More than one-quarter of U.S. adults report past-year binge drinking and 5.6% meet criteria for a past-year
alcohol use disorder (AUD). These traits are associated with psychosocial disruption, medical co-morbidities,
and nearly 10% of all U.S. deaths annually. Alcohol consumption and AUD have an estimated twin heritability
of 43% and 49%, respectively. Genome-wide association studies (GWAS) of these traits have identified 380
genome-wide significant SNPs in 155 loci. In this revised application, we propose to use a “variant to gene
mapping to clinical impact” approach, intersecting GWAS data with ATAC-seq and high-resolution promoter-
focused Capture C neuronal datasets derived from human induced pluripotent stem cells (iPSCs) to identify
potential effector genes. To prioritize SNPs, we will focus on those residing in open chromatin and with
enhancer epigenetic marks; we will also use eQTL resources, as needed. We will validate these and newly
identified candidates in three ways: 1) in established Drosophila models of alcohol behavioral effects, 2) initially
with human iPSC-derived cortical and dopaminergic neurons and subsequently with neurons of other
neurotransmitter systems, as indicated, to delete the genomic neighborhood harboring a putative causal
variant to determine its effects on the gene's expression, and 3) with data from 4 biobanks to corroborate the
association of the functionally validated genes with alcohol-related phenotypes. In Aim 1, we will expand our
preliminary set of 43 loci from iPSC-derived cortical neurons, initially by applying similar methods to iPSC-
derived dopaminergic neurons and other neuronal types, as indicated. In Aim 2a, we will screen candidate
genes identified in preliminary findings and those identified in Aim 1 by knocking down or over-expressing their
orthologs in fly models of alcohol consumption, preference, sensitivity, and tolerance. In Aim 2b, we will use
SNP-CRISPR in human iPSC-derived neurons to test whether deleting the genomic neighborhood of putative
causal variants from preliminary data or those identified in Aim 1 affects expression of their target gene(s). In
Aim 3, we will use array and exome sequence data from both European- and African-ancestry individuals to
validate all putative effector genes by examining their association with alcohol-related traits, conducting
downstream analyses, testing rare variant effects, and examining pleiotropy in phenome-wide association
studies. This project will identify novel genetic variants and the corresponding effector genes that contribute to
alcohol-related traits, thereby shedding light on the biological pathways that influence the development of the
traits. Study results will have fundamental implications for novel approaches to the diagnosis, prevention, and
treatment of heavy drinking and AUD.
