RAB3IL1: a key regulator in the autophagy-ferroptosis axis of male infertility

Abstract Ferroptosis, an iron-dependent form of regulated cell death characterized by phospholipid peroxidation, plays a pivotal role in various diseases. However, its involvement in male infertility, particularly in idiopathic non-obstructive azoospermia (iNOA), remains largely unexplored. Idiopathic non-obstructive azoospermia accounts for 10–15% of male infertility cases, presenting a significant clinical challenge due to its unknown origins. This study investigated the potential link between ferroptosis and iNOA, revealing excessive activation of both ferroptosis and autophagy in the testes of iNOA patients, along with severe disruption of the blood-testis barrier (BTB). The BTB, formed by tight junctions between Sertoli cells, is essential for maintaining the spermatogenesis microenvironment. Using the ferroptosis inducer erastin, an antagonist of the cystine antiporter Solute Carrier Family 7 Member 11 (SLC7A11), we established a ferroptosis model in mouse Sertoli cells, demonstrating that ferroptosis activation led to cytoskeletal disarray and BTB disruption, accompanied by excessive autophagy activation. Notably, inhibition of autophagy using 3-methyladenine significantly rescued erastin-induced ferroptosis and restored BTB integrity, highlighting a crucial cross-talk between ferroptosis and autophagy in maintaining the spermatogenic microenvironment. Furthermore, through combined transcriptome analysis of iNOA testes and Sertoli cells, we identified RAB3IL1 as a novel regulator of ferroptosis. Mechanistically, Rab3il1 knockdown induced ferroptosis via translocation of SLC7A11, resulting in cytoskeletal defects and BTB damage. Our findings underscore the critical role of the ferroptosis-autophagy axis, regulated by RAB3IL1, in preserving the spermatogenic microenvironment, offering a potential therapeutic target for restoring spermatogenesis in iNOA patients.