Nicotinic acetylcholine receptor subunits and associated proteins in human sperm

We demonstrated previously the involvement of a nicotinic acetylcholine receptor containing an alpha7 subunit in the human sperm acrosome reaction (a modified exocytotic event essential to fertilization). Here we report the presence in human sperm of alpha7, alpha9, alpha3, alpha5, and beta4 nicotinic acetylcholine receptor subunits and the following proteins known to be associated with the receptor in the somatic cell: rapsyn and the tyrosine kinases c-SRC and FYN. The alpha7 subunit appears to exist as a homomer in the posterior post-acrosomal and neck regions of sperm and is probably linked to the cytoskeleton via rapsyn. The alpha3, alpha5, and beta4 subunits are present in the sperm flagellar mid-piece of sperm and possibly exist as alpha3alpha5beta4 and/or alpha3beta4 channels. The alpha9 subunit is present in the sperm mid-piece. We detected the FYN and c-SRC tyrosine kinases in the flagellar mid-piece region. Both co-precipitated only with the nicotinic acetylcholine receptor beta4 subunit. Immunolocalization with a C-terminal SRC kinase antibody, which recognizes several members of SRC kinase family, detected a SRC kinase co-localized with the alpha7 subunit in the neck region of sperm. Immunoprecipitation studies with that antibody demonstrated that the alpha7 subunit is associated with a SRC kinase. Antagonists of tyrosine phosphorylation inhibited the acetylcholine-initiated acrosome reaction, suggesting the involvement of a SRC kinase in the acrosome reaction.