Mouse Epididymal Spam1 (PH‐20) Is Released in the Luminal Fluid With its Lipid Anchor

Cumulus cell layers of expanded cumulus oocyte complexes (COCs) are interlinked with networks of hyaluronic acid, chondroitin sulfate B proteoglycans and link proteins, and they can be dispersed by sperm surface hyaluronidases. In this report, we showed that arylsulfatase A (AS-A), existing on the sperm head surface, also had this dispersion action. Purified AS-A free of protease, hyaluronidase and chondroitinase activities could disperse the cumulus matrix of expanded COCs. However, this COC dispersion action was not associated with AS-A desulfation activity, assayed by using p-nitrocatecholsulfate (artificial substrate). COCs incubated for 1 h with sperm pretreated with anti-AS-A IgG in the presence of apigenin (a hyaluronidase inhibitor) did not exhibit matrix dispersion, whereas several cumulus layers were already dispersed in COCs incubated with sperm pretreated with preimmune IgG. Furthermore, sperm from AS-A null mice showed a significant delay in COC dispersion, compared with wild-type sperm. Within 1 h of sperm-COC co-incubation, the size of COCs incubated with AS-A null sperm was 65% of the original dimension, whereas that of COCs inseminated with wild-type sperm was only 17%. A further delay in COC dispersion by AS-A(-/-) mouse sperm was observed when apigenin was present in the co-incubation. We also showed for the first time that AS-A had a specific affinity for chondroitin sulfate B, a component of cumulus matrix proteoglycan networks; this might provide a mechanism of cumulus matrix destabilization induced by sperm surface AS-A.

Sperm surface arylsulfatase A can disperse the cumulus matrix of cumulus oocyte complexes

Wu¹,

Anupriwan²,

Iamsaard³

et al. 2007

Proteomic and functional analysis of human sperm detergent resistant membranes

Nixon

Mitchell

Anderson

et al. 2011

Mammalian spermatozoa attain the ability to fertilize an oocyte as they negotiate the female reproductive tract. This acquisition of functional competence is preceded by an intricate cascade of biochemical and functional changes collectively known as "capacitation." Among the universal correlates of the capacitation process is a remarkable remodeling of the lipid and protein architecture of the sperm plasma membrane. While the mechanisms that underpin this dynamic reorganization remain enigmatic, emerging evidence has raised the prospect that it may be coordinated, in part, by specialized membrane microdomains, or rafts. In the present study we have demonstrated that human spermatozoa express recognized markers of membrane rafts. Further, upon depletion of membrane cholesterol through either physiological (capacitation) or pharmacological (methyl-β-cyclodextrin) intervention, these membrane rafts appear to undergo a polarized redistribution to the peri-acrosomal region of the sperm head. This finding encourages speculation that membrane rafts represent platforms for the organization of proteins involved in sperm-oocyte interactions. Support for this notion rests with the demonstration that membrane rafts isolated on the basis of their biochemical composition in the form of detergent resistant membranes (DRMs), possess the ability to adhere to homologous zona pellucidae. Furthermore a comprehensive proteomic analysis of the DRMs identified a number of proteins known for their affinity for the zona pellucida in addition to other candidates putatively involved in the mediation of downstream binding and/or fusion with the oolemma. Collectively these data afford novel insights into the subcellular localization and potential functions of membrane rafts in human spermatozoa.

Plasma membrane calcium ATPase 4 (PMCA4) co‐ordinates calcium and nitric oxide signaling in regulating murine sperm functional activity

Olli

Galileo

et al. 2017

Reduced sperm motility (asthenospermia) and resulting infertility arise from deletion of the Plasma Membrane Ca 2+ -ATPase 4 (Pmca4) gene which encodes the highly conserved Ca 2+ efflux pump, PMCA4. This is the major Ca 2+ clearance protein in murine sperm. Since the mechanism underlying asthenospermia in PMCA4's absence or reduced activity is unknown, we investigated if sperm PMCA4 negatively regulates nitric oxide synthases (NOSs) and when absent NO, peroxynitrite, and oxidative stress levels are increased. Using co-immunoprecipitation (Co-IP) and Fluorescence Resonance Energy Transfer (FRET), we show an association of PMCA4 with the NOSs in elevated cytosolic [Ca 2+ ] in capacitated and Ca 2+ ionophore-treated sperm and with neuronal (nNOS) at basal [Ca 2+ ] (ucapacitated sperm). FRET efficiencies for PMCA4-eNOS were 35% and 23% in capacitated and uncapacitated sperm, significantly (p < 0.01) different, with the molecules being <10 nm apart. For PMCA4-nNOS, this interaction was seen only for capacitated sperm where FRET efficiency was 24%, significantly (p < 0.05) higher than in uncapacitated sperm (6%). PMCA4 and the NOSs were identified as interacting partners in a quaternary complex that includes Caveolin1, which co-immunoprecipitated with eNOS in a Ca 2+ -dependent manner. In Pmca4 −/− sperm NOS activity was elevated twofold in capacitated/uncapacitated sperm (vs. wild-type), accompanied by a twofold increase in peroxynitrite levels and significantly (p < 0.001) increased numbers of apoptotic germ cells. The data support a quaternary complex model in which PMCA4 co-ordinates Ca 2+ and NO signaling to maintain motility, with increased NO levels resulting in asthenospermia in Pmca4 −/− males.They suggest the involvement of PMCA4 mutations in human asthenospermia, with diagnostic relevance.asthenospermia, calcium efflux pump, infertility, oxidative stress, sperm damageCapacitation is the series of changes that occur in the final maturation of mammalian sperm in the female in order for them to gain fertilizing competence, and subsequently undergo the acrosome reaction. Both of these processes are dependent on elevated levels of cytosolic Ca 2+ concentration ([Ca 2+ ] c ) (de Lamirande, Leclerc, & Gagnon, 1997;Fraser, 1987). After the calcium influx required to meet this high calcium demand, there needs to be a return to resting cytosolic calcium concentration ([Ca 2+ ] c ) levels (50-100 nM [Herrick et al., 2005]). In mice, in the absence of the major calcium efflux pump, Plasma Membrane Calcium ATPase 4 (PMCA4) (Wennemuth, Babcock, & Hille, 2003), calcium levels remain elevated with loss of sperm motility (Asthenospermia, AS) and resulting male infertility (Okunade et al., 2004;Schuh et al., 2004 (de Lamirande et al., 1997;Fraser, 1987) which rapidly activates them (Knowles & Moncada, 1994). Thus, as PMCA4 extrudes Ca 2+ , the NOSs are held in a locale or microdomain with a relatively low [Ca 2+ ] c, compared to the surrounding environment where it is globally high. In this Ca 2+ -extruding locale, NO...