Comparative analysis reveals distinctive epigenetic features of the human cerebellum

Abstract

Identifying the molecular underpinnings of the neural specializations that underlie human cognitive and behavioral traits has long been of considerable interest. Much research on human-specific changes in gene expression and epigenetic marks has focused on the prefrontal cortex, a brain structure distinguished by its role in executive functions. The cerebellum shows expansion in great apes and is gaining increasing attention for its role in motor skills and cognitive processing, including language. However, relatively few molecular studies of the cerebellum in a comparative evolutionary context have been conducted. Here, we identify human-specific methylation in the lateral cerebellum relative to the dorsolateral prefrontal cortex, in a comparative study with chimpanzees (Pan troglodytes) and rhesus macaques (Macaca mulatta). Specifically, we profiled genome-wide methylation levels in the three species for each of the two brain structures and identified human-specific differentially methylated genomic regions unique to each structure. We further identified which differentially methylated regions (DMRs) overlap likely regulatory elements and determined whether associated genes show corresponding species differences in gene expression. We found greater human-specific methylation in the cerebellum than the dorsolateral prefrontal cortex, with differentially methylated regions overlapping genes involved in several conditions or processes relevant to human neurobiology, including synaptic plasticity, lipid metabolism, neuroinflammation and neurodegeneration, and neurodevelopment, including developmental disorders. Moreover, our results show some overlap with those of previous studies focused on the neocortex, indicating that such results may be common to multiple brain structures. These findings further our understanding of the cerebellum in human brain evolution.

Fig 1. Species relationships, brain structures, and sample clustering.

A = Phylogenetic tree of species included in this study from TimeTree.org [113], B = brain structures included in this study depicted on a chimpanzee brain illustration modified from [114], C and D = hierarchical clustering of samples from the dorsolateral prefrontal cortex (C) and cerebellum (D) based on genome-wide methylation after filtering and normalizing using correlations between samples as distance. DLPFC = dorsolateral prefrontal cortex.

Fig 2. DLPFC-specific human regDMRs associated with ADAM30 and FAM193A.

A = ADAM30, B = FAM193A. Lines are loess smoothed methylation values for each species across the DMR and each point represents raw methylation values for each individual at each CpG site within the DMR ranges. TSS200 = within 200 bp of a transcription start site and TSS1500 within 1500 bp of a transcription start site. Chromosomal coordinates are in base pairs and refer to human genome build hg19.

Fig 3. Cerebellum-specific human regDMRs associated with PLCH1, ANKS1B, DLGAP1, and PHACTR1.

A = PLCH1, B = ANKS1B, C = DLGAP1, D = PHACTR1. Lines are loess smoothed methylation values for each species across the DMR and each point represents raw methylation values for each individual at each CpG site within the DMR ranges. TSS200 = within 200 bp of a transcription start site and TSS1500 within 1500 bp of a transcription start site. Chromosomal coordinates are in base pairs and refer to human genome build hg19.

Fig 4. Cerebellum-specific human regDMRs and gene expression levels for CHAD, CPNE6, and STK33.

A = CHAD methylation, B = CHAD gene expression, C = CPNE6 methylation, D = CPNE6 gene expression, E = STK33 methylation, and F = STK33 gene expression. Lines are loess smoothed methylation values for each species across the DMR and each point represents raw methylation values for each individual at each CpG site within the DMR ranges. TSS200 = within 200 bp of a transcription start site and TSS1500 within 1500 bp of a transcription start site. Chromosomal coordinates are in base pairs and refer to human genome build hg19.