Endocrine Abstracts

Primary ovarian insufficiency (POI) is a highly heterogeneous condition defined by the occurrence of amenorrhoea, hypoestrogenism and hypergonadotropinism in women under 40. POI onset can be triggered by multiple factors, such as iatrogenic events, environmental conditions, autoimmunity or genetic alterations. When the ovarian insufficiency occurs as a consequence of either chromosomal or genetic alterations, it can be associated with other congenital abnormalities and classified as part of a syndrome. Genetic alterations of the transcription factor FOXL2 are associated with the onset of the blepharophimosis, ptosis and epicantus inversus syndrome (BPES), an autosomal dominantly inherited condition in which eyelid dysplasia occur either in association (BPES type 1) or not (BPES type 2) with POI. By genetic screening of several BPES-affected women we identified two novel nonsense FOXL2 variants: p.E92* variant in a fertile woman with regular menses at the time of sampling and her prepuberal daughter, and p.Y186* variant in a woman with primary amenorrhea. To unravel the impact of the novel variants on protein function, the subcellular localization of WT and mutant FOXL2 was investigated in transiently transfected HeLa cells by confocal microscopy. Surprisingly, while both FOXL2-WT and FOXL2-p.Y186* signals were confined to the nucleus, mislocalization of the FOXL2-p.E92* variant to mitochondria was revealed by colocalization with the mitochondrial marker TOM20 and confirmed by cellular fractionation. Moreover, evaluation of mitochondrial morphology in FOXL2-p.E92* cells revealed a significant shift from an interconnected network of mitochondria to a more fragmented state compared to FOXL2-WT- and FOXL2-p.Y186* cells. On this regard, it is widely known that mitochondrial fusion-to-fission rate is finely tuned in response to metabolic or environmental stress, and that an increase in mitochondrial fission enables the removal of damaged mitochondria and facilitates apoptosis. The increase in mitochondrial fragmentation in FOXL2-p.E92* cells therefore suggests that this variant might have a toxic effect on cell metabolism. This hypothesis is further supported by its increased degradation by the ubiquitin-proteasome system, as confirmed by treatment with the proteasome inhibitor MG132.In all the experiments conducted thus far, the FOXL2-p.Y186* variant behaviour did not differ from that of the WT, and further studies are underway to characterize its functionality inside the nucleus. On the other hand, given the importance of mitochondrial dynamics in both oocyte metabolism and tissue aging, our experimental data suggest that the FOXL2-p.E92* might eventually determine the onset of POI in the BPES-affected patients.