Abstract
BackgroundFertilization of echinoderm eggs is accompanied by dynamic changes of the actin cytoskeleton and by a drastic increase of cytosolic Ca2+. Since the plasma membrane-enriched phospholipid phosphatidylinositol 4,5-bisphosphate (PIP2) serves as the precursor of inositol 1,4,5 trisphosphate (InsP3) and also regulates actin-binding proteins, PIP2 might be involved in these two processes.Methodology/Principal FindingsIn this report, we have studied the roles of PIP2 at fertilization of starfish eggs by using fluorescently tagged pleckstrin homology (PH) domain of PLC-δ1, which has specific binding affinity to PIP2, in combination with Ca2+ and F-actin imaging techniques and transmission electron microscopy. During fertilization, PIP2 increased at the plasma membrane in two phases rather than continually decreasing. The first increase was quickly followed by a decrease about 40 seconds after sperm-egg contact. However, these changes took place only after the Ca2+ wave had already initiated and propagated. The fertilized eggs then displayed a prolonged increase of PIP2 that was accompanied by the appearance of numerous spikes in the perivitelline space during the elevation of the fertilization envelope (FE). These spikes, protruding from the plasma membrane, were filled with microfilaments. Sequestration of PIP2 by RFP-PH at higher doses resulted in changes of subplasmalemmal actin networks which significantly delayed the intracellular Ca2+ signaling, impaired elevation of FE, and increased occurrences of polyspermic fertilization.Conclusions/SignificanceOur results suggest that PIP2 plays comprehensive roles in shaping Ca2+ waves and guiding structural and functional changes required for successful fertilization. We propose that the PIP2 increase and the subsequent formation of actin spikes not only provide the mechanical supports for the elevating FE, but also accommodate increased membrane surfaces during cortical granule exocytosis.
Highlights
Starfish oocytes arrested at the first prophase of meiosis are characterized by a large nucleus
Mature eggs of A. aranciacus microinjected with either RFP-pleckstrin homology (PH) or R40A mutant proteins (330 mM, pipette concentration) were stimulated for Ca2+ signaling and cortical granule exocytosis by fertilization or photoactivation of uncaged InsP3
Mouse eggs displayed an increase of PIP2 at the plasma membrane [31], but an apparent decrease of PIP2 was subsequently reported in sea urchin eggs [29]
Summary
Starfish oocytes arrested at the first prophase of meiosis are characterized by a large nucleus (germinal vesicle, GV). When exposed to the maturation hormone (1-methyladenine, 1-MA), the oocytes reenter the cell cycle and proceed with meiosis to become mature eggs. Starfish eggs are adequate to monitor other cytological changes occurring at fertilization [2]. The massive Ca2+ release in fertilized eggs in part facilitates exocytosis of cortical granules. The initial rise of Ca2+ induced by the sperm occurs at the egg cortex (cortical flash), and is followed by the propagation of Ca2+ waves starting from the site of sperm interaction [3,4]. Fertilization of echinoderm eggs is accompanied by dynamic changes of the actin cytoskeleton and by a drastic increase of cytosolic Ca2+. Since the plasma membrane-enriched phospholipid phosphatidylinositol 4,5bisphosphate (PIP2) serves as the precursor of inositol 1,4,5 trisphosphate (InsP3) and regulates actin-binding proteins, PIP2 might be involved in these two processes
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
