Single-nuclei RNA-seq of neural organoids generated from wildtype and FNDC4 KO iPSCs

Large-cohort genome-wide association studies (GWAS) for alcohol use disorder (AUD) risk and drug treatment outcomes have identified significant genetic loci that are splicing quantitative trait loci (sQTLs) for the FNDC4 gene in multiple human brain regions. However, in spite of the fact that FNDC4 is highly expressed in the brain, its function and how it might contribute to AUD pathophysiology remain unknown. In the precent study, we characterized FNDC4 function using CRISPR/cas9 gene editing, the creation of human induced pluripotent stem cell (iPSC)-derived neural organoids and with single-nucleus RNA sequencing (snRNA-seq). Overall design: Specifically, we generated FNDC4 homozygous knock-out (KO) iPSC lines and differentiated two single-colony KO iPSCs, together with wildtype (WT) iPSCs, to generate forebrain organoids. Those neural organoids were harvested at three time points of differentiation (45, 90, and 150 days) for snRNA-seq using a “split-pool” combinatorial barcoding technology (Parse Bio).