ESP13.2, a member of the beta-defensin family, is a macaque sperm surface-coating protein involved in the capacitation process.
Female macaques produced isoantibodies to a limited number of sperm surface proteins following immunization with sperm components released by phosphatidylinositol-specific phospholipase C (PI-PLC). Washed, acrosome-intact, fixed sperm injected into rabbits elicited a major immune response to one of the same PI-PLC-released proteins, which was shown to be a sperm surface-coating protein. After purification and digestion of the glycoprotein, four peptides were analyzed for amino acid sequence, and all had 100% homology with an epididymal secretory protein, ESP13.2, reported previously to be a small, cationic-rich peptide and a member of the beta-defensin family. Antibodies to purified ESP13.2 recognized a number of protein bands on Western blots of nonreduced PI-PLC-released sperm components and nonreduced whole-sperm extracts. After chemical disulfide reduction, only a single, broad band from 31 to 35 kDa was recognized by anti-ESP13.2 antibodies. Indirect immunofluorescence showed ESP13.2 over the entire surface of ejaculated macaque sperm. Fluorescence was only slightly reduced after sperm were washed through 80% Percoll. A 24-h incubation in capacitating medium significantly reduced the amount of ESP13.2 over the head and midpiece, whereas exposure of the incubated sperm to dbcAMP and caffeine (capacitation activators) resulted in almost complete loss of ESP13.2 from the sperm surface. After activation, ESP13.2 was the primary component released into the medium as judged electrophoretically. Lignosulfonic acid, a potent inhibitor of macaque fertilization in vitro, completely blocked release of ESP13.2 from the sperm surface, even following treatment with activators. These findings suggest that the beta-defensin, ESP13.2, has a function in the capacitation of macaque spermatozoa and may modulate sperm surface-receptor presentation at the time of fertilization.
Yudin
Ashley I
AI
Department of Obstetrics and Gynecology, Division of Reproductive Biology, University of California, Davis 94923, USA.
Tollner
Theodore L
TL
Li
Ming-Wen
MW
Treece
Cathy A
CA
Overstreet
James W
JW
Cherr
Gary N
GN
D000595:Amino Acid Sequence; D000818:Animals; D002110:Caffeine; D002462:Cell Membrane / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D005260:Female; D005837:Genitalia, Male / Q000737:chemistry; D007518:Isoantibodies / Q000276:immunology; D008031:Lignin / Q000031:analogs & derivatives* / Q000494:pharmacology; D008252:Macaca fascicularis / Q000502:physiology*; D008297:Male; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D011817:Rabbits; D013075:Sperm Capacitation / Q000187:drug effects / Q000502:physiology*; D013094:Spermatozoa / Q000737:chemistry* / Q000187:drug effects / Q000502:physiology; D023083:beta-Defensins / Q000737:chemistry / Q000187:drug effects / Q000302:isolation & purification / Q000502:physiology*
D007518:Isoantibodies; D023083:beta-Defensins; D002110:Caffeine; C001545:lignosulfuric acid; D008031:Lignin