Interactome network of the protein EPPIN in human spermatozoa: functional characterization and applications on the development of pharmacological strategies for male contraception

Abstract

Almost 30% of couples rely on the two currently available options of male contraception: condoms and vasectomy, which have issues regarding effectiveness and reversibility, respectively. Thus, there is an unmet need for new male contraceptive methods, which will contribute to decreasing unintended pregnancy rates and to improve family planning, health and well-being. Protein-protein interactions regulating sperm function are compelling targets for non-hormonal, fast-onset male contraceptive drugs. The sperm-bound protein EPPIN (epididymal protease inhibitor) is a hub for the interaction of semenogelin-1 (SEMG1), the most abundant protein of the human seminal plasma, which leads to a transient inhibition of their motility after ejaculation. The demonstration that EPPIN ligands inhibited human sperm motility confirmed EPPIN druggability, thus triggering the development of first-in-class spermostatic compounds with male contraceptive potential. In this scenario, further understanding EPPIN's roles and mechanisms underlying EPPIN roles on the regulation of sperm motility are crucial for the development of safe and effective male contraceptive EPPIN-binding drugs. This project aims to investigate the EPPIN interactome network in human spermatozoa, to understand its molecular interactions and mechanisms governing sperm function. We will co-immunoprecipitate EPPIN protein complexes from human spermatozoa and identify them using state-of-the-art quantitative label-free mass spectrometry. We will validate the results and determine EPPIN sequences playing key roles in the interaction with its binding partners using molecular and protein-protein interaction assays. Finally, we will investigate the relative stability of EPPIN, and its co-localization with binding partners after coitus using mice as experimental models. Our proposal will pave the way for investigations on sperm protein-protein interactions as drug targets for male contraception, thus contributing to radical innovation of birth-control methods. (AU)