Journal of Andrology, Vol 23, Issue 1 48-63, Copyright © 2002 by The American Society of Andrology

JOURNAL ARTICLE

Localization of the sperm protein SP22 and inhibition of fertility in vivo and in vitro

G. R. Klinefelter, J. E. Welch, S. D. Perreault, H. D. Moore, R. M. Zucker, J. D. Suarez, N. L. Roberts, K. Bobseine and S. Jeffay
US Environmental Protection Agency, Office of Research and Development, National Health and Environmental Effects Research Laboratory, Research Triangle Park, North Carolina 27711, USA. klinefelter.gary@epa.gov

We previously established that levels of the sperm membrane protein, SP22, are highly correlated with the fertility of sperm from the cauda epididymidis of rats exposed to both epididymal and testicular toxicants, and that a testis-specific SP22 transcript is expressed in postmeiotic germ cells. In this study, polyclonal and monoclonal antibodies were generated to study the expression of SP22 in the testis and epididymis, and to determine whether SP22 plays a coincidental or causal role in fertility. Polyclonal antiserum was raised in sheep against full-length recombinant rat SP22 (rSP22). Hybridoma clones were generated from mice immunized with rSP22 and boosted with native SP22; positive clones were used for ascites production. Immunoblots indicated that affinity-purified anti-rSP22 immunoglobulin (Ig) and ascites Ig recognized denatured and native SP22, respectively. Linear epitope mapping of the 189-amino acid SP22 sequence revealed 3 distinct peptide sequences recognized by anti-rSP22 Ig, and 1 sequence recognized by ascites Ig. Cytoplasm of round spermatids and heads of elongating/elongated spermatids immunostained with both anti-rSP22 and ascites antibodies. Isolated rete testis sperm revealed discrete staining over the cytoplasmic droplet, whereas staining was apparent over the equatorial segment of the head by the time sperm reached the caput epididymidis. Clear cells were, interestingly, immunostained along the length of the epididymis. Ascites Ig and anti-SP22 Ig each recognized the equatorial segment of sperm heads from rat, hamster, bull, rabbit, and human. Ascites Ig and affinity-purified anti-rSP22 Ig each significantly inhibited the fertility of cauda epididymal sperm from the rat in vivo, as well as the fertilization rates of cauda epididymal sperm in vitro. Moreover, affinity-purified anti-rSP22 significantly inhibited in vitro fertilization of both zona-intact and zona-free hamster oocytes, suggesting that SP22 may play a role in both the zona penetration and membrane fusion steps of fertilization.

This article has been cited by other articles:

				A. M. Benoit, H. A. LaVoie, G. L. McCoy, and C. A. Blake Expression of Sperm Protein 22 (SP22) in the Rat Ovary During Different Reproductive States Experimental Biology and Medicine, July 1, 2007; 232(7): 910 - 920. [Abstract] [Full Text] [PDF]

				A. M. Benoit, G. L. McCoy, and C. A. Blake Localization of Fertility Factor SP22 to Specific Cell Types Within the Anterior Pituitary Gland Experimental Biology and Medicine, November 1, 2005; 230(10): 721 - 730. [Abstract] [Full Text] [PDF]

				A Cesari, M R Katunar, M A Monclus, A Vincenti, J C de Rosas, and M W Fornes Serine protease activity, bovine sperm protease, 66 kDa (BSp66), is present in hamster sperm and is involved in sperm-zona interaction Reproduction, March 1, 2005; 129(3): 291 - 298. [Abstract] [Full Text] [PDF]

				G. R. Klinefelter, L. F. Strader, J. D. Suarez, N. L. Roberts, J. M. Goldman, and A. S. Murr Continuous Exposure to Dibromoacetic Acid Delays Pubertal Development and Compromises Sperm Quality in the Rat Toxicol. Sci., October 1, 2004; 81(2): 419 - 429. [Abstract] [Full Text] [PDF]

				E. H. Kaydos, J. D. Suarez, N. L. Roberts, K. Bobseine, R. Zucker, J. Laskey, and G. R. Klinefelter Haloacid Induced Alterations in Fertility and the Sperm Biomarker SP22 in the Rat Are Additive: Validation of an ELISA Toxicol. Sci., October 1, 2004; 81(2): 430 - 442. [Abstract] [Full Text] [PDF]

				J. A. Olzmann, K. Brown, K. D. Wilkinson, H. D. Rees, Q. Huai, H. Ke, A. I. Levey, L. Li, and L.-S. Chin Familial Parkinson's Disease-associated L166P Mutation Disrupts DJ-1 Protein Folding and Function J. Biol. Chem., February 27, 2004; 279(9): 8506 - 8515. [Abstract] [Full Text] [PDF]

				D. W. Miller, R. Ahmad, S. Hague, M. J. Baptista, R. Canet-Aviles, C. McLendon, D. M. Carter, P.-P. Zhu, J. Stadler, J. Chandran, et al. L166P Mutant DJ-1, Causative for Recessive Parkinson's Disease, Is Degraded through the Ubiquitin-Proteasome System J. Biol. Chem., September 19, 2003; 278(38): 36588 - 36595. [Abstract] [Full Text] [PDF]

				K. Honbou, N. N. Suzuki, M. Horiuchi, T. Niki, T. Taira, H. Ariga, and F. Inagaki The Crystal Structure of DJ-1, a Protein Related to Male Fertility and Parkinson's Disease J. Biol. Chem., August 15, 2003; 278(33): 31380 - 31384. [Abstract] [Full Text] [PDF]

				C. Bohring and W. Krause Immune infertility: towards a better understanding of sperm (auto)-immunity: The value of proteomic analysis Hum. Reprod., May 1, 2003; 18(5): 915 - 924. [Abstract] [Full Text] [PDF]

				T. Niki, K. Takahashi-Niki, T. Taira, S. M.M. Iguchi-Ariga, and H. Ariga DJBP: A Novel DJ-1-Binding Protein, Negatively Regulates the Androgen Receptor by Recruiting Histone Deacetylase Complex, and DJ-1 Antagonizes This Inhibition by Abrogation of This Complex Mol. Cancer Res., February 1, 2003; 1(4): 247 - 261. [Abstract] [Full Text] [PDF]

				G. R. Klinefelter, L. F. Strader, J. D. Suarez, and N. L. Roberts Bromochloroacetic Acid Exerts Qualitative Effects on Rat Sperm: Implications for a Novel Biomarker Toxicol. Sci., July 1, 2002; 68(1): 164 - 173. [Abstract] [Full Text] [PDF]