Hostname: page-component-586b7cd67f-dsjbd Total loading time: 0 Render date: 2024-11-22T20:18:09.881Z Has data issue: false hasContentIssue false
  • English
  • Français

Evidence that acrosin activity is important for the development of fusibility of mammalian spermatozoa with the oolemma: inhibitor studies using the golden hamster

Published online by Cambridge University Press:  26 September 2008

Hiroko Takano ,
R. Yanagimachi  and
Umbert A. Urch
Hiroko Takano
Affiliation:
University of Hawaii and University of California, USA
R. Yanagimachi*
Affiliation:
University of Hawaii and University of California, USA
Umbert A. Urch
Affiliation:
University of Hawaii and University of California, USA
*
R. Yanagimachi, Department of Anatomy and Reproductive Biology, University of Hawaii School of Medicine, Honolulu, HI 96822, USA. Fax: (808) 956-5474.
Get access Rights & Permissions [Opens in a new window]

Summary

The sperm plasma membrane over the equatorial segment of the acrosome gains the ability to fuse with the oolemma some time during, or after, the acrosome reaction. Since acrosin is a major component of the acrosome matrix that dissolves during the acrosome reaction, we sought to determine the effect of acrosin inhibitors on the sperm's ability to fuse with the oolemma. Five acrosin inhibitors (soybean trypsin inhibitor (SBTI), leupeptin, benzamidine, N-p-tosyl-1-lysin-chloromethyl ketone (TLCK) and phenylmethylsulphonyl fluoride (PMSF) and one non-acrosin inhibitor (N-p-tosyl-1-phenylalanine chloromethyl ketone (TPCK) were tested at non-toxic levels (below motility-disturbing concentrations). These inhibitors were added at three different times: (1) during the acrosome reaction of spermatozoa, (2) during sperm-oocyte contact and fusion, and (3) soon after sperm-oocyte fusion was completed. TLCK prevented sperm-oocyte fusion by inhibiting the acrosome reaction.PMSF inhibited gamete fusion, without inhibiting the acrosome reaction. SBTI, leupeptin and benzamidine also inhibited gamete fusion, but they had no effect if spermatozoa were allowed to acrosome-react in inhibitor-free medium. TPCK was without any inhibitory effects, suggesting that chymotrypsin-like enzymes are not involved in gamete fusion. Although acrosin inhibitors prevented acrosome-reacted spermatozoa from becoming fusion-competent, acrosin (and trypsin) alone could not make the plasma membrane of acrosome-intact spermatozoa fusion-competent. The data suggest that (1) the plasma membrane of the acrosomal region first undergoes dramatic changes immediately before or during the acrosome reaction and (2) acrosin released from the acrosome during the acrosome reaction further alters biophysical and biochemical characteristics of the plasma membrane over the equatorial segment. Such dual changes make the plasma membrane of this specialised region of the spermatozoon competent to fuse with the oolemma. Acrosin may not be the only acrosomal enzyme to participate in these changes.

Keywords

AcrosinFertilisationFusionHamsterOocyteSperm
Type
Article
Information
Zygote , Volume 1 , Issue 1 , February 1993 , pp. 79 - 91
Copyright
Copyright © Cambridge University Press 1993

