MeCP2 and non-CG DNA methylation stabilize the expression of long genes that distinguish closely related neuron types

The diversity of mammalian neurons is delineated by subtle gene expression differences that may require specialized mechanisms to be maintained. Neurons uniquely express the longest genes in the genome and use non-CG DNA methylation (mCA), together with the Rett syndrome protein methyl-CpG-binding protein 2 (MeCP2), to control gene expression. However, whether these distinctive gene structures and molecular machinery regulate neuronal diversity remains unexplored. Here, we use genomic and spatial transcriptomic analyses to show that MeCP2 maintains transcriptomic diversity across closely related neuron types. We uncover differential susceptibility of neuronal populations to MeCP2 loss according to global mCA levels and dissect methylation patterns driving shared and distinct MeCP2 gene regulation. We show that MeCP2 regulates long, mCA-enriched, ‘repeatedly tuned’ genes, that is, genes differentially expressed between many closely related neuron types, including across spatially distinct, vision-dependent gene programs in the visual cortex. Thus, MeCP2 maintains neuron type-specific gene programs to facilitate cellular diversity in the brain.