Identification of functional noncoding variants regulating Fc gamma receptor expression in human natural killer cells 2260713

Abstract Introduction Antibody-dependent cellular cytotoxicity (ADCC) is an important effector mechanism through which antibody-based therapies drive target cell clearance. Natural Killer (NK) cells mediate ADCC through Fc gamma receptor IIIa (FcγRIIIa) expressed on their surface, with variation in receptor abundance affecting cytotoxic activity and treatment efficacy. While coding polymorphisms in FcγR genes have been linked to altered receptor function, genome-wide association studies (GWAS) have also associated variation in noncoding regions of the low-affinity FcγR cluster with complex disease and treatment outcomes. However, the functional impact of these noncoding variants remains largely uncharacterized. Here, we aim to investigate noncoding variants in the Fc region with allele-specific protein binding that may influence receptor abundance and modulate NK cell-mediated ADCC. Methods To identify functional variants in the Fc region, we screened an oligonucleotide library containing sequences from 4,323 single nucleotide polymorphisms (SNPs) using high-throughput Regulatory Element Sequencing (Reel-seq). Unlike GWAS, which identifies context-specific associations, Reel-seq is an unbiased assay that detects allele-specific protein-DNA interactions through electrophoresis mobility shift assays (EMSA). This allows for the identification of variants with potential regulatory activity based on direct protein binding. Results Using Reel-seq, we identified a set of functional SNPs in human NK cell lines (NK-92, NKL). Ongoing work aims to attribute these variants to differences in FcγR expression and to map regions of regulatory activity within Fc that may influence receptor abundance. Conclusion Altogether, this work will define regulatory variants affecting FcγR abundance, with implications for antibody-based therapies. Funding Source National Institutes of Health Grant #1R01AI187318-01, North Carolina State University College of Veterinary Medicine Topic Categories Innate Immune Responses and Host Defense: Molecular Mechanisms (INM)