Exocytosis Of Acrosomal Contents Research Articles

Blend of Roselle Calyx and Selected Fruit Modulates Testicular Redox Status and Sperm Quality of Diabetic Rats

Mammalian fertilization, a species-specific event, is the net result of a highly orchestrated process that collectively results in the fusion of two radically different-looking haploid cells, sperm and egg, to form a diploid zygote, a cell with somatic chromosome numbers. Prior to the interaction of the opposite gametes, mammalian spermatozoa undergo many fascinating changes during development in the testis, maturation in the epididymis, and capacitation in the female genital tract. Only the capacitated spermatozoa interact with the extracellular coat, the zona pellucida, which surrounds the mammalian oocyte. The tight and irreversible binding of the opposite gametes in the mouse and many other mammals studied, including human, starts a ca2+- dependent signal transduction pathway that results in the exocytosis of acrosomal contents at the site of the sperm binding. The hydrolytic action of the acrosomal glycohydrolases and proteinases, released at the site of the sperm-egg binding, along with the enhanced thrust generated by the hyperactivated spermatozoon, are important factors that regulate the penetration of the zona pellucida and fusion of the oppositegametes. The purpose of this editorial is to highlight the well programmed molecular events that are necessary before sperm-egg adhesion. In addition, my intention is to discuss the increasing controversy about the mechanism(s) that regulate mammalian sperm-egg interactions.

Mechanisms of Mammalian Sperm-Egg Interaction Leading to Fertilization

Mammalian fertilization, a species-specific event, is the net result of a highly orchestrated process that collectively results in the fusion of two radically different-looking haploid cells, sperm and egg, to form a diploid zygote, a cell with somatic chromosome numbers. Prior to the interaction of the opposite gametes, mammalian spermatozoa undergo many fascinating changes during development in the testis, maturation in the epididymis, and capacitation in the female genital tract. Only the capacitated spermatozoa interact with the extracellular coat, the zona pellucida, which surrounds the mammalian oocyte. The tight and irreversible binding of the opposite gametes in the mouse and many other mammals studied, including human, starts a ca2+- dependent signal transduction pathway that results in the exocytosis of acrosomal contents at the site of the sperm binding. The hydrolytic action of the acrosomal glycohydrolases and proteinases, released at the site of the sperm-egg binding, along with the enhanced thrust generated by the hyperactivated spermatozoon, are important factors that regulate the penetration of the zona pellucida and fusion of the oppositegametes. The purpose of this editorial is to highlight the well programmed molecular events that are necessary before sperm-egg adhesion. In addition, my intention is to discuss the increasing controversy about the mechanism(s) that regulate mammalian sperm-egg interactions.

Fertilization with acrosome-reacted mouse sperm: Implications for the site of exocytosis

For successful mammalian fertilization, sperm penetrate the outer investments of female gametes in the ampulla of the oviduct and fuse with the egg plasma membrane. The “fertilizing” sperm traverse the cumulus mass surrounding ovulated eggs, bind temporarily, and then penetrate through the zona pellucida, a structured extracellular glycomatrix, to enter into the perivitelline space (Fig. 1A). Underlying the anterior plasma membrane of the mouse sperm head is the acrosome, a Golgi-derived subcellular organelle. Sperm that approach ovulated eggs in the female reproductive tract have an intact acrosome. Before fertilization, the outer acrosomal membrane fuses with the sperm plasma membrane resulting in exocytosis of acrosomal contents (Fig. 1B). Only acrosome-reacted sperm are observed in the perivitelline space between the egg plasma membrane and the inner aspect of the zona pellucida and only acrosome-reacted sperm fuse with the egg. Thus, at some point during sperm penetration through the investments of ovulated eggs, acrosome exocytosis must occur. But where (Fig. 1A)? This is the issue that concerns Inoue et al. (1) in their study in PNAS.

Capacitated acrosome-intact mouse spermatozoa bind to Sepharose beads coated with functional neoglycoproteins

Capacitated acrosome-intact mouse spermatozoa bind to the egg’s extracellular coat, the zona pellucida (ZP), in a carbohydrate-mediated receptor–ligand manner. The tight irreversible binding of the opposite gametes triggers a signal transduction pathway resulting in the exocytosis of acrosomal contents (i.e., induction of the acrosome reaction [AR]). Previously, we demonstrated that a hexose (mannose) and two amino sugars ( N-acetylglucosamine and N-acetylgalactosamine), when covalently conjugated to bovine serum albumin (BSA) (functional neoglycoproteins, ngps), mimicked mZP3 and induced the AR [Biol. Reprod. 60 (1999) 94–101]. To further elucidate the specificity of sperm–ngp interaction and the mZP3 mimicking role of the functional ngps, we have examined binding of the mouse spermatozoa to Sepharose 4B beads coated with the functional and non-functional ngps as well as BSA, ovalbumin (OVA), or asialofetuin (ASF). A significantly greater number of capacitated acrosome-intact spermatozoa bound to the beads coated with functional ngps than the beads coated with non-functional ngps, BSA, OVA, or ASF. The binding was temperature-sensitive and was highest when the sperm–bead assay was carried out at 37 °C. Blocking of in vitro capacitation, by including calmodulin antagonists in the incubation medium, prevented sperm from binding to the beads. Furthermore, inclusion of free sugars (mannose, N-acetylglucosamine, or N-acetylgalactosamine) in the binding assay, either individually or as a mixture, inhibited sperm–bead binding in a concentration-dependent manner. Taken together, our data provide evidence strongly suggesting that binding of capacitated spermatozoa to the ngp-coated Sepharose beads is specific. The beads that mimic zona-intact eggs provide an excellent tool for examining pharmacological effects of reagents that alter the sperm function. In addition, the immobilized ngp(s) will be useful as an affinity medium to isolate the sperm surface receptor(s) that recognize and bind to the sugar residues.

