Abstract

Systemic characterization of genes and pathways underlying the genetic architecture of type 2 diabetes (T2D) requires scalable functional genomics approaches. Molecular readouts from CRISPR perturbations can effectively uncover the mechanistic effects of underexplored genes. Here we performed single-cell RNA sequencing on pooled CRISPR screens (Perturb-seq) of 61 T2D-associated genes and 40 ribosome-associated quality control (RQC) genes in human pancreatic β cells (EndoC-βH1) for investigations of insulin production and T2D pathology. We identified 21 functional genes, including the uncharacterized KLHL42 and ZZEF1 . Findings from global and β cell-specific knockout male mice, islet organoids and human islets reveal that ZZEF1 is a regulator of insulin synthesis and β cell stress through ribosomal stress-surveillance pathways in working and stress status-defined β cell subtypes. ZZEF1 deficiency impairs β cell function by inhibiting the RQC sensor EDF1, which could be improved by azoramide and ISRIB treatments. These Perturb-seq analyses and identification of functional RQC-related genes can provide potential therapeutic targets for T2D.