Distinct disease mutations in DNMT3A result in a spectrum of behavioral, epigenetic, and transcriptional deficits.

Phenotypic heterogeneity in monogenic neurodevelopmental disorders can arise from differential severity of variants underlying disease, but how distinct alleles drive variable disease presentation is not well understood. Here, we investigate missense mutations in DNA methyltransferase 3A (DNMT3A), a DNA methyltransferase associated with overgrowth, intellectual disability, and autism, to uncover molecular correlates of phenotypic heterogeneity. We generate a Dnmt3aP900L/+ mouse mimicking a mutation with mild to moderate severity and compare phenotypic and epigenomic effects with a severe R878H mutation. P900L mutants exhibit core growth and behavioral phenotypes shared across models but show subtle epigenomic changes, while R878H mutants display extensive disruptions. We identify mutation-specific dysregulated genes that may contribute to variable disease severity. Shared transcriptomic disruption identified across mutations overlaps dysregulation observed in other developmental disorder models and likely drives common phenotypes. Together, our findings define central drivers of DNMT3A disorders and illustrate how variable epigenomic disruption contributes to phenotypic heterogeneity in neurodevelopmental disease.

Distinct disease mutations in DNMT3A result in a spectrum of behavioral, epigenetic, and transcriptional deficits.

Phenotypic heterogeneity is a common feature of monogenic neurodevelopmental disorders that can arise from differential severity of missense variants underlying disease, but how distinct alleles impact molecular mechanisms to drive variable disease presentation is not well understood. Here, we investigate missense mutations in the DNA methyltransferase DNMT3A associated with variable overgrowth, intellectual disability, and autism, to uncover molecular correlates of phenotypic heterogeneity in neurodevelopmental disease. We generate a DNMT3A P900L/+ mouse model mimicking a disease mutation with mild-to-moderate severity and compare phenotypic and epigenomic effects with a severe R878H mutation. We show that the P900L mutation leads to disease-relevant overgrowth, obesity, and social deficits shared across DNMT3A disorder models, while the R878H mutation causes more extensive epigenomic disruption leading to differential dysregulation of enhancers elements. We identify distinct gene sets disrupted in each mutant which may contribute to mild or severe disease, and detect shared transcriptomic disruption that likely drives common phenotypes across affected individuals. Finally, we demonstrate that core gene dysregulation detected in DNMT3A mutant mice overlaps effects in other developmental disorder models, highlighting the importance of DNMT3A-deposited methylation in neurodevelopment. Together, these findings define central drivers of DNMT3A disorders and illustrate how variable disruption of transcriptional mechanisms can drive the spectrum of phenotypes in neurodevelopmental disease.

Functional and epigenetic phenotypes of humans and mice with DNMT3A Overgrowth Syndrome.

Germline pathogenic variants in DNMT3A were recently described in patients with overgrowth, obesity, behavioral, and learning difficulties (DNMT3A Overgrowth Syndrome/DOS). Somatic mutations in the DNMT3A gene are also the most common cause of clonal hematopoiesis, and can initiate acute myeloid leukemia (AML). Using whole genome bisulfite sequencing, we studied DNA methylation in peripheral blood cells of 11 DOS patients and found a focal, canonical hypomethylation phenotype, which is most severe with the dominant negative DNMT3AR882H mutation. A germline mouse model expressing the homologous Dnmt3aR878H mutation phenocopies most aspects of the human DOS syndrome, including the methylation phenotype and an increased incidence of spontaneous hematopoietic malignancies, suggesting that all aspects of this syndrome are caused by this mutation.

DNMT3A Haploinsufficiency Results in Behavioral Deficits and Global Epigenomic Dysregulation Shared across Neurodevelopmental Disorders.

Mutations in DNA methyltransferase 3A (DNMT3A) have been detected in autism and related disorders, but how these mutations disrupt nervous system function is unknown. Here, we define the effects of DNMT3A mutations associated with neurodevelopmental disease. We show that diverse mutations affect different aspects of protein activity but lead to shared deficiencies in neuronal DNA methylation. Heterozygous DNMT3A knockout mice mimicking DNMT3A disruption in disease display growth and behavioral alterations consistent with human phenotypes. Strikingly, in these mice, we detect global disruption of neuron-enriched non-CG DNA methylation, a binding site for the Rett syndrome protein MeCP2. Loss of this methylation leads to enhancer and gene dysregulation that overlaps with models of Rett syndrome and autism. These findings define the effects of DNMT3A haploinsufficiency in the brain and uncover disruption of the non-CG methylation pathway as a convergence point across neurodevelopmental disorders.