Dissecting ELANE neutropenia pathogenicity by human HSC gene editing

Severe congenital neutropenia (SCN) is a life-threatening disorder most often caused by dominant mutations of ELANE that interfere with neutrophil maturation. We conducted a pooled CRISPR screen in human hematopoietic stem and progenitor cells (HSPCs) that correlated ELANE mutations with neutrophil maturation potential. Highly efficient gene editing of early exons elicited nonsense-mediated decay (NMD), overcame neutrophil maturation arrest in HSPCs from ELANE-mutant SCN patients, and produced normal hematopoietic engraftment function. Conversely, terminal exon frameshift alleles that mimic SCN-associated mutations escaped NMD, recapitulated neutrophil maturation arrest, and established an animal model of ELANE-mutant SCN. Surprisingly, only −1 frame insertions or deletions (indels) impeded neutrophil maturation, whereas −2 frame late exon indels repressed translation and supported neutrophil maturation. Gene editing of primary HSPCs allowed faithful identification of variant pathogenicity to clarify molecular mechanisms of disease and encourage a universal therapeutic approach to ELANE-mutant neutropenia, returning normal neutrophil production and preserving HSPC function. [Display omitted] •Primary cell CRISPR screen reveals ELANE mutations that arrest neutrophil maturation•Gene-edited xenograft establishes animal model of ELANE-mutant neutropenia•Indels triggering nonsense-mediated decay (NMD) circumvent neutropenia mutations•–2 frame indels that escape NMD repress translation and enable neutrophil maturation Rao et al. perform comprehensive gene editing in human primary cells to associate ELANE mutations with neutrophil maturation potential. Indels that trigger nonsense-mediated decay (NMD) circumvent severe congenital neutropenia (SCN) mutations. Indels that escape NMD model SCN by xenograft, although −2 frame indels repress translation and enable neutrophil maturation.