Sphingosine 1-Phosphate and Sphingosine Kinase Are Involved in a Novel Signaling Pathway Leading to Acrosomal Exocytosis

Abstract

Regulated secretion is a central issue for the specific function of many cells; for instance, mammalian sperm acrosomal exocytosis is essential for egg fertilization. Sphingosine 1-phosphate is a bioactive sphingolipid that regulates crucial physiological processes. Here we report that this lipid triggers acrosomal exocytosis in human sperm by a mechanism involving a G i -coupled receptor. Real-time imaging showed a remarkable increase of cytosolic calcium upon activation with sphingosine 1-phosphate and pharmacological experiments indicate that the process requires extracellular calcium influx through voltage and store-operated calcium channels and efflux from intracellular stores through inositol 1,4,5-trisphosphate-sensitive calcium channels. Sphingosine 1-phosphate-induced exocytosis requires phospholipase C and protein kinase C activation. We investigated possible sources of the lipid. Western blot indicates that sphingosine kinase 1 is present in spermatozoa. Indirect immunofluorescence showed that phorbol ester, a potent protein kinase C activator that can also trigger acrosomal exocytosis, redistributes sphingosine kinase 1 to the acrosomal region. Functional assays showed that phorbol ester-induced exocytosis depends on the activation of sphingosine kinase 1. Furthermore, incorporation of 32 P to sphingosine demonstrates that cells treated with the phorbol ester increase their sphingosine kinase activity that yields sphingosine 1-phosphate. We present here the first evidence indicating that human spermatozoa produce sphingosine 1-phosphate when challenged with an exocytic stimulus. These observations point to a new role of sphingosine 1-phosphate in a signaling cascade that facilitates acrosome reaction providing some clues about novel lipid molecules involved in exocytosis.Sphingosine 1-phosphate (S1P) 2 is a bioactive sphingolipid metabolite that regulates a variety of cellular processes. For years, researchers thought that the biological effects of S1P were due to its actions as an intracellular second messenger. However, surface receptors have recently been identified, and, since then, many downstream effects of S1P receptor activation have been demonstrated (1).

ADP Ribosylation Factor 6 (ARF6) Promotes Acrosomal Exocytosis by Modulating Lipid Turnover and Rab3A Activation

“…Permeabilized cells showed staining at the acrosome region and midpiece (Fig. 8A), which is very different from the cytoplasmic distribution observed in nonpermeabilized spermatozoa (55). As shown in Fig.…”

“…The acrosome behaves as an internal store of releasable calcium (19,47,53,54); efflux from this reservoir through IP 3 -sensitive channels is required for the AR initiated by all inducers (22,28,33,55). Results presented in Figs.…”

Calcium Channels in the Development, Maturation, and Function of Spermatozoa

“…Other cellular ligands can induce the AR such as progesterone (31, 55, 355, 506), ␥-aminobutyric acid (GABA) (280,355,559), glycine (301), EGF (133), ATP (339,442), acetylcholine (66), and sphingosine-1-phosphate (495). The role of many of these "alternative" agonists is not clear.…”

Role of Ion Channels in the Sperm Acrosome Reaction

“…New findings have engaged the field in reevaluating other cellular ligands able to trigger the AR, since the physiological role of ZP in triggering this reaction in mouse sperm has been questioned (Jin et al 2011;Tateno et al 2013). These include progesterone (Thomas and Meizel 1989;Blackmore et al 1990), γ-aminobutyric acid (GABA) (Wistrom and Meizel 1993), glycine (Kumar and Meizel 2008), EGF (Daniel et al 2010), ATP (Foresta et al 1992;Luria et al 2002), acetylcholine (Bray et al 2005), and sphingosine-1-phosphate (Suhaiman et al 2010).…”