G2 phase. To investigate the neuronal phenotype of HSANIE mutations, we performed DNMT1 rescue assays and could show that cells expressing mutated DNMT1 were prone to apoptosis and failed to differentiate into neuronal lineage. Our results provide insights into the molecular basis of DNMT1 dysfunction in HSANIE patients and emphasize the importance of the TS domain in the regulation of DNA methylation in pluripotent and differentiating cells.

dnmt1function is required to maintain retinal stem cells within the ciliary marginal zone of the zebrafish eye

“…dnmt1 is expressed in RSCs at 4dpf ( Figure 4I,J), consistent with dnmt1's known requirements in stem cell populations in vivo 27,30,31,50,51 . Loss of Dnmt1 function results in aberrant gene expression in a number of contexts 45,50,52,53 and therefore we wanted to determine if CMZ-specific gene expression was altered in the dnmt1 -/-CMZ. Previous reports have characterized the expression/distribution of the CMZ-specific genes: col15a1b, cyclinD1, cdkn1c, and atoh7 14,15,54 .…”

A novel mutation in the DNMT1 gene in a patient presenting with pure cerebellar ataxia

“…A recently published study investigating mutations in exon 20 found that these mutations affect DNMT1 interaction with the essential cofactor UHRF1 (ubiquitin-like with PHD and ring finger domains 1), which is believed to cause cell cycle-dependent degradation of DNMT1 and lead to impaired neuronal differentiation [7]. Further studies reporting different cohorts with a variety of mutations in DNMT1 need to be published to further understand the pathogenic mechanism.…”

Genetics meets DNA methylation in rare diseases

“…In fact, DNMT1 is recruited to DNA by its obligate partner UHRF1 (ubiquitin‐like, containing PHD and RING finger domains 1) throughout S phase, an interaction suggested as promoting conformational changes and the unmasking of the DNMT1 catalytic domain . Indeed, HSAN1E mutations in the RFTS domain reproduced in the mouse disrupt the Dnmt1/Uhrf1 interaction, affect DNMT1's proper folding, and cause a cell cycle‐dependent degradation of Dnmt1that results in defective DNAme maintenance . They also affect the sub‐cellular localization of DNMT1 in neurons, promoting its translocation to the cytoplasm and preventing its interaction with DNA.…”