Identification of osteopontin (OPN) mRNA and protein in the rat testis and epididymis, and on sperm.

Abstract

We have identified a bone cell adhesion molecule, osteopontin, in the rat testis and epididymis by Northern analysis, RT-PCR, Western immunoblot analysis and immunocytochemistry. A polyclonal antibody raised against rat epididymal fluid proteins was used to detect fusion proteins produced by a testis lambda gt11 cDNA library. Sequence analysis of one of four positive cDNA clones, designated as pREP5, revealed identity with the rat osteopontin (OPN) cDNA. The partial cDNA clone pREP5 encompasses 64% of the 1,457 residues reported by Oldberg et al. (1986; Proc Natl Acad Sci USA 83:8819-8823). Immunoblot analysis with a monoclonal antibody against OPN detects the presence of immunoreactive polypeptides in rat testis homogenates as well as in epididymal fluid and sperm extracts. Immunocytochemical localization to the basal and adluminal region of the seminiferous tubule suggests that OPN could be a Sertoli cell product. Indeed, Northern blot analysis of testicular cell preparations demonstrated positive hybridization to Sertoli cell-enriched RNA, but not to RNA isolated from interstitial cell preparations or to isolated germ cell RNA preparations. OPN is also detected in the rat epididymis and on epididymal spermatozoa. This is the first report on the presence of OPN mRNA and protein in rat testis and epididymis and on the presence of OPN on the surface of epididymal spermatozoa. The characterization of this protein in other tissue suggests that OPN could play a role in testicular cell adhesion during spermatogenesis and/or epididymal maturation, although other potential functions in the male reproductive tract are discussed.