Sperm-egg recognition in the mouse: characterization of sp56, a sperm protein having specific affinity for ZP3.

Cheng, Ann‐Lii; Le, Tung B. K.; Palacios, Marcelo Marcet; Bookbinder, L H; Wassarman, Paul M.; Suzuki, Fumie; Bleil, Jeffrey D.

doi:10.1083/jcb.125.4.867

Cited by 159 publications

(102 citation statements)

References 20 publications

(36 reference statements)

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“…Capacitated Acr ϩ/ϩ and Acr Ϫ/Ϫ mouse sperm were treated with calcium ionophore A23187, and the time course of protein release from the acrosome was monitored by indirect immunofluorescent staining using two monoclonal antibodies against mouse acrosomal proteins (mAb sp56 (12,13) and mAb MC101 (8); Fig. 1).…”

Section: Resultsmentioning

confidence: 99%

Acrosin Accelerates the Dispersal of Sperm Acrosomal Proteins during Acrosome Reaction

Yamagata¹,

Murayama²,

Okabe³

et al. 1998

Journal of Biological Chemistry

154

108

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Using homologous recombination, we have previously produced male mice carrying a disruptive mutation (Acr ؊/؊ ) in the acrosin gene. Although Acr ؊/؊ mouse sperm lacking the acrosin protease activity still penetrated the zona pellucida and fertilized the egg, the mutant sperm exhibited a delay in penetration of the zona pellucida solely at the early stages after insemination. To further elucidate the role of acrosin in fertilization, we have examined the involvement of acrosin in the acrosome reaction of sperm using the Acr ؊/؊ mutant mice. When the ability of sperm to adhere (attach) and bind to the zona pellucida of cumulus-free eggs was assessed in vitro, no significant difference was observed among Acr ؉/؉ , Acr ؉/؊ , and Acr ؊/؊ mouse sperm. Immunocytochemical analysis demonstrated that the release of several acrosomal proteins from the acrosome of Acr ؊/؊ mouse sperm was significantly delayed during the calcium ionophore-and solubilized zona pellucidainduced acrosome reaction, despite normal membrane vesiculation. These data indicate that the delayed sperm penetration of the zona pellucida in the Acr ؊/؊ mouse results from the altered rate of protein dispersal from the acrosome and provide the first evidence that the major role of acrosin is to accelerate the dispersal of acrosomal components during acrosome reaction.The acrosome reaction of sperm, a fusion (vesiculation) event between the overlying plasma and outer acrosomal membranes, occurs following the binding of sperm to the zona pellucida (ZP), 1 an extracellular glycoprotein matrix surrounding the egg. This exocytotic reaction is required for fertilization, because only acrosome-reacted sperm are capable of penetrating ZP and of fusing with the egg plasma membrane (for review see Ref. 1). The acrosomal components, including hydrolytic enzymes, are released by the acrosome reaction and then interact initially with ZP to facilitate the sperm penetration of the glycoprotein matrix.Acrosin, an endoprotease with a trypsin-like substrate specificity, is localized in the acrosomal matrix as an enzymatically inactive zymogen, proacrosin, that is then converted into the active form as a consequence of the acrosome reaction (2-4). The physiological role of acrosin in fertilization has long been believed to be the limited proteolysis of the ZP, thus enabling the sperm to penetrate the ZP. Using homologous recombination, we have successfully produced male mice carrying a disruptive mutation in the acrosin gene (Acr) and found that the mouse sperm lacking the acrosin protease activity (Acr Ϫ/Ϫ ) still penetrate ZP and normally fertilize the egg (5). These data provide evidence that acrosin is not essential for sperm penetration of the ZP. However, as compared with Acr ϩ/ϩ and Acr ϩ/Ϫ mice, Acr Ϫ/Ϫ mouse sperm showed a delay in sperm penetration of the ZP solely at the early stages after insemination (5). A recent report using separate lines of Acr Ϫ/Ϫ mice (6) has confirmed that sperm lacking acrosin exhibit the delayed fertilization. Thus, these results imply ...

