Identification of novel biallelic variants in BMP15 in two siblings with premature ovarian insufficiency.

Background: Premature ovarian insufficiency (POI) occurs in women before the age of 40 years, accompanied by amenorrhea, hypoestrogenism, hypergonadotropinism, and infertility. The pathology of POI is complex and the molecular genetic mechanisms are poorly understood. Bone morphogenetic protein 15 (BMP15) plays a crucial role in oocyte maturation and follicular development through the activation of granulosa cells. Dysfunction of BMP15 causes ovarian dysgenesis and is related to POI. Identifying pathogenic variants contributes to revealing genetic mechanisms and making clinical diagnoses of POI.

Methods: The study involved two sisters diagnosed with POI. Whole-exome sequencing (WES) was performed to identify causative genes. Sanger sequencing was used to validate the mutations in patients with POI and members of the family with no clinical signs or symptoms. The effect of the novel mutations on the BMP15 structure was analyzed by PSIPRED. By over-expressing wild-type (WT) or mutant BMP15 plasmids in vitro, a functional study of the BMP15 mutant was conducted by real-time qPCR and western blotting. Through cocultivation with HEK293T cells, the effects of secreted BMP15 WT and variants on granulosa cell proliferation and apoptosis were detected through a cell counting kit-8 assay and flow cytometric analysis.

Results: We identified biallelic variants in BMP15, c.791G > A (p. R264Q) and c.1076C > T (p. P359L), in two siblings with POI. Both sisters carried the same biallelic variants, while the other female members of their family carried only one of them. Structural prediction showed that the variants have not affected the secondary structure of BMP15 but may change the conformation of water molecules around protein surfaces and thermal stability of BMP15. Real-time qPCR showed no significant difference in mRNA levels among WT and the two variants. Western blotting indicated a reduction in BMP15 expression with the c.791G > A and c.1076C > T variants compared to WT. Moreover, mutants 791G > A and 1076C > T impaired the function of secreted BMP15 in promoting granulosa cell proliferation and suppressing cell apoptosis caused by reactive oxygen species.

Conclusions: This study identified novel biallelic variants, c.791G > A and c.1076C > T, of BMP15 in two siblings with POI. Both missense variants reduced the level of the BMP15 protein and impaired the function of BMP15 in promoting granulosa cell proliferation in vitro. Taken together, our findings provide a novel molecular genetic basis and potential pathogenesis of BMP15 variants in POI.