Este artículo es de Acceso Abierto y puede ser descargado en su versión final desde nuestro repositorio
Consulte el artículo en la página del editor
Consulte la política de Acceso Abierto del editor


Mammalian fertilization is a complex multi-step process mediated by different molecules present on both gametes. Epididymal protein CRISP1, a member of the Cysteine-RIch Secretory Protein (CRISP) family, was identified by our laboratory and postulated to participate in both sperm-zona pellucida (ZP) interaction and gamete fusion by binding to egg-complementary sites. To elucidate the functional role of CRISP1 in vivo, we disrupted the Crisp1 gene and evaluated the effect on animal fertility and several sperm parameters. Male and female Crisp1-/- animals exhibited no differences in fertility compared to controls. Sperm motility and the ability to undergo a spontaneous or progesterone-induced acrosome reaction were neither affected in Crisp1-/- mice. However, the level of protein tyrosine phosphorylation during capacitation was clearly lower in mutant sperm than in controls. In vitro fertilization assays showed that Crisp1-/- sperm also exhibited a significantly reduced ability to penetrate both ZP-intact and ZP-free eggs. Moreover, when ZP-free eggs were simultaneously inseminated with Crisp1+/+ and Crisp1-/- sperm in a competition assay, the mutant sperm exhibited a greater disadvantage in their fusion ability. Finally, the finding that the fusion ability of Crisp1-/- sperm was further inhibited by the presence of CRISP1 or CRISP2 during gamete co-incubation, supports that another CRISP cooperates with CRISP1 during fertilization and might compensate for its lack in the mutant mice. Together, these results indicate that CRISP proteins are players in the mammalian fertilization process. To our knowledge this is the first knockout mice generated for a CRISP protein. The information obtained might have important functional implications for other members of the widely distributed and evolutionarily conserved CRISP family. © 2008 Elsevier Inc.


Documento: Artículo
Título:Impaired sperm fertilizing ability in mice lacking Cysteine-RIch Secretory Protein 1 (CRISP1)
Autor:Da Ros, V.G.; Maldera, J.A.; Willis, W.D.; Cohen, D.J.; Goulding, E.H.; Gelman, D.M.; Rubinstein, M.; Eddy, E.M.; Cuasnicu, P.S.
Filiación:Instituto de Biología y Medicina Experimental (IBYME-CONICET), Vuelta de Obligado 2490, Buenos Aires, 1428, Argentina
Gamete Biology Section, Laboratory of Reproductive and Developmental Toxicology, National Institute of Environmental Health Sciences, Research Triangle Park, NC 27709, United States
Instituto de Investigaciones en Ingenieria Genetica y Biologia Molecular (INGEBI-CONICET), Buenos Aires, 1428, Argentina
Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Buenos Aires, 1428, Argentina
Palabras clave:CRISP; Egg; Fertilization; Sperm; cysteine rich secretory protein 1; DNA; RNA; secretory protein; tyrosine; acrosome reaction; animal cell; animal experiment; animal tissue; article; controlled study; female; fertility; fertilization; fertilization in vitro; gene disruption; genotype; immunoblotting; male; mouse; mutant; nonhuman; priority journal; protein phosphorylation; reverse transcription polymerase chain reaction; sperm; spermatozoon capacitation; spermatozoon motility; zona pellucida; Animalia; Mammalia; Mus
Página de inicio:12
Página de fin:18
Título revista:Developmental Biology
Título revista abreviado:Dev. Biol.
