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J. Biol. Chem., Vol. 280, Issue 27, 25928-25935, July 8, 2005

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Nicotinic Acetylcholine Receptor Subunits and Associated Proteins in Human Sperm^*

Priyadarsini Kumar and Stanley Meizel

From the Department of Cell Biology and Human Anatomy, School of Medicine, University of California, Davis, California 95616

We demonstrated previously the involvement of a nicotinic acetylcholine receptor containing an 7 subunit in the human sperm acrosome reaction (a modified exocytotic event essential to fertilization). Here we report the presence in human sperm of 7, 9, 3, 5, and 4 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 7 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 3, 5, and 4 subunits are present in the sperm flagellar mid-piece of sperm and possibly exist as 354 and/or 34 channels. The 9 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 4 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 7 subunit in the neck region of sperm. Immunoprecipitation studies with that antibody demonstrated that the 7 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.

Received for publication, March 4, 2005 , and in revised form, April 18, 2005.

^* This work was supported by National Institutes of Health Grant HD 33368 (to S. M.) and a postdoctoral fellowship (to P. K.) from National Institutes of Health Fertilization and Early Development Training Grant 5 T32 HD007131. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

To whom correspondence should be addressed: Dept. of Cell Biology and Human Anatomy, University of California School of Medicine, Davis, CA 95616. Tel.: 530-752-3213; Fax: 530-752-8520; E-mail: pkumar{at}ucdavis.edu.

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