Mapping the human sperm proteome across compartments, donors and individual cells

The human spermatozoon is among the most specialised cells in the body, yet where its proteins reside, where they come from and how they vary between and within men remain largely unresolved. Here we present the deepest proteomic atlas of human spermatozoa to date, identifying 10,828 proteins across six donors. Abundances correlate well between individuals and are anchored by a conserved core of mitochondrial energy-metabolism proteins, while a smaller fraction varies between donations from the same man. By mechanically separating heads from tails, we assigned thousands of previously unlocalised proteins to a compartment, and by comparing seminal fluid from vasectomised and non-vasectomised men we distinguished sperm-and epididymis-derived proteins from accessory-gland secretions. Using a single-cell analysis workflow we further show that the proteomes of individual spermatozoa can be measured, recovering up to ∼1,000 protein groups per cell. In a cohort with total fertilisation failure, we find recurrent mitochondrial rather than sperm–egg fusion dysregulation. Together or data provides novel insights into the dynamics of the sperm proteome and male infertility