Rare variants of

Introduction: Recurrent spontaneous abortion (RSA) is associated with embryonic chromosomal abnormalities, yet the maternal genetic factors predisposing to the production of aneuploid gametes remain incompletely understood. Identifying germline variants that disrupt meiotic chromosome segregation may elucidate the genetic etiology of RSA. Methods: Peripheral blood DNA was collected from 100 RSA patients for whom aneuploidy of embryonic origin had been confirmed in prior miscarriage specimens, and whole-exome sequencing (WES) was performed. Rare missense variants in candidate meiotic genes were identified and prioritized for functional analysis. The effect of GPSM2 loss-of-function was modeled via siRNA-mediated knockdown in germ cell lines, and meiotic progression, ploidy outcomes, and spindle-associated protein localization were assessed in vitro and in vivo. Results: Two patients harbored rare heterozygous missense point mutations in GPSM2 (G protein signaling modulator 2). GPSM2 knockdown resulted in a marked increase in diploid spermatocytes (NC: 10.6% vs. GPSM2-siRNA: 53.8%) and a corresponding decrease in haploid spermatids (NC: 57.8% vs. GPSM2-siRNA: 5.49%), indicating failure of the first meiotic division, while earlier spermatogenic stages remained unaffected. Mechanistically, GPSM2 was found to interact with the nuclear mitotic apparatus (NUMA) protein; upon GPSM2 depletion, chromosome separation was arrested at metaphase in GC-2 cells, and NUMA failed to localize to the spindle poles. Discussion: These findings suggest that rare GPSM2 missense mutations predicted to impair gene function may compromise first meiotic chromosome segregation through disruption of the GPSM2-NUMA interaction at the spindle pole, thereby predisposing carriers to produce aneuploid germ cells. Fertilization of such aneuploid gametes would yield chromosomally abnormal embryos, providing a mechanistic basis for RSA in these patients. However, definitive causal attribution awaits direct functional characterization of the specific missense variants identified, as the current study employed siRNA-mediated knockdown as a surrogate loss-of-function model.