Chemical Characterization of the Predominant Proteins Secreted by Mouse Seminal Vesicles

Abstract

Mouse seminal vesicles secrete four major protein components with estimated molecular masses of 95, 38, 17, and 16 kDa. Amino acid sequencing revealed that the 95-kDa component represents a protein with an unknown structure, while the 38-kDa component was identified as semenoclotin, the 17-kDa component as seminal-vesicle-secreted protein IV, and the 16-kDa component as seminal-vesicle-secreted protein V. Semenoclotin and the 95-kDa component were readily cross-linked by transglutaminase, suggesting that the two proteins are involved in the formation of the mouse copulatory plug. Treatment of mouse seminal vesicle fluid with human prostate-specific antigen rapidly degraded semenoclotin, indicating a structural resemblance of this protein to human semenogelins, despite the vast difference in primary structure. As previously reported for other seminal-vesicle-secreted proteins, the semenoclotin transcripts are shown to be under androgen control.Keywords: seminal vesicle ; semen ; androgen; transglutaminase ; coagulation.Mammalian semen is formed by the mixing of spermatozoarich fluid from epididymis with secretions from the accessory sex glands during ejaculation, that mainly consists of fluids from the seminal vesicles and the prostate (Mann and Lutwark-Mann, 1981). In most species, the major contribution is provided by the seminal vesicles, amounting to more than half of the semen volume. The newly ejaculated sperm is highly viscous or gellike, but within a short time it will either liquefy, as in man, or transform into a firm coagulum, yielding a copulatory plug through the action of a prostate-secreted transglutaminase, as seen in rodents.The proteins responsible for gel and clot formation are present at very high concentrations in the seminal vesicle fluid. Somewhat unexpectedly, the primary structure of these proteins are highly dissimilar between species (Lundwall and Lazure, 1995; Lundwall and Ulvsback, 1996). This is caused by a relatively recent evolution of an ancestral gene, involving duplications, exon expansion, and varying selection of splice sites. The genes constitute the REST gene family (abbreviation for rapidly evolving substrates for transglutaminase) as most members seem to encode substrates for transglutaminase (Lundwall, 1996 : Lundwall and Ulvsback, 1996).The rat seminal vesicle secretion consists of five predominant protein components, designated seminal-vesicle-secreted proteins I-V (SVS I-V) in order of size (Ostrowski et al., 1979). The SVS IV and V components are also referred to as S and F proteins by some authors (Higgins et al., 1976). The strucCorrespondence to A. Lundwall,

Identification of prostatic-secreted proteins in mice by mass spectrometric analysis and evaluation of lobe-specific and androgen-dependent mRNA expression

“…The results of mass spectrometric identification of SV proteins were generally in agreement with previous reports, i.e. SVS2, 4, 5, and 6, as well as SPI-KT3 (Lai et al 1991, Lundwall et al 1997. Except SPI-KT3, SV proteins are specifically expressed in the SV and not in the prostate gland, which differs from the rat case, where SVS2, for instance, is highly expressed in DLP/AP.…”

Unexpected Virilization in Male Mice Lacking Steroid 5α-Reductase Enzymes

“…For example, inhibition of 5α-reductase in man causes a decrease in prostatic DHT and a corresponding increase in T (39), as seen in 5α-reductase knockout mice. Treatment of male rats with 5α-reductase inhibitors causes a decrease in copulatory plug formation (26,27), and the same phenomenon is observed in the knockout mice together with a decrease in the mRNA encoding transglutaminase, an enzyme associated with plug formation (40). These similarities suggest that the phenotypes of the knockout mice may accurately reflect the endocrine roles of 5α-reductase and DHT in this species.…”

Genetic and phenotypic influences on copulatory plug survival in mice

“…Male mate choice (Dewsbury, 1982;Drickamer et al, 2003;Edward and Chapman, 2011;Ramm and Stockley, 2014) and the dynamic adjustment of ejaculate allocation (Wedell et al, 2002;Delbarco-Trillo and Ferkin, 2004) suggest that ejaculates are costly to produce and conserved when possible. Plug-forming proteins account for nearly one third of the total protein abundance of the ejaculate in mice, suggesting that this structure is a major reproductive investment for males (Lundwall et al, 1997;Lin et al, 2002;Dean et al, 2011). As large plugs also seem to survive shorter periods of time, and ejaculates are likely to be costly, our study begs the question of why males would ever invest in large plugs.…”

“…The full exomes will be published as part of a larger study elsewhere, but we focus here on these five genes as they are either necessary for copulatory plug formation or present in seminal vesicles and plugs at high abundance (Lundwall et al, 1997;Dean et al, , 2011Dean, 2013;Kawano et al, 2014). DNA was sheared using a Bioruptor UCD-200 (Diagenode, Denville, NJ, USA) with 7 rounds of sonication (7 min per round on high, 30 s on and 30 s off) and genomic DNA libraries were constructed using a previously described protocol designed to facilitate multiplexed exome capture (Rohland and Reich, 2012).…”

“…Coomassie-stained polyacrylamide gels of seminal vesicle fluids revealed four very abundant proteins (Supplementary Figure 3 and Table 2). We did not attempt to identify these four bands using mass spectrometry, but based on previous studies (Lundwall et al, 1997;Lin et al, 2002;Dean et al, 2011;Tseng et al, 2011) and the match to known molecular mass, these four bands most likely consisted of the seminal vesicle-secreted proteins Svs1, Svs2, Svs4 and Svs5 (Supplementary Figure 3). The exact identity of the proteins is not critical as we are only using them as a biomarker of plug composition.…”