Schizophrenia (SZ) genome-wide association studies (GWAS) have identified >100 genome-wide significant (GWS) risk loci. However, risk loci typically span 1 gene and multiple GWS index and proxy SNPs in linkage disequilibrium, leaving causal genes/variants largely unknown, which hinders translating GWAS findings into disease biology and drug targets. It is thus imperative to identify at each locus the functional risk variants, the affected risk gene/s and the associated cellular phenotype changes. Most risk variants are noncoding, and likely influence gene expression through modulating chromatin accessibility to transcription factors (TF). Accessible (open) chromatin overlaps with cis-regulatory sequences and is enriched for common disease risk variants. Our pilot study in neurons derived from induced pluripotent stem cells (iPSCs) also showed an enrichment of SZ GWS index and proxy SNPs in open chromatin flanking TF footprints. We hypothesize that many causal variants at SZ loci modulate TF binding in open chromatin, thereby altering transcriptional and neuronal phenotypes relevant to SZ pathophysiology. We present an innovative approach to identify chromatin-modulating variants at each SZ locus, for both GWS index SNPs and proxy variants. We will directly compare the allele-specific effect of a heterozygous SNP on the quantitative measurements of open chromatin (i.e., ASoC) within the same individual, which minimizes experimental variation and increases assay sensitivity, allowing an effective study using a smaller sample size. We have identified a set of 20 super-heterozygous (super-het) subjects with >80% power to detect ASoC at 70 SZ loci. We found that chromatin openness correlated with gene expression changes from iPSC?neurons at loci of interest, and ASoC was prevalent in iPSC-neurons of a single subject. We propose to extend the ASoC assay to the 20 super-het subjects, and pursue three specific aims: (1) We will map open chromatin by ATAC-seq (Assay for Transposase-Accessible Chromatin by sequencing) in pathophysiologically relevant iPSC-derived neuronal stem cells, dopaminergic and glutmatergic neurons, and search for regulatory SZ-risk variants that present ASoC in sequences flanking TF footprints. (2) We will use multiplex CRISPR-Cas9 genome editing to generate pairs of isogenic iPSC lines that differ only for every 6~7 regulatory SZ-risk variants, and identify the gene(s) cis regulated by each variant by comparing expression differences between the pair of isogenic iPSC-derived neuronal cells. (3) We will prioritize synaptic genes cis-regulated by a regulatory SZ-risk variant and generate isogenic iPSCs differing only for the regulatory variant, and compare neuronal morphological, biochemical, and electrophysiological phenotypes. SZ is a devastating disorder afflicting 1% of the population without cure. This project will help move the field beyond SZ GWAS to deciphering causal mechanisms which will aid in the development of more effective treatments, and will also create a rich research resource of iPSCs carrying SZ GWS risk alleles.

Public Health Relevance

We will systemically identify open chromatin-modulating regulatory variants and their cis-regulated risk gene/s at loci implicated by schizophrenia in genome-wide association studies. The knowledge gained will substantially advance our understanding of the causal molecular mechanisms underlying these risk loci, which will facilitate disease risk prediction and treatment.

Agency
National Institute of Health (NIH)
Institute
National Institute of Mental Health (NIMH)
Type
Research Project (R01)
Project #
5R01MH106575-05
Application #
9853838
Study Section
Molecular Neurogenetics Study Section (MNG)
Program Officer
Panchision, David M
Project Start
2016-05-04
Project End
2021-01-31
Budget Start
2020-02-01
Budget End
2021-01-31
Support Year
5
Fiscal Year
2020
Total Cost
Indirect Cost
Name
Northshore University Healthsystem
Department
Type
DUNS #
069490621
City
Evanston
State
IL
Country
United States
Zip Code
60201
Kos, Mark Z; Duan, Jubao; Sanders, Alan R et al. (2018) Dopamine perturbation of gene co-expression networks reveals differential response in schizophrenia for translational machinery. Transl Psychiatry 8:278
Zhang, Siwei; Moy, Winton; Zhang, Hanwen et al. (2018) Open chromatin dynamics reveals stage-specific transcriptional networks in hiPSC-based neurodevelopmental model. Stem Cell Res 29:88-98
Doostparast Torshizi, Abolfazl; Duan, Jubao; Wang, Kai (2018) Transcriptional network analysis on brains reveals a potential regulatory role of PPP1R3F in autism spectrum disorders. BMC Res Notes 11:489
Duan, Jubao; Göring, Harald H H; Sanders, Alan R et al. (2018) Transcriptomic signatures of schizophrenia revealed by dopamine perturbation in an ex vivo model. Transl Psychiatry 8:158
Sanders, A R; Drigalenko, E I; Duan, J et al. (2017) Transcriptome sequencing study implicates immune-related genes differentially expressed in schizophrenia: new data and a meta-analysis. Transl Psychiatry 7:e1093
Forrest, Marc P; Zhang, Hanwen; Moy, Winton et al. (2017) Open Chromatin Profiling in hiPSC-Derived Neurons Prioritizes Functional Noncoding Psychiatric Risk Variants and Highlights Neurodevelopmental Loci. Cell Stem Cell 21:305-318.e8
Stueve, Theresa Ryan; Li, Wen-Qing; Shi, Jianxin et al. (2017) Epigenome-wide analysis of DNA methylation in lung tissue shows concordance with blood studies and identifies tobacco smoke-inducible enhancers. Hum Mol Genet 26:3014-3027
Blizinsky, Katherine D; Diaz-Castro, Blanca; Forrest, Marc P et al. (2016) Reversal of dendritic phenotypes in 16p11.2 microduplication mouse model neurons by pharmacological targeting of a network hub. Proc Natl Acad Sci U S A 113:8520-5
Shi, Jianxin; Park, Ju-Hyun; Duan, Jubao et al. (2016) Winner's Curse Correction and Variable Thresholding Improve Performance of Polygenic Risk Modeling Based on Genome-Wide Association Study Summary-Level Data. PLoS Genet 12:e1006493