Biological Effects of Recombinant Human Zona Pellucida Proteins on Sperm Function1

Caballero-Campo, Pedro; Chirinos, Mayel; Fan, Xue Jun; González-González, María Elena; Galicia-Chavarría, Myrna; Larrea, Fernando; Gerton, George L.

doi:10.1095/biolreprod.105.047522

Cited by 58 publications

(50 citation statements)

References 63 publications

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“…The importance and site of ZP protein glycosylation is also under revision (191). ZP3 is considered the main physiological inductor of the mammalian sperm AR (177,550) though in the human system ZP4 and ZP1 can induce this reaction to a lesser extent (76,200,282). In spite of these, recent experiments in which oocytes expressing genetically modified mouse ZPs are used show that very few spermatozoa bound to ZP have undergone AR after 1 h (28, 191).…”

Section: A Generalitiesmentioning

confidence: 99%

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

Darszon

Nishigaki

Beltrán

et al. 2011

Physiological Reviews

314

313

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-permeable channels and located them at distinct sites in spermatozoa so that they can help fertilize the egg. New tools to study sperm ionic currents, and image intracellular Ca 2ϩ with better spatial and temporal resolution even in swimming spermatozoa, are revealing how sperm ion channels participate in fertilization. This review critically examines the involvement of Ca 2ϩ channels in multiple signaling processes needed for spermatozoa to mature, travel towards the egg, and fertilize it. Remarkably, these tiny specialized cells can express exclusive channels like CatSper for Ca 2ϩ and SLO3 for K ϩ , which are attractive targets for contraception and for the discovery of novel signaling complexes. Learning more about fertilization is a matter of capital importance; societies face growing pressure to counteract rising male infertility rates, provide safe male gamete-based contraceptives, and preserve biodiversity through improved captive breeding and assisted conception initiatives.

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Section: A Generalitiesmentioning

confidence: 99%

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

Darszon

Nishigaki

Beltrán

et al. 2011

Physiological Reviews

314

313

View full text Add to dashboard Cite

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“…87,101 E. coli-expressed recombinant human ZP1, ZP3 and ZP4 fail to induce any significant increase in the AR, whereas baculovirusexpressed recombinant ZP1, ZP3 and ZP4 induce dose-dependent increases in the AR. 31,32,52,53,61 These studies suggest that glycosylation of human zona proteins is critical for induction of the AR. Expression of recombinant human ZP3 and ZP4 using a baculovirus expression system in the presence of tunicamycin made available these proteins with reduced N-linked oligosaccharides.…”

Section: Zona-induced Acrosome Reaction Sk Gupta and B Bhandari 102mentioning

confidence: 92%

“…58 In hamsters and humans, ZP3 performs the function of primary sperm receptor. 52,53,55,59 Studies employing purified native human ZP3, 60 as well as baculovirus-expressed recombinant ZP3, 32,52,53,61 have shown dose-dependent increases in acrosomal exocytosis. Moreover, human ZP3 expressed in mammalian cells also leads to an increase in the AR.…”

Section: Zp3mentioning

confidence: 99%

“…E. coli-expressed recombinant human ZP4 binds to the anterior head of capacitated acrosome-intact human spermatozoa but does not induce the AR. 53 On the other hand, baculovirus-expressed recombinant human ZP4 not only binds to the anterior head of capacitated acrosome-intact spermatozoa, but also induces a dose-dependent increase in the AR 32,52,53,61 (Table 1). These observations were further confirmed by employing immunoaffinity purified native human ZP4 from solubilized human ZP.…”

Section: Zp4mentioning

confidence: 99%

“…As discussed above, in humans (in addition to ZP3), ZP1 and ZP4 also mediate the induction of the AR. 31,32,52,53,60,61 Using pharmacological inhibitors, subtle differences in the downstream signalling pathways used by the ZP glycoproteins were observed, which are summarized in Table 2. ZP3-mediated induction of the AR in humans is inhibited by pertussis toxin, whereas pertussis toxin does not inhibit ZP1-or ZP4-mediated acrosomal exocytosis, which indicates that ZP1/ZP4 act through a G i protein-independent pathway.…”

Section: Zp4mentioning

confidence: 99%

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Acrosome reaction: relevance of zona pellucida glycoproteins

Gupta

Bhandari²

2010

Asian J Androl

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During mammalian fertilisation, the zona pellucida (ZP) matrix surrounding the oocyte is responsible for the binding of the spermatozoa to the oocyte and induction of the acrosome reaction (AR) in the ZP-bound spermatozoon. The AR is crucial for the penetration of the ZP matrix by spermatozoa. The ZP matrix in mice is composed of three glycoproteins designated ZP1, ZP2 and ZP3, whereas in humans, it is composed of four (ZP1, ZP2, ZP3 and ZP4). ZP3 acts as the putative primary sperm receptor and is responsible for AR induction in mice, whereas in humans (in addition to ZP3), ZP1 and ZP4 also induce the AR. The ability of ZP3 to induce the AR resides in its C-terminal fragment. O-linked glycans are critical for the murine ZP3-mediated AR. However, N-linked glycans of human ZP1, ZP3 and ZP4 have important roles in the induction of the AR. Studies with pharmacological inhibitors showed that the ZP3-induced AR involves the activation of the G i -coupled receptor pathway, whereas ZP1-and ZP4-mediated ARs are independent of this pathway. The ZP3-induced AR involves the activation of T-type voltage-operated calcium channels (VOCCs), whereas ZP1-and ZP4-induced ARs involve both T-and L-type VOCCs. To conclude, in mice, ZP3 is primarily responsible for the binding of capacitated spermatozoa to the ZP matrix and induction of the AR, whereas in humans (in addition to ZP3), ZP1 and ZP4 also participate in these stages of fertilisation. Keywords: acrosome reaction; fertilisation; oocyte; signalling pathways; spermatozoa; zona pellucida glycoproteins INTRODUCTION Mammalian fertilisation is a highly synchronized process that involves a complex series of interactions between the spermatozoon and the egg, culminating in their unison. The initial steps in fertilisation involves the binding of the spermatozoon to the zona pellucida (ZP) matrix surrounding the egg, followed by induction of the acrosome reaction (AR) in the zona-bound spermatozoon, a pre-requisite for penetration of the ZP matrix by the spermatozoon. The spermatozoon acrosome is a Golgi-derived organelle that forms a cap over the anterior two-thirds of its nucleus. The AR involves the fusion of the sperm membrane with the outer acrosomal membