Phospholipase C isoforms in mammalian spermatozoa: potential components of the sperm factor that causes Ca2+ release in eggs

Abstract

ResearchInjection of a soluble protein factor from mammalian spermatozoa triggers Ca 2+ oscillations in mammalian eggs similar to those seen at fertilization. This sperm factor also generates inositol 1,4,5-trisphosphate and causes Ca 2+ release in sea urchin egg homogenates and frog eggs. Recent studies have indicated that the sperm factor may be an inositol-specific phospholipase C (PLC) activity. This study investigated whether any of the commonly known PLC isoforms are components of the sperm factor. PLCβ, PLCγ and PLCδ isoforms were shown to be present in boar sperm extracts. However, upon column fractionation of sperm extracts, none of the PLC isoforms detected correlated with the ability to cause Ca 2+ release in eggs. In addition to our previous work on recombinant PLCs, it was also shown that PLCδ3, PLCδ4 and its splice variant PLCδ4 Alt1 fail to cause Ca 2+ release. The recently discovered 255 kDa PLCε isoform also appears unlikely to be a component of the sperm factor, as fractionation of sperm extracts on a gel filtration column demonstrated that the peak of Ca 2+ -releasing activity was associated with fractions of 30-70 kDa. These findings indicate that the sperm factor that triggers Ca 2+ release in eggs does not appear to have a known PLC isoform as one of its components.

Mammalian Fertilization: From Sperm Factor to Phospholipase Cζ

“…Besides the sperm-specific localization of this PLC, several other unique features make PLCf a very attractive candidate to be SF. First, the molecular weight of PLCf of approximately 70 kDa is within the 30-70 kDa range that has long been anticipated by chromatographic fractionation of sperm extracts to be the molecular weight of SF (Wu et al, 1998;Parrington et al, 2002). Second, injection of PLCf mRNA into mouse eggs was shown to trigger fertilization-like [Ca 2+ ] i oscillations, whereas injection of PLCd1 mRNA at the same pipette concentration could not initiate [Ca 2+ ] i oscillations Saunders et al, 2002), indicating that PLCf has a unique PLC activity.…”

“…Moreover, PLCs are well known to be involved in cell growth, differentiation, and cytoskeletal organization (Divecha and Irvine, 1995;Rhee, 2001). To date, 12 PLC isozymes (b1-4, c1-2, d1-4, e, f) have been identified, and expression of PLCb1, b3, c1, and c2 has been noted in mouse eggs (Dupont et al, 1996;Mehlmann et al, 1998;Wang et al, 1998), whereas expression of b1-3, c1-2, d2, d4, and f has been detected in sperm Fukami et al, 2001;Wu et al, 2001b;Parrington et al, 2002;Saunders et al, 2002). Importantly, discriminating whether a PLC(s) from eggs or sperm, or a combination of PLCs from both gametes, plays a role in mammalian fertilization has proven an elusive task.…”

Phospho-Signaling at Oocyte Maturation and Fertilization: Set Up for Embryogenesis and Beyond Part II. Kinase Regulators and Substrates

“…It should be noted that Src-dependent activation of PLC involves a new function of PLC as GEF for small G-protein Ras (Bivona et al 2003), suggesting that other means of cellular function contributes to egg activation in these species. On the other hand, Ca 2+ release associated with mammalian fertilization does not seem to involve tyrosine kinase activity and PLC activation, probably because sperm-derived PLC activity is necessary and sufficient for sperm-induced Ca 2+ release in these species (Kurokawa et al 2004;Parrington et al 2002;Saunders et al 2002).…”

Flipping the switch: How a sperm activates the egg at fertilization

“…A breakthrough came with the discovery that the mammalian sperm factor was a sperm-specific PLC that had very distinctive enzymatic properties compared to other known PLCs (Jones et al, 1998(Jones et al, , 2000Parrington et al, 1999Parrington et al, , 2002). This discovery was made possible by the use of a cell-free eggbased Ca 2ϩ release assay, the sea urchin egg homogenate, which enabled a biochemical dissection of the sperm factor's mechanism of action in a way that would have been much more difficult using an intact egg.…”