Epigenomic landscape of the developing human rhombic lip reveals gene regulatory network and non-coding loci of developmental, evolutionary, and disease relevance.

The cerebellar rhombic lip neural progenitor niche of the prenatal hindbrain is an anatomical structure critical for cerebellar glutamatergic neurogenesis. Humans have elaborated the rhombic lip niche to include a rhombic lip subventricular zone (RL-SVZ) not seen in mice or macaques. Although developmental disruptions of this progenitor zone can cause cerebellar growth abnormalities - from malformations to tumors - the gene regulatory networks underpinning this unique progenitor niche are unknown. Here we provide a predicted gene regulatory network for the human cerebellar rhombic lip, inferred from epigenomic maps of the developing human cerebellum. We generated DNA methylomes of neuroanatomically-dissected mid-gestation human rhombic lip ventricular zone and RL-SVZ (N=9 samples; 15-16 post conception weeks) using low-input Enzymatic MethylSeq. We also mapped histone modifications marking active promoters and enhancers in the whole mid-gestation human fetal cerebellum (N=6 samples; 14 and 18 post-conception weeks). Integrating these data, we identified 9,855 differentially-methylated regions (DMR) which converge on binding sites of over three hundred transcription factors, including master regulators of rhombic lip neurogenesis, ATOH1, NEUROD1, and NEUROD2. DMRs hypomethylated in the RL-SVZ are enriched in active enhancers and in human accelerated regions, and are depleted in active promoters. We inferred 81,844 transcription factor-enhancer-gene links, covering 41 transcription factors active in the rhombic lip, and 4,610 target genes that include drivers of cerebellar neurogenesis and pediatric hindbrain cancer. Twenty-five DMRs overlap human accelerated regions located near genes associated with intellectual disability, autism spectrum disorders, and neurological deficits. DMRs are also statistically enriched in copy number aberrations in medulloblastoma, a malignant pediatric hindbrain cancer with subtypes hypothesized to originate in the rhombic lip. Close to one-quarter of the DMRs overlap known copy number aberrations in medulloblastoma, nominating potential enhancer and promoter elements impacted by these genomic aberrations. Collectively, our data provide a rich resource to start decoding the functional impact of non-coding variation on gene regulation in the developing cerebellum and on genomic dysregulation in diseases of cerebellar growth.