VCY2 protein interacts with the HECT domain of ubiquitin-protein ligase E3A

VCY2 locates in the AZFc region on chromosome Yq and is frequently deleted in infertile men with severe oligozoospermia or azoospermia. VCY2 is a testis-specific protein with unknown function. This study was to identify the protein that interacts with VCY2. We used the full-length VCY2 as bait to screen the human testis cDNA library using yeast two-hybrid approach. We identified a number of positive-interacting clones that encode ubiquitin-protein ligase E3A (UBE3A). UBE3A contains a HECT domain that binds VCY2. The specificity of the interaction was confirmed by co-immunoprecipitation and yeast mating. Northern blot analyses revealed two UBE3A transcripts 1.4 and 2kb that were abundantly expressed in human testis. We also showed that both VCY2 and UBE3A mRNAs were expressed in ejaculated human spermatozoa, indicating that both genes localize in the germ cell compartment. These data suggest that UBE3A ubiquitination may be required for VCY2 function.