Fig. 1

Schematic representation of the bioinformatic and experimental pipeline. Whole-exome sequencing data from 118 women with ovarian failure (cases) and 32 healthy controls collected in a previous study (Henarejos-Castillo et al., 2021) was uploaded to the OpenCGA platform to prioritize single nucleotide variants (SNVs) according to their predictive deleteriousness (REVEL and CADD predictors), minor allele frequency in the general population (IGSR and GnomAD databases), and statistical differences assessed by Fisher tests between the cases and control populations (including pseudo-control populations using female samples from IGSR and GnomAD). Next, a burden test prioritized genes with a significant presence of SNVs in the OF population compared to controls. To validate the impact of compromised genes with deleterious SNVs on female fertility, Drosophila females with mutations in orthologues of the prioritized genes were generated via RNAi-mediated silencing and gene-knockout and compared with control females (y, w). Silencing and knockout efficiency was evaluated by RT-qPCR, and the flies reproductive potential was assessed by histological analysis of ovarian morphology, proportion of mature oocytes and number of offspring produced