Sperm of both mammals and invertebrates move toward specific sites in the female reproductive tract. However, molecular mechanisms for sperm to follow directional cues are unknown. Here, we report genetic analysis of Drosophila Pkd2 at 33E3 (Pkd2, CG6504), which encodes a Ca(2+)-activated, nonselective cation channel homologous to the human Pkd2 autosomal dominant polycystic kidney disease (ADPKD) gene. The PKD2 family of genes has been implicated in sensory responses through protein localization on primary cilia of epithelia and neurons. In renal tubules, cilium-associated PKD2 appears to mediate Ca(2+) influx in response to fluid flow, and the loss of fluid sensation probably contributes to cyst growth and ADPKD. Sperm tails or flagella are specialized cilia essential for movement. Drosophila Pkd2 is abundantly associated with the tail and the acrosome-containing head region of mature sperm. Targeted disruption of Pkd2 results in male sterility without affecting spermatogenesis. The mutant sperm are motile but fail to swim into the storage organs in the female. Rare mutant sperm that reach the storage organs are able to fertilize the egg and produce viable progeny. Our data demonstrate that the Drosophila PKD2 cation channel operates in sperm for directional movement inside the female reproductive tract.