Demonstration of a Glycoprotein Derived From the Ceacam10 Gene in Mouse Seminal Vesicle Secretions1

Abstract

CEACAM10 was purified from mouse seminal vesicle secretions by a series of purification steps that included ion exchange chromatography on a DEAE-Sephacel column and ion exchange high-performance liquid chromatography on a sulfopropyl column. It was shown to be a 36-kDa glycoprotein with an N-linked carbohydrate moiety. The circular dichromoism spectrum of CEACAM10 in 50 mM phosphate buffer at pH 7.4 appeared as one negative band arising from the beta form at 217 nm. CEACAM10 was expressed predominantly in seminal vesicles of adult mice. Both CEACAM10 and its mRNA were demonstrated on the luminal epithelium of the mucosal folds in the seminal vesicle. The amount of Ceacam10 mRNA in the seminal vesicle was correlated with the stage of animal maturation. Castration of adult mice resulted in cessation of Ceacam10 expression, while treatment of castrated mice with testosterone propionate in corn oil restored Ceacam10 expression in the seminal vesicle. During the entire course of pregnancy, Ceacam10 might be silent in the embryo. A cytochemical study illustrated the presence of the CEACAM10 binding region on the entire surface of mouse sperm. CEACAM10-sperm binding greatly enhanced sperm motility in vitro.

SPINKL, a Kazal-type serine protease inhibitor-like protein purified from mouse seminal vesicle fluid, is able to inhibit sperm capacitation

“…Many gene expressions in this tissue have been demonstrated to be regulated by androgen (Chen et al 1987, 1998, Yu et al 1993, Li et al 2005. In this study, regulation of the Spinkl gene was also the case (Fig.…”

“…They were housed under controlled lighting (14h light:10h darkness) at 21-22 8C and were provided with water and NIH-31 laboratory chow ad libitum. To study the developmental profile and tissue distribution of the protein, we followed our previously described methods (Li et al 2005(Li et al , 2006. To investigate the androgenic effects, male mice were castrated and administrated testosterone as previously described (Li et al 2005(Li et al , 2006.…”

“…After centrifugation at 10 000 g for 15 min, the supernatant was resolved by ionexchange chromatography on a diethylaminoethyl (DEAE)-Sephacel (Amersham Pharmacia Biotech, Uppsala, Sweden) column (12!2.6 cm) into four fractions (Fig. 1A), as described previously (Li et al 2005(Li et al , 2006. Fraction IV collected from three separations was concentrated and subjected to a Sephadex G-75 (Sigma Chemical Co.) column (2.6!120 cm) pre-equilibrated with 50 mmol/l Tris-HCl and 150 mmol/l NaCl (pH 7.4).…”

“…Immunostaining was performed according to our previously described methods (Li et al 2005(Li et al , 2006. In brief, tissues slides were blocked with 10% (v/v) normal goat serum in PBS (blocking solution) for 1 h at room temperature and then incubated with anti-SPINKL antiserum diluted 1:1000 in the blocking solution at 4 8C for 16 h. After washing, the slides were treated with biotin-conjugated goat anti-rabbit IgG (w1 mg/ml; Kirkegaard & Perry Laboratories, Gaithersburg, MD, USA) in the blocking solution for 1 h at room temperature.…”

“…Thus, the components of SVSs may affect sperm function. In fact, several proteins purified from murine SVSs have been demonstrated to affect sperm physiology (Chen et al 1998, Huang et al 1999, Luo et al 2001, Li et al 2005. However, other components of SVSs remain to be identified.…”