SNARE proteins and caveolin-1 in stallion spermatozoa: possible implications for fertility

Abstract

Proteins implicated in the "SNARE hypothesis" for membrane fusion have been characterized in the acrosome of several mammalian species, and a functional role for these proteins during the acrosome reaction has been proposed. We have investigated the presence of SNAREs in equine sperm, using semen samples obtained from stallions with varying fertility. Immunocytochemical analysis revealed that members of different SNARE families can be detected on the acrosome of equine sperm, notably in the acrosomal cap and equatorial segment. These proteins include the t-SNARE syntaxin, the v-SNARE synaptobrevin/VAMP, the calcium sensor synaptotagmin, and the ATPase NSF. Also present is caveolin-1, a component of lipid rafts. Stallions with fertility problems presented the worst quality of sperm and acrosomal membrane, and had less sperm cells stained positively for SNAREs and caveolin-1, than sperm from fertile donors (p < 0.001). Ubiquitin surface staining was also performed and it seemed to inversely correlate with stallion fertility, supporting data obtained with the negative staining technique. A male-related problem was confirmed when mares that had failed to impregnate with samples from an infertile stallion were successfully inseminated with sperm from a fertile donor. Furthermore NSF, synaptotagmin and caveolin-1 staining seemed to be useful in predicting stallion fertility, i.e. significantly more sperm cells stained positively for these proteins in samples from fertile males. Although these results need to be expanded on a larger scale, they suggest that acrosomal and surface staining of equine sperm with novel probes may constitute useful tools in predicting stallion fertility.

Remodelling of Lipid Rafts during In vitro Capacitation and Acrosome Reaction of Ram Spermatozoa

“…Caveolin-1 was located at the acrosomal region, although not all sperm were stained. These results corroborate our previous observations about the heterogeneity of the sperm sample obtained by swim-up [51,52], and it is also consistent with those reported for stallion semen in which a relationship between a lack of positive caveolin-1 staining and a reduction in fertility was found [50]. Our results also showed an increase in the proportion of stained sperm at the acrosome, occasionally including the equatorial region, after capacitation.…”

“…Caveolin-1 is a cholesterol-binding protein [46], and a key raft organizer that has been identified in the acrosomal region of the mouse [48,49], guinea pig [49], stallion [50]and boar [30] spermatozoa. We have identified, by immunocytochemical analysis, caveolin-1 in fresh ram spermatozoa freed from seminal plasma by swim-up.…”

Implication of transcriptome profiling of spermatozoa for stallion fertility

“…Furthermore, Giesecke et al (2011) chose three genes, namely angiotensin-converting enzyme (ACE), sperm autoantigenic protein 17 (SPA17) and FSH beta subunit (FSHB), to test as candidates for determining Hanoverian stallion fertility, finding that haplotypes of all three genes significantly contributed to the paternal and embryonic fertility components of the pregnancy rate per oestrus. In addition, Gamboa and Ramalho-Santos (2005) investigated the presence of soluble N-ethylmaleimide-sensitive factor activating protein receptor (SNARE) proteins in equine spermatozoa and found that, stallions with fertility problems had the worst sperm and acrosomal membrane quality, as well as fewer sperm cells that stained positive for SNAREs and caveolin1 compared with spermatozoa from fertile donors. In the present study, we used the microarray approach to identify molecular pathways relevant to stallion fertility.…”

Membrane Dynamics of Spermatozoa during Capacitation: New Insight in Germ Cells Signalling

“…Botto et al (2010) demonstrated that the amount of Cav-1 significantly increases in the insoluble membrane fraction in spermatozoa incubated in vitro under capacitating conditions (from 35 and 20% to 60 and 70%, respectively) when compared with freshly ejaculated sperm cells [34]. More recently, Baltiérrez-Hoyos et al confirmed the idea that CAV-1 could be involved in biochemical machinery that controls capacitation and the AR.…”

“…Interestingly, in stallion spermatozoa, it has been found that SNARE protein colocalizes with Cav-1 and that fertility seems to be related with the percentage spermatozoa immunopositive for synaptotagmin (a calcium sensor), NSF (a SNARE complex disassembler) and caveolin-1 (a signalling pathways organizer) [34].…”