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

References

Ahkong, Q.F., Blow, A.M., Botham, G.M., Launder, J.M.,Quirk, S.J. & Lucy, J.A. (1978). Proteinases and cell fusion. FEBS Lett. 95, 147–52.CrossRefGoogle ScholarPubMed
Austin, C.R. & Bishop, M.W.H. (1958). Role of rodent acrosome and perforatorium in fertilization. Proc. R. Soc. Lond. B 149, 241–8.Google ScholarPubMed
Bavister, B.D.(1989).A consistently successful procedure for in vitro fertilization of golden hamster eggs. Gamete Res. 23, 139–58.CrossRefGoogle ScholarPubMed
Bedford, J.M., Moore, H.D.M. &Franklin, L.E. (1979).Significance of the equatorial segment of the acrosome of the spermatozoa in eutherian mammals. Exp. Cell Res. 119, 119–26.CrossRefGoogle ScholarPubMed
Blobel, C.P., Myles, D.G., Primakoff, P. & White, J.M.(1990). Proteolytic processing of a protein involved in sperm-egg fusion correlates with acquisition of fertilization com petence. J. Cell Biol. 111, 69–78.CrossRefGoogle Scholar
Blobel, C.P., Wolfsberg, T.G., Turck, C.W., Myles, D.G., makoff, P. & White, J.M. (1992). A potential fusion peptide and an integrin ligand domain in a protein active in sperm-egg fusion. Nature 356 248–52.CrossRefGoogle Scholar
Cherr, G.N., Lambert, H., Meizel, S. & Katz, D.F. (1986). In vitro studies of the golden hamster sperm acrosome reaction:completion on the zona pellucida and induction by homologous solubilized zonae pellucidae. Dev. Biol. 114, 119–31.CrossRefGoogle ScholarPubMed
Cowan, A.E., Myles, D.G. & Koppel, D.E. (1991). Migration of the guinea pig sperm membrane protein PH-20 from one localized surface domain to another does not occur by a simple diffusion-trapping mechanism. Dev. Biol. 144, 189–98.CrossRefGoogle Scholar
Diaz-Perez, E. & Meizel, S.(1992). Importance of mammalian sperm metalloprotease activity during the acrosome reaction to subsequent sperm-egg fusion: inhibitor studies with human sperm and zona-free hamster eggs. Mol. Reprod. Dev. 31, 122–30.CrossRefGoogle ScholarPubMed
Diaz-Perez, E.,Thomas, P. & Meizel, S. (1988).Evidence suggesting a role for sperm metalloprotease activity in penetration of zona-free hamster eggs by human sperm. J. Exp. Zool. 248, 213–21.CrossRefGoogle ScholarPubMed
Dravland, J.E. & Meizel, S.(1982). The effect of inhibitors of trypsin and phospholipase A2 on the penetration of zona pellucida-free hamster eggs by acrosome-reacted hamster sperm. J. Androl. 3, 388–95.CrossRefGoogle Scholar
Draviand, J.E., Llanos, M., Munn, R. & Meizel, S. (1984). Evidence for the involvement of a sperm trypsin-like enzyme in the membrane events of the hamster sperm acrosome reaction. J. Exp. Zool. 232, 117–28.CrossRefGoogle Scholar
Fleming, A.D.& Yanagimachi, R.(1980). Superovulation and superpregnancy in the golden hamster. Dev. Growth Differ. 22, 103–12.CrossRefGoogle ScholarPubMed
Fraser, L. (1982). p-Aminobenzamidine, an acrosin inhibitor, inhibits mouse sperm penetration of the zona pellucida but not the acrosome reaction. J. Reprod. Fert. 65, 1–11.CrossRefGoogle Scholar
Green, D.P. (1978). The activation of proteolysis in the acrosome reaction of guinea pig sperm. J. Cell Sci. 32, 153–64.CrossRefGoogle ScholarPubMed
Greenwald, G.S. (1962). Analysis of superovulation in the adult hamster. Endocrinology 71, 378–89.CrossRefGoogle ScholarPubMed
Hedrick, J.L., Urch, U.A. & Hardy, D.M. (1989). Structure-functional properties of the sperm enzyme acrosin. In: Biocatalysis in Agricultural Biotechnology, ed. Whitaker, J.R. & Sonnet, P.E., pp. 215–29. Washington, DC: American Chemical Society.CrossRefGoogle Scholar
Huang, T.T.F., Hardy, D., Yanagimachi, H., Tuschor, C., Tung, K., Wild, G. & Yanagimachi, R. (1985). pH and protease control of acrosomal content stasis and release during the guinea pig sperm acrosome reaction. Biol. Reprod. 32, 451–62.CrossRefGoogle ScholarPubMed
Huang-Yang, Y.H.H. & Meizel, S. (1975). Purification of rabbit testis proacrosin and studies of its active form. Biol. Reprod. 12, 232–8.CrossRefGoogle ScholarPubMed
Kopf, G. & Gerton, G.L. (1991). The mammalian sperm acrosome and the acrosome reaction. In: Elements of Mammalian Fertilization, ed.Wassarman, P., pp. 153–203. Boca Raton, Florida: CRC Press.Google Scholar
Leibfried, M.L. & Bavister, B.D. (1981). The effects of taurine and hypotaurine on in vitro fertilization in the golden hamster. Gamete Res. 4, 57–63.CrossRefGoogle Scholar
Liu, W. & Meizel, S. (1979). Further evidence in support of a role of hamster sperm hydrolytic enzymes in the acrosome reaction. J. Exp. Zool. 207, 173–86.Google Scholar
