Pluto Bioinformatics

GSE152177: RNA-Seq and ATAC-Seq of HEK239 cell line carrying different genotypes at rs4420550 by CRISPR/Cas9 editing

Bulk RNA sequencing

Genome-wide association studies (GWAS) have reported hundreds of genomic loci associated with schizophrenia, yet identifying the functional risk variations is a key step in elucidating the genetic mechanisms. In the present study, we have identified the functional variations in the schizophrenia risk chromosome 16p11.2 region using bioinformatics and molecular approaches. We find that the schizophrenia GWAS risk variations in 16p11.2 show significant associations with mRNA expression of multiple genes in human brain, and one of the leading expression quantitative trait loci (eQTL) genes MAPK3 locates ~200-kb away in the genome. Further analyses using epigenome marks in human brain and cell lines implicate that a noncoding single nucleotide polymorphism (SNP) rs4420550 locates in the DNA enhancer region, which is validated via in vitro luciferase assays. Precise CRISPR/Cas9 editing in cells followed by RNA-Seq further confirm the regulatory effects of rs4420550 on 16p11.2 genes, among which MAPK3 is the leading affected gene whose mRNA and protein levels are both influenced. Chromosome conformation capture (3C) experiment shows that the rs4420550 region exhibits physical interaction with the MAPK3 promoter, and assay for transposase-accessible chromatin followed by sequencing (ATAC-Seq) further demonstrates that rs4420550 affects chromatin accessibility at the 16p11.2 region. These results together suggest that rs4420550 is a functional risk variation, which regulates MAPK3 expression in a long-range chromatin looping manner. This study provides valuable insights into the biological mechanisms underlying schizophrenia GWAS, and illustrates both an example of comprehensive functional characterization of GWAS risk loci and alerts for the complexity of SNP-gene interactions. SOURCE: Ming Li (limingkiz@mail.kiz.ac.cn) - Kunming Institute of Zoology, Chinese Academy of Sciences