Assessment of acrosomal status in rat spermatozoa: studies on carbohydrate and non-carbohydrate agonists

In the mouse and several other species, including man, capacitated acrosome-intact spermatozoa interact with natural [soluble zona pellucida (ZP) and progesterone (P4)] and synthetic [neoglycoproteins (ngps) and calcium (Ca 2+) ionophore] agonists, prior to the initiation of a Ca 2+-dependent signal transduction cascade. The net result is the fusion of the sperm plasma membrane overlying the outer acrosomal membrane at multiple sites and exocytosis of acrosomal contents [i.e., induction of the acrosome reaction (AR)]. This step is believed to be a prerequisite that enables the acrosome-reacted spermatozoon to penetrate the ZP and fertilize the egg. Although the rat is one of the most commonly used laboratory animals, very little is known about the chemical nature of agonists that induce the AR in this species. The lack of this information is primarily due to the fact that the rat sperm acrosome is a relatively thin structure. Thus, it is difficult to assess the status of the sperm acrosome in this species. In this report, we describe the use of a Coomassie brilliant blue dye staining procedure to assess the status of the rat sperm acrosome by light microscopy. The procedure is highly reproducible and has allowed us to determine the effects of carbohydrate (ngps and mouse ZP) and noncarbohydrate (P4 and Ca 2+ ionophore) agonists on capacitated spermatozoa. In addition, we have used a pharmacological approach to examine the functional significance of calmodulin (CaM), a Ca 2+-binding protein, in induction of the AR in spermatozoa. Data presented in this report demonstrate that several ngps, solubilized mZP, P4, and Ca 2+ ionophores induce the AR in rat spermatozoa. Furthermore, we demonstrate that, whereas CaM antagonists blocked P4-induced AR, most of the inhibitors used had no significant effect on the Ca 2+ ionophore-induced (nonphysiological) AR.

Calmodulin Signals Capacitation and Triggers the Agonist-Induced Acrosome Reaction in Mouse Spermatozoa

Capacitated acrosome-intact spermatozoa interact with specific sugar residues on neoglycoproteins (ngps) or solubilized zona pellucida (ZP), the egg's extracellular glycocalyx, prior to the initiation of a signal transduction cascade that results in the fenestration and fusion of the sperm plasma membrane and the outer acrosomal membrane at multiple sites and exocytosis of acrosomal contents (i.e., induction of the acrosome reaction (AR)). The AR releases acrosomal contents at the site of sperm–zona binding and is thought to be a prerequisite event that allows spermatozoa to penetrate the ZP and fertilize the egg. Since Ca2+/calmodulin (CaM) plays a significant role in several cell signaling pathways and membrane fusion events, we have used a pharmacological approach to examine the role of CaM, a calcium-binding protein, in sperm capacitation and agonist-induced AR. Inclusion of CaM antagonists (calmodulin binding domain, calmidazolium, compound 48/80, ophiobolin A, W5, W7, and W13), either in in vitro capacitation medium or after sperm capacitation blocked the npg-/ZP-induced AR. Purified CaM largely reversed the AR blocking effects of antagonists during capacitation. Our results demonstrate that CaM plays an important role in priming (i.e., capacitation) of mouse spermatozoa as well as in the agonist-induced AR. These data allow us to propose that CaM regulates these events by modulating sperm membrane component(s).

Mammalian sperm molecules that are potentially important in interaction with female genital tract and egg vestments.

Fertilisation is a highly programmed process by which two radically different cells, sperm and egg, unite to form a zygote, a cell with somatic chromosome numbers. Development of the zygote begins immediately after sperm and egg haploid pronuclei come together, pooling their chromosomes to form a single diploid nucleus with the parental genes. Mammalian fertilisation is the net result of a complex set of molecular events which allow the capacitated spermatozoa to recognise and bind to the egg's extracellular coat, the zona pellucida (ZP), undergo the acrosome reaction, and fuse with the egg plasma membrane. Sperm-zona (egg) interaction leading to fertilisation is a species-specific carbohydrate-mediated event which depends on glycan-recognising proteins (glycosyltransferases/glycosidases/lectin-like molecules) on sperm plasma membrane (receptors) and their complementary glycan units (ligands) on ZP. The receptor-ligand interaction event initiates a signal transduction pathway resulting in the exocytosis of acrosomal contents. The hydrolytic action of the sperm glycohydrolases and proteases released at the site of sperm-egg interaction, along with the enhanced thrust generated by the hyperactivated beat pattern of the bound spermatozoon, are important factors regulating the penetration of egg investments. This review focuses on sperm molecules believed to be important for the interaction with the female genital tract, passage through cumulus oophorus and attachment to ZP, induction of the acrosome reaction, secondary binding events, and passage through the ZP. An understanding of the expression and modifications of molecules thought to be important in multiple events leading to fertilisation will allow new strategies to block these modifications and alter sperm function.