Characterization and Localization of Cysteine‐Rich Secretory Protein 3 (CRISP‐3) in the Human Male Reproductive Tract

Abstract

Mammalian members of the cysteine-rich secretory protein (CRISP) family are expressed predominantly in the male reproductive tract and are implicated in the process of reproduction from spermiogenesis, posttesticular sperm maturation, and capacitation to oocyte-sperm fusion, and possibly also penetration of the zona pellucida. Rodents express only 2 CRISPs (CRISP-1 and CRISP-2) in their male reproductive system, whereas humans and horses express an additional third member named CRISP-3. We have previously demonstrated that this protein is present in human seminal plasma as well as in other exocrine secretions, in blood plasma, and in neutrophilic granulocytes. To characterize the protein in seminal plasma and localize the production of CRISP-3 in the human male reproductive tract, we performed immunoblotting and enzyme-linked immunosorbent assay measurements of seminal plasma and immunohistochemistry and in situ hybridization of tissue specimens. We were able to show that human CRISP-3 is a quantitatively minor seminal plasma protein not associated with prostasomes. Furthermore, CRISP-3 expression was found in the secretory epithelium throughout the male genital tract, with particularly high expression in the cauda epididymis and ampulla vas deferens. Examination of seminal plasma from vasectomized males indicates that organs downstream of the epididymis are probably the major sources of seminal plasma CRISP-3.

Low salivary dehydroepiandrosterone and androgen‐regulated cysteine‐rich secretory protein 3 levels in Sjögren's syndrome

“…Hidden antigenic epitopes were revealed after pretreatment with Buffer for Antigen Retrieval (TechMate; Dako, Glostrup, Denmark) using a microwave oven twice for 5 minutes each at 700W. Slides were installed in (27) was inserted into pBluescript II KS (Stratagene, La Jolla, CA) and, after linearization with Xho I and Spe I, was used as a template for in vitro transcription to generate 35 S-labeled antisense and sense probes, respectively. After transcription, the RNA probes were ultrafiltrated (Micron 100; Amicon, Beverly, MA) before hybridization.…”

Cysteine‐rich secretory proteins are not exclusively expressed in the male reproductive tract

“…Crisp3 is regulated by androgens in mice, but to a lesser extent in humans (Haendler et al, 1993;Udby et al, 2005). Crisp3 was initially identified in the mouse salivary gland (Haendler et al, 1993) and later in the human as SGP28 (Kjeldsen et al, 1996).…”

β-Microseminoprotein binds CRISP-3 in human seminal plasma

“…A minor proportion was found in the fractions with CRISP-3, corresponding to the complex. This was expected, as the concentration of MSP in seminal plasma (600-900 mg/L) is >20-fold higher than the concentration of CRISP-3 (3-30 mg/L) [7,15].…”

“…We have recently studied CRISP-3 in the human male reproductive tract, and found mRNA and protein expression throughout the secretory epithelium from the epididymis to the prostate gland [15]. In that work, it was erroneously assumed that CRISP-3 is present in a noncomplexed form in seminal plasma as gel filtration suggested a size close to its monomeric form.…”

“…A minor proportion was found in the fractions with CRISP-3, corresponding to the complex. This was expected, as the concentration of MSP in seminal plasma (600-900 mg/L) is >20-fold higher than the concentration of CRISP-3 (3-30 mg/L) [7,15].To evaluate the capacity of CRISP-3 binding, a small sample of seminal plasma was supplemented with a slight molar excess of pure CRISP-3, i.e., a final concentration of 2 mg CRISP-3 per milliliter seminal plasma, and evaluated by gel filtration. Compared to the gel filtration of normal seminal plasma, a new peak exactly corresponding to the complex formed with the pure proteins was observed in the UV-chromatogram (Fig.…”