Endogenous fine-mapping of functional regulatory elements in complex genetic loci in HAP1 [scRNA-seq]
The vast majority of genetic loci associated with polygenic complex traits are located in non-coding regions of the human genome. Many of these regions exhibit high-order gene regulatory relationships and complicated linkage disequilibrium (LD) configurations, which bring challenges to accurately identify causal variants and their target genes controlling specific molecular processes or traits. We employed multiplexed single-cell CRISPR interference and activation perturbations to explore the links between cis-regulatory element (CRE) and target gene expression within tight LD in the endogenous chromatin context. We validated the prevalence of multiple causality in perfect LD (pLD) for independent expression quantitative trait locus (eQTL), and revealed fine-grained genetic effects on gene expression within pLD. These effects are difficult to decipher using conventional eQTL fine-mapping or to predict via existing computational methods. We found that nearly half of the casual CREs lack classical epigenetic markers, potentially affecting gene expression through hidden regulatory mechanisms. Integrative analysis on different types of perturbation effects suggested a high regulatory plasticity of the human genome. These findings will propel further in-depth exploration of functional genomic elements, facilitating a more comprehensive understanding of gene expression regulatory patterns and the development of complex traits. Overall design: Four scRNA-seq datasets of a moderate multiplicity-of-infection (MOI) sgRNA screen and high MOI sgRNA screeen in dCas9-KRAB-blast or dCAS9-VP64-blast stably expressed HAP1 clones , in addition to three 4C-seqs used to to valify the regulatory relationship of the causal CRE and target genes in HAP1 cells.