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J. Biol. Chem., Vol. 278, Issue 33, 30506-30515, August 15, 2003

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SAMP14, a Novel, Acrosomal Membrane-associated, Glycosylphosphatidylinositol-anchored Member of the Ly-6/Urokinase-type Plasminogen Activator Receptor Superfamily with a Role in Sperm-Egg Interaction^*

Jagathpala Shetty, Michael J. Wolkowicz, Laura C. Digilio, Kenneth L. Klotz, Friederike L. Jayes, Alan B. Diekman, V. Anne Westbrook, Erin M. Farris, Zhonglin Hao, Scott A. Coonrod, Charles J. Flickinger and John C. Herr 

From the Department of Cell Biology, Center for Research in Contraceptive and Reproductive Health, University of Virginia, Charlottesville, Virginia 22908

We report a new member of the Ly-6/urokinase-type plasminogen activator receptor (uPAR) superfamily of receptors, SAMP14, which is retained on the inner acrosomal membrane of the human spermatozoan following the acrosome reaction and may play a role in fertilization. The SAMP14 sequence predicted a glycosylphosphatidylinositol (GPI)-anchored protein with a signal peptide, a transmembrane domain near the carboxyl terminus, and a putative transamidase cleavage site in the proprotein. Attachment of SAMP14 to the membrane by a lipid anchor was confirmed by its sensitivity to phosphatidylinositol phospholipase C. SAMP14 has a single functional domain similar to the Ly-6 and urokinase plasminogen activator receptor superfamily of proteins, and the gene mapped to 19q13.33, near the PLAUR locus for uPAR at 19q13.2. Northern and dot blotting showed that SAMP14 expression was testis-specific. Indirect immunofluorescence and immunoelectron microscopy with antisera to purified recombinant SAMP14 localized the protein to outer and inner acrosomal membranes as well as the acrosomal matrix of ejaculated human sperm. Acrosome-reacted sperm demonstrated SAMP14 immunofluorescence, indicating its retention on the inner acrosomal membrane following the acrosome reaction. However, SAMP14 localized to the entire sperm when unwashed swim-up sperm from the ejaculate were stained, indicating that some SAMP14 is loosely associated with the plasma membrane. Antibodies against recombinant SAMP14 inhibited both the binding and the fusion of human sperm to zona free hamster eggs, suggesting that SAMP14 may have a role in sperm-egg interaction. SAMP14 represents a GPI-anchored putative receptor in the Ly-6/uPAR family that is exposed on the inner acrosomal membrane after the acrosome reaction.

Received for publication, February 19, 2003 , and in revised form, May 29, 2003.

The nucleotide sequence(s) reported in this paper has been submitted to the GenBank^TM/EBI Data Bank with accession number(s) AF353721.

^* This work was supported by D43 TW/HD 00654 from the Fogarty International Center, National Institutes of Health Grant HD U54 29099, the Andrew W. Mellon Foundation, Schering A.G., and Office of Justice Programs, National Institute of Justice, United States Department of Justice Grant 2000-IJ-CX-K013. 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, Center for Research in Contraceptive and Reproductive Health, P.O. Box 800732, University of Virginia, Charlottesville, VA 22908. Tel.: 434-924-2007; Fax: 434-982-3912; E-mail: jch7k{at}virginia.edu.

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