Single substitution in H3.3G34 alters DNMT3A recruitment to cause progressive neurodegeneration [multiome]

Germline histone H3.3 amino acid substitutions, including H3.3G34R/V, cause severe neurodevelopmental syndromes. To understand how these mutations impact brain development, we generated H3.3G34R/V/W knock-in mice and identified strikingly distinct developmental defects for each mutation. H3.3G34R-mutants uniquely exhibited progressive microcephaly and neurodegeneration, with abnormal accumulation of disease-associated microglia and concurrent neuronal depletion. G34R severely decreased H3K36me2 on the mutant H3.3 tail, impairing recruitment of DNA methyltransferase DNMT3A. Redistribution of DNMT3A on chromatin promoted sustained expression of complement and other innate immune genes through loss of non-CG (CH) methylation, and silencing of neuronal gene promoters through aberrant CG methylation. Persistent complement expression in G34R neurons led to neuroinflammation possibly accounting for progressive neurodegeneration. Our study reveals that H3.3G34-substitutions have differential impact on the epigenome, which underlie the diverse phenotypes observed, and uncovers unappreciated roles for H3K36me2 and DNMT3A-dependent CH-methylation in modulating synaptic pruning and neuroinflammation in post-natal brains. 6 multiome samples (expression and ATAC-seq). Comparison of mutant (H3f3a +/G34R, n = 3) and wild type (WT, n = 3) murine cortex