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Section: Resultsmentioning

confidence: 99%

Acrosin Accelerates the Dispersal of Sperm Acrosomal Proteins during Acrosome Reaction

Yamagata¹,

Murayama²,

Okabe³

et al. 1998

Journal of Biological Chemistry

154

108

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“…75 Immunization of female BALB/c mice with E. coli-expressed recombinant Sp56 fusion protein led to sub-fertility. 76 However, sp56 null mice showed normal fecundity and thus undermining its role during fertilization.…”

Section: Sp56mentioning

confidence: 99%

Milestones in contraceptive vaccines development and hurdles in their application

Gupta

Shrestha

Minhas

2013

Human Vaccines & Immunotherapeutics

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“…ZP3R/sp56 was initially discovered by photoaffinity crosslinking with ZP3 (9) and later characterized as having specific affinity for mouse ZP3 oligosaccharides involved in sperm binding (10). The cloning of ZP3R/sp56 demonstrated that it is a member of the superfamily of C3/C4 binding proteins, possessing multiple short consensus repeats (so-called "sushi" domains) characteristic of complement regulatory proteins; however, the protein lacked an obvious carbohydrate recognition domain (8).…”

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confidence: 99%

“…ZP3R/sp56 is first detectable during late meiosis although during the course of spermatid differentiation, the acrosomal glycoprotein undergoes post-translational modifications, which may include the modification of carbohydrate side chains (14). Initial reports suggested that ZP3R/sp56 is a plasma membrane protein (10,15). However, our laboratory later demonstrated by conventional immuno-electron microscopy that ZP3R/sp56 is an intra-acrosomal protein and is, in fact, part of a stable acrosomal matrix (14,16).…”

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confidence: 99%

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Recombinant Mouse Sperm ZP3-binding Protein (ZP3R/sp56) Forms a High Order Oligomer That Binds Eggs and Inhibits Mouse Fertilization in Vitro

Buffone

Zhuang²,

Ord

et al. 2008

Journal of Biological Chemistry

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Many candidates have been proposed as zona pellucida-binding proteins. Without precluding a role for any of those candidates, we focused on mouse sperm protein ZP3R/sp56, which is localized in the acrosomal matrix. The objective of this study was to analyze the role of ZP3R/sp56 in mouse fertilization. We expressed recombinant ZP3R/sp56 as a secreted protein in HEK293 cells and purified it from serum-free, conditioned medium. In the presence of reducing agents, the recombinant ZP3R/sp56 exhibited a molecular weight similar to that observed for the native ZP3R/sp56. Reminiscent of the native protein, recombinant ZP3R/sp56 formed a high molecular weight, disulfide cross-linked oligomer consisting of six or more monomers under non-reducing conditions. Recombinant ZP3R/sp56 bound to the zona pellucida of unfertilized eggs but not to 2-cell embryos, indicating that the changes that take place in the zona pellucida at fertilization affected the interaction of this protein with the zona pellucida. The extent of in vitro fertilization was reduced in a dose-dependent manner when unfertilized eggs were preincubated with recombinant ZP3R/sp56 (74% drop at the maximum concentrations assayed). Eggs incubated with the recombinant protein showed an absence of or very few sperm in the perivitelline space, suggesting that the reduction in the fertilization rate is caused by the inhibition of sperm binding and/or penetration through the zona pellucida. These results indicate that sperm ZP3R/sp56 is important for sperm-zona interactions during fertilization and support the concept that the acrosomal matrix plays an essential role in mediating the binding of sperm to the zona pellucida. Sperm-zona pellucida (ZP)3 binding is an important step of gamete interaction in many species. The actual mechanism of binding is still unresolved. For mammals, some researchers favor a two-step process whereby primary binding with the ZP occurs through a plasma membrane receptor(s) of sperm with intact acrosomes, and secondary binding utilizes intraacrosomal proteins that are exposed during the course of or following acrosomal exocytosis ("Acrosome Reaction Model") (1, 2). We and others feel that is difficult to ascertain the definitive acrosomal status at the time of binding to the zona pellucida. It is possible that during sperm capacitation subtle changes occur to the acrosomal region that initiate the acrosomal exocytotic process, thereby exposing acrosomal proteins involved in binding to the ZP (Acrosomal Exocytosis Model) (1, 3).While there are only a few ZP sulfoglycoproteins involved in these binding events, a large number of sperm molecules have been shown to possess ZP binding activity (2). The list includes ␤-1,4 galactosyltransferase (4), zonadhesin (5), SED-1 (6), spermadhesins (7), and ZP3R/sp56 (formerly sp56) (8). Without precluding a role for any of these candidates in sperm-ZP interactions, we focused on mouse sperm protein ZP3R/sp56, because of our interest in defining the function of the acrosomal matrix.ZP3R/sp56 was initially ...

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Sperm-egg recognition in the mouse: characterization of sp56, a sperm protein having specific affinity for ZP3.

Cited by 159 publications

References 20 publications

Acrosin Accelerates the Dispersal of Sperm Acrosomal Proteins during Acrosome Reaction

Acrosin Accelerates the Dispersal of Sperm Acrosomal Proteins during Acrosome Reaction

Milestones in contraceptive vaccines development and hurdles in their application

Recombinant Mouse Sperm ZP3-binding Protein (ZP3R/sp56) Forms a High Order Oligomer That Binds Eggs and Inhibits Mouse Fertilization in Vitro

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