Dynamic resolution of acrosomal exocytosis in human sperm

Abstract

An essential step in mammalian fertilisation is the sperm acrosome reaction (AR) – exocytosis of a single large vesicle (the acrosome) that surrounds the nucleus at the apical sperm head. The acrosomal and plasma membranes fuse, resulting in both the release of factors that might facilitate penetration of the zona pellucida (which invests the egg) and the externalisation of membrane components required for gamete fusion. Exocytosis in somatic cells is a rapid process – typically complete within milliseconds – yet acrosomal enzymes are required throughout zona penetration – a period of minutes. Here, we present the first studies of this crucial and complex event occurring in real-time in individual live sperm using time-lapse fluorescence microscopy. Simultaneous imaging of separate probes for acrosomal content and inner acrosomal membrane show that rapid membrane fusion, initiated at the cell apex, is followed by exceptionally slow dispersal of acrosomal content (up to 12 minutes). Cells that lose their acrosome prematurely (spontaneous AR), compromising their ability to penetrate the egg vestments, are those that are already subject to a loss of motility and viability. Cells undergoing stimulus-induced AR (progesterone or A23187) remain viable, with a proportion remaining motile (progesterone). These findings suggest that the AR is a highly adapted form of exocytosis.

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

“…The correlation with high spatiotemporal resolution [Ca 2ϩ ] i changes and AR in single spermatozoa is important to understand this fundamental process required for fertilization. Now it is possible to image the acrosome status using a transgenic mouse which expresses GFP in the acrosome (379) or fluorescence probes (186,240,509).…”

CaMKIIα interacts with multi-PDZ domain protein MUPP1 in spermatozoa and prevents spontaneous acrosomal exocytosis

“…A special form of Ca 2+ -regulated exocytosis is the sperm acrosome reaction, a mandatory event in the fertilization process, initiating two essential events: release of hydrolyzing enzymes, necessary for sperm to penetrate the egg glycoprotein matrix, the zona pellucida (ZP) (Wassarman and Litscher, 2008), and exposure of the inner acrosomal membrane, which thereby becomes accessible for the egg plasma membrane during the ultimate fusion event (Florman et al, 2008;Harper et al, 2008). Although acrosomal exocytosis in sperm cells differs from exocytotic events in other cellular systems, mainly because the acrosome is a huge and single secretory vesicle, certain remarkable parallels have been outlined between the acrosome reaction in sperm and vesicle fusion in neurons and neuroendocrine or exocrine cells Tulsiani and Abou-Haila, 2004).…”

A model of the acrosome reaction progression via the acrosomal membrane-anchored protein equatorin

“…zonadhesin (Hardy et al 1991, Gao & Garbers 1998, Olson et al 2004, ACRV1/SP-10 (Foster et al 1994), AM67/ZP3R/sp56 (Cheng et al 1994, Foster et al 1997, Kim et al 2001a, ZPBP1/sp38/IAM38 (Mori et al 1995, Yu et al 2006, AM50 (Westbrook-Case et al 1994, Kim et al 2001b, MC41 (Tanii et al 2002), SAMP32 (Hao et al 2002), ESP , SAMP14 , and acrosomal enzymes (reviewed by Moreno & Alvarado (2006)). In previous reports relating acrosomal molecules as described above, the status of the acrosome and the acrosome reaction progression were often measured by acrosome-related lectin stains such as Arachis hypogaea agglutinin (PNA), concanavalin A, and Pisum sativum agglutinin (Kallajoki et al 1986, Mortimer et al 1987, Holden et al 1990, Holden & Trounson 1991, Kinger & Rajalakshmi 1995, De Blas et al 2005, Harper et al 2008, Mugnier et al 2009, and in transgenic mice such as acrosin-green fluorescent protein (GFP) transgenic mice (Nakanishi et al 1999, Kim & Gerton 2003, Buffone et al 2009b. In human sperm, the acrosome reaction progression was identified as having six stages, with the intermediate ones based on transmission electron microscopic observations, in which acrosomal matrix material was lost while outer membranes appeared to retain their integrity (Stock & Fraser 1987, Yudin et al 1988.…”