CAS:DNA, 9007-49-2; RNA, 63231-63-0; tyrosine, 16870-43-2, 55520-40-6, 60-18-4


  • Baba, D., Kashiwabara, S., Honda, A., Yamagata, K., Wu, Q., Ikawa, M., Okabe, M., Baba, T., Mouse sperm lacking cell surface hyaluronidase PH-20 can pass through the layer of cumulus cells and fertilize the egg (2002) J. Biol. Chem., 277, pp. 30310-30314
  • Berditchevski, F., Gilbert, E., Griffiths, M.R., Fitter, S., Ashman, L., Jenner, S.J., Analysis of the CD151-alpha3beta1 integrin and CD151-tetraspanin interactions by mutagenesis (2001) J. Biol. Chem., 276, pp. 41165-41174
  • Busso, D., Cohen, D.J., Maldera, J.A., Dematteis, A., Cuasnicu, P.S., A novel function for CRISP1 in rodent fertilization: involvement in sperm-zona pellucida interaction (2007) Biol. Reprod., 77, pp. 848-854
  • Busso, D., Goldweic, N.M., Hayashi, M., Kasahara, M., Cuasnicu, P.S., Evidence for the involvement of testicular protein CRISP2 in mouse sperm-egg fusion (2007) Biol. Reprod., 76, pp. 701-708
  • Cameo, M.S., Blaquier, J.A., Androgen-controlled specific proteins in rat epididymis (1976) J. Endocrinol., 69, pp. 317-324
  • Cohen, D.J., Ellerman, D.A., Cuasnicú, P.S., Mammalian sperm-egg fusion: evidence that epididymal protein DE plays a role in mouse gamete fusion (2000) Biol. Reprod., 63, pp. 462-468
  • Cohen, D.J., Rochwerger, L., Ellerman, D.A., Morgenfeld, M., Busso, D., Cuasnicu, P.S., Relationship between the association of rat epididymal protein DE with spermatozoa and the behavior and function of the protein (2000) Mol. Reprod. Dev., 56, pp. 180-188
  • Cohen, D.J., Ellerman, D.A., Busso, D., Morgenfeld, M., Piazza, A., Hayashi, M., Young, E.T., Cuasnicu, P.S., Evidence that human epididymal protein ARP plays a role in gamete fusion through complementary sites on the surface of the human egg (2001) Biol. Reprod., 65, pp. 1000-1005
  • Cohen, D.J., Da Ros, V.G., Busso, D., Ellerman, D.A., Maldera, J.A., Goldweic, N., Cuasnicu, P.S., Participation of epididymal cysteine-rich secretory proteins in sperm-egg fusion and their potential use for male fertility regulation (2007) Asian J. Androl., 9, pp. 528-532
  • Da Ros, V.G., Munuce, M.J., Cohen, D.J., Marin-Briggiler, C.I., Busso, D., Visconti, P.E., Cuasnicu, P.S., Bicarbonate is required for migration of sperm epididymal protein DE (CRISP-1) to the equatorial segment and expression of rat sperm fusion ability (2004) Biol. Reprod., 70, pp. 1325-1332
  • Drabent, B., Saftig, P., Bode, C., Doenecke, D., Spermatogenesis proceeds normally in mice without linker histone H1t (2000) Histochem. Cell Biol., 113, pp. 433-442
  • Ellerman, D.A., Cohen, D.J., Da Ros, V.G., Morgenfeld, M.M., Busso, D., Cuasnicu, P.S., Sperm protein "DE" mediates gamete fusion through an evolutionarily conserved site of the CRISP family (2006) Dev. Biol., 297, pp. 228-237
  • Fraser, L.R., Drury, L.M., The relationship between sperm concentration and fertilization In vitro of mouse eggs (1975) Biol. Reprod., 13, pp. 513-518
  • Garberi, J.C., Kohane, A.C., Cameo, M.S., Blaquier, J.A., Isolation and characterization of specific rat epididymal proteins (1979) Mol. Cell. Endocrinol., 13, pp. 73-82
