Non-obstructive azoospermia (NOA) is a common cause of male infertility characterized by the absence of sperm due to spermatogenic failure. NOA is frequently associated with deletions in Azoospermia factor (AZF) regions a, b and c of the human Y chromosome. Although AZF deletions are the most common genetic cause of male infertility, the individual AZF gene(s) responsible for NOA remain unknown. Kdm5d is a single copy gene located in the AZFb deleted region and encodes a histone demethylase expressed in pre-meiotic/early meiotic testicular germ cells. My preliminary data in mice lacking Kdm5d demonstrates that these mice have azoospermia and meiotic arrest of testicular germ cells, mirroring observations in men carrying AZFb deletions. The goal of my studies is to uncover the molecular basis of Kdm5d-dependent NOA in Kdm5d null mice and determine if disruption of KDM5D is responsible for NOA in humans. I will achieve this goal through the completion of three independent aims. In Aim 1, I will assess the necessity of Kdm5d for male fertility, spermatogenesis and in specific meiotic processes by characterizing reproductive phenotypes and meiotic progression in mice lacking Kdm5d. In Aim 2, I will test if Kdm5d is necessary for removal of chromatin methylation modifications at programmed meiotic double strand break sites (DSB). I will utilize cleavage under targets and release using nuclease (CUT & RUN) sequencing combined with FACS sorting of distinct staged meiotic cells to determine whether a chromatin modification that specifies meiotic DSBs remains present in the absence of Kdm5d. Finally, in Aim 3 I will determine if loss of KDM5D underlies NOA in men carrying potential loss of function point mutations in KDM5D. I will generate mouse models of Kdm5d missense mutations found in men with NOA using a conditional gene targeting by inversion (“COIN”) approach to determine if these mutant mouse lines recapitulate phenotypes found in Kdm5d null male mice. Overall my studies will determine the role of Kdm5d in NOA and will provide insight into the elusive genetic underpinnings of AZFb deletions in infertile men. These studies also have potential to inform our understanding of the propensity for single copy Y- linked genes located within AZF regions to contribute to male infertility and may thus provide a new gene for diagnostic testing of men with NOA.