Longo, F. & Yanagimachi, R. (1993). Detection of sperm-egg fusion. In: Methods in Enzymology, ed. Duzgunes, N.New York: Academic Press (in press).Google Scholar
Meizel, S. (1984). The importance of hydrolytic enzymes to an exocytotic event, the mammalian sperm acrosome reaction. Biol. Rev. 59, 125–57.CrossRefGoogle Scholar
Meizel, S. & Mukerji, S.K. (1976). Biochemical studies of proacrosin and acrosin from hamster cauda epididymal spermatozoa. Biol. Reprod. 14, 440–50.CrossRefGoogle ScholarPubMed
Mrsny, R., Waxman, L. & Meizel, S. (1979). Taurine main tains and stimulates motility of hamster sperm during capacitation in vitro. J. Exp. Zool. 210, 123–8.CrossRefGoogle Scholar
Parrish, R.F. & Polakoski, K.L. (1978). Boar alpha-acrosin: purification and characterization of the initial active enzyme resulting from the conversion of boar proacrosin to acrosin. J. Biol. Chem. 253, 8424–32.CrossRefGoogle Scholar
Perreault, S.D., Zerkin, B.R., and Rogers, B.J. (1982).Effect of trypsin inhibitors on acrosome reaction of guinea pig spermatozoa. Biol. Reprod. 26, 343–351.CrossRefGoogle ScholarPubMed
Phelps, B.M., Koppel, D.E., Primakoff, P. & Myles, D.G. (1990). Evidence that proteolysis of the surface is an initial step in the mechanism of formation of sperm cell surface domains. J. Cell Biol. 111, 1839–47.CrossRefGoogle ScholarPubMed
Richardson, R.T., Nikolajczyk, B.S., Abdullah, L.H., Beaver, J.C. & O'Rand, M.G.(1991). Localization of rabbit sperm acrosin during the acrosome reaction induced by immobilized zona matrix. Biol. Reprod. 45, 20–6.CrossRefGoogle ScholarPubMed
Talbot, P. &Chacon, R. (1981).Detection of modification in the tail of capacitated guinea pig sperm using lectins. J. Exp. Zool. 216, 435–44.CrossRefGoogle ScholarPubMed
Tesarik, J., Drahorad, J., Testart, J. & Mendoza, C. (1990).Acrosin activation follows its surface exposure and precedes membrane fusion in human sperm acrosome reaction. Development 110, 391–400.CrossRefGoogle ScholarPubMed
Topfer-Petersen, E., Cedhova, D., Henchen, A., Steinberger, M.M., Fries, A.E. & Zucker, A. (1990).Cell biology of acrosomal proteins. Andrologia 22, 110–21.CrossRefGoogle ScholarPubMed
Urch, U.A. (1991). Biochemistry and function of acrosin. In: Elements of Mammalian Fertilization, ed. Wassarman, P., 1, pp. 233–48. Boca Raton, Florida: CRC Press.Google Scholar
Wolf, D.P. (1977). Involvement of a trypsin-like activity in sperm penetration into zona-free mouse ova. J. Exp. Zool. 199, 149–56.CrossRefGoogle ScholarPubMed
Yanagimachi, R. (1970). The movement of golden hamster spermatozoa before and after capacitation. J. Reprod. Pert. 23, 193–6.CrossRefGoogle ScholarPubMed
Yanagimachi, R. (1978). Calcium requirement for sperm-egg fusion in mammals. Biol. Reprod. 19, 949–58.CrossRefGoogle ScholarPubMed
Yanagimachi, R. (1981). Mechanisms of fertilization in mammals. In: Fertilization and Embryonic Development, ed. Mastroianni, L. & Biggers, J.D., pp. 81–181. New York: Plenum Press.CrossRefGoogle Scholar
Yanagimachi, R.(1982). In vitro sperm capacitation and fertilization of golden hamster eggs in a chemically defined medium. In: In Vitro Fertilization and Embryo Transfer, ed. Hafez, E.S.E. & Semm, K., pp.65–76. Lancaster, MTP Press.Google Scholar
Yanagimachi, R. (1988 a). Mammalian fertilization. In: The Physiology of Reproduction, ed. Knobil, E. & Neill, J.D., pp. 135–85. New York: Raven Press.Google Scholar
Yanagimachi, R. (1988 b). Sperm-egg fusion. In: Current Topics in Membranes and Transport, vol. 32, Membrane Fusion in Fertilization, Cellular Transport and Viral Infection, ed. Duz gunes, N. & Bronner, F., pp. 3–43.New York: Academic Press.Google Scholar
Yanagimachi, R. & Noda, Y.D. (1970 a). Physiological changes in the post-nuclear cap region of mammalian spermatozoa: a necessary preliminary to the membrane fusion between sperm and egg cells. J. Ultrastruct. Res. 31, 486–93.CrossRefGoogle Scholar
Yanagimachi, R. & Noda, Y.D.(1970 b). Electron microscope studies of sperm incorporation into hamster egg. Am. J. Anat. 126, 429–62.CrossRefGoogle Scholar
Zaneveld, L.J.D. & De Jonge, C.J. (1991). Mammalian sperm acrosomal enzymes and the acrosome reaction. In: A Com parative Overview of Mammalian Fertilization, ed. Dunbar, B.S. & O'Rand, M.G., pp. 63–79. New York: Raven Press.CrossRefGoogle Scholar
Zaneveld, L.J.D., Polakoski, K.L., Robertson, R.T. & Williams, W.L. (1975). Trypsin inhibitors and fertilization. In: Proteinose and Biological Control, ed. Fritz, H. & Tschesche, H., pp. 236–42. New York: Walter de Gruyter.Google Scholar