  • Garberi, J.C., Fontana, J.D., Blaquier, J.A., Carbohydrate composition of specific rat epididymal protein (1982) Int. J. Androl., 5, pp. 619-626
  • Guo, M., Teng, M., Niu, L., Liu, Q., Huang, Q., Hao, Q., Crystal structure of the cysteine-rich secretory protein stecrisp reveals that the cysteine-rich domain has a K+ channel inhibitor-like fold (2005) J. Biol. Chem., 280, pp. 12405-12412
  • Hayashi, M., Fujimoto, S., Takano, H., Ushiki, T., Abe, K., Ishikura, H., Yoshida, M., Kasahara, M., Characterization of a human glycoprotein with potential role in sperm-egg fusion: cDNA cloning, immunohistochemical localization, and chromosomal assignment of the gene (AEGL1) (1996) Genomics, 32, pp. 367-374
  • Jalkanen, J., Huhtaniemi, I., Poutanen, M., Mouse cysteine-rich secretory protein 4 (CRISP4): a member of the crisp family exclusively expressed in the epididymis in an androgen-dependent manner (2005) Biol. Reprod., 72, pp. 1268-1274
  • Kjeldsen, L., Cowland, J.B., Johnsen, A.H., Borregaard, N., SGP28, a novel matrix glycoprotein in specific granules of human neutrophils with similarity to a human testis-specific gene product and a rodent sperm-coating glycoprotein (1996) FEBS Lett., 380, pp. 246-250
  • Kohane, A.C., Gonzalez Echeverria, F., Piñeiro, L., Blaquier, J.A., Interaction of proteins of epididymal origin with spermatozoa (1980) Biol. Reprod., 23, pp. 737-742
  • Kohane, A.C., Piñeiro, L., Blaquier, J.A., Androgen-controlled synthesis of specific protein in the rat epididymis (1983) Endocrinology, 112, pp. 1590-1596
  • Laemmli, U.K., Cleavage of structural proteins during the assembly of the head of bacteriophage T4 (1970) Nature, 227, pp. 680-685
  • Lu, Q.X., Shur, B.D., Sperm from β1,4-galactosyltransferase-null mice are refractory to ZP3-induced acrosome reactions and penetrate the zona pellucida poorly (1997) Development, 124, pp. 4121-4131
  • Nicolson, G., Yanagimachi, H., Yanagimachi, R., Ultrastructural localization of lectin-binding sites on the zonae pellucidae and plasma membrane of mammalian egg (1975) J. Cell Biol., 66, pp. 263-273
  • Nolan, M.A., Babcock, D.F., Wennemuth, G., Brown, W., Burton, K.A., McKnight, G.S., Sperm-specific protein kinase A catalytic subunit Cα2 orchestrates cAMP signaling for male fertility (2004) Proc. Natl. Acad. Sci. U. S. A., 101, pp. 13483-13488
  • Nolan, M.A., Wu, L., Bang, H.J., Jelinsky, S.A., Roberts, K.P., Turner, T.T., Kopf, G.S., Johnston, D.S., Identification of rat cysteine-rich secretory protein 4 (Crisp4) as the ortholog to human CRISP1 and mouse Crisp4 (2006) Biol. Reprod., 74, pp. 984-991
  • Okabe, M., Cummins, J.M., Mechanisms of sperm-egg interactions emerging from gene-manipulated animals (2007) Cell. Mol. Life Sci., 64, pp. 1945-1958
  • Roberts, K.P., Wamstad, J.A., Ensrud, K.M., Hamilton, D.W., Inhibition of capacitation-associated tyrosine phosphorylation signaling in rat sperm by epididymal protein crisp-1 (2003) Biol. Reprod., 69, pp. 572-581
  • Roberts, K.P., Ensrud, K.M., Wooters, J.L., Nolan, M.A., Johnston, D.S., Hamilton, D.W., Epididymal secreted protein Crisp-1 and sperm function (2006) Mol. Cell. Endocrinol., 250, pp. 122-127
