Developmental Epigenetic Modification Regulates Stochastic Expression of Clustered Protocadherin Genes, Generating Single Neuron Diversity

•Pcdh promoters are differentially methylated by Dnmt3b at early embryonic stages•Promoter methylation controls the allocation of Pcdh isoform expression•Dnmt3b-KO alters the stochastic expression of Pcdh isoforms in Purkinje cells•Dnmt3b-KO Purkinje cells have defective dendritic arborization

SummaryIn the brain, enormous numbers of neurons have functional individuality and distinct circuit specificities. Clustered Protocadherins (Pcdhs), diversified cell-surface proteins, are stochastically expressed by alternative promoter choice and affect dendritic arborization in individual neurons. Here we found that the Pcdh promoters are differentially methylated by the de novo DNA methyltransferase Dnmt3b during early embryogenesis. To determine this methylation’s role in neurons, we produced chimeric mice from Dnmt3b-deficient induced pluripotent stem cells (iPSCs). Single-cell expression analysis revealed that individual Dnmt3b-deficient Purkinje cells expressed increased numbers of Pcdh isoforms; in vivo, they exhibited abnormal dendritic arborization. These results indicate that DNA methylation by Dnmt3b at early embryonic stages regulates the probability of expression for the stochastically expressed Pcdh isoforms. They also suggest a mechanism for a rare human recessive disease, the ICF (Immunodeficiency, Centromere instability, and Facial anomalies) syndrome, which is caused by Dnmt3b mutations.