  • Rochwerger, L., Cuasnicu, P.S., Redistribution of a rat sperm epididymal glycoprotein after In vivo and in vitro capacitation (1992) Mol. Reprod. Dev., 31, pp. 34-41
  • Rochwerger, L., Cohen, D.J., Cuasnicú, P.S., Mammalian sperm-egg fusion: the rat egg has complementary sites for a sperm protein that mediates gamete fusion (1992) Dev. Biol., 153, pp. 83-90
  • Rubinstein, E., Ziyyat, A., Prenant, M., Wrobel, E., Wolf, J.P., Levy, S., Le Naour, F., Boucheix, C., Reduced fertility of female mice lacking CD81 (2006) Dev. Biol., 290, pp. 351-358
  • Schwidetzky, U., Schleuning, W.D., Haendler, B., Isolation and characterization of the androgen-dependent mouse cysteine-rich secretory protein-1 (CRISP-1) gene (1997) Biochem. J., 321, pp. 325-332
  • Stein, K.K., Go, J.C., Primakoff, P., Myles, D.G., Defects in secretory pathway trafficking during sperm development in Adam2 knockout mice (2005) Biol. Reprod., 73, pp. 1032-1038
  • Towbin, H., Staehelin, T., Gordon, J., Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications (1979) Proc. Natl. Acad. Sci. U. S. A., 76, pp. 4350-4360
  • Udby, L., Bjartell, A., Malm, J., Egesten, A., Lundwall, A., Cowland, J.B., Borregaard, N., Kjeldsen, L., Characterization and localization of cysteine-rich secretory protein 3 (CRISP-3) in the human male reproductive tract (2005) J. Androl., 26, pp. 333-342
  • Wright, M.D., Geary, S.M., Fitter, S., Moseley, G.W., Lau, L.M., Sheng, K.C., Apostolopoulos, V., Ashman, L.K., Characterization of mice lacking the tetraspanin superfamily member CD151 (2004) Mol. Cell. Biol., 24, pp. 5978-5988
  • Xie, F., Garcia, M.A., Carlson, A.E., Schuh, S.M., Babcock, D.F., Jaiswal, B.S., Gossen, J.A., Conti, M., Soluble adenylyl cyclase (sAC) is indispensable for sperm function and fertilization (2006) Dev. Biol., 296, pp. 353-362
  • Yamazaki, Y., Morita, T., Structure and function of snake venom cysteine-rich secretory proteins (2004) Toxicon, 44, pp. 227-231
  • Yanagimachi, R., Mammalian fertilization (1994) The Physiology of Reproduction, pp. 189-317. , Knobil E., and Neill J.D. (Eds), Raven Press, New York


---------- APA ----------
Da Ros, V.G., Maldera, J.A., Willis, W.D., Cohen, D.J., Goulding, E.H., Gelman, D.M., Rubinstein, M.,..., Cuasnicu, P.S. (2008) . Impaired sperm fertilizing ability in mice lacking Cysteine-RIch Secretory Protein 1 (CRISP1). Developmental Biology, 320(1), 12-18.
---------- CHICAGO ----------
Da Ros, V.G., Maldera, J.A., Willis, W.D., Cohen, D.J., Goulding, E.H., Gelman, D.M., et al. "Impaired sperm fertilizing ability in mice lacking Cysteine-RIch Secretory Protein 1 (CRISP1)" . Developmental Biology 320, no. 1 (2008) : 12-18.
---------- MLA ----------
Da Ros, V.G., Maldera, J.A., Willis, W.D., Cohen, D.J., Goulding, E.H., Gelman, D.M., et al. "Impaired sperm fertilizing ability in mice lacking Cysteine-RIch Secretory Protein 1 (CRISP1)" . Developmental Biology, vol. 320, no. 1, 2008, pp. 12-18.
---------- VANCOUVER ----------
Da Ros, V.G., Maldera, J.A., Willis, W.D., Cohen, D.J., Goulding, E.H., Gelman, D.M., et al. Impaired sperm fertilizing ability in mice lacking Cysteine-RIch Secretory Protein 1 (CRISP1). Dev. Biol. 2008;320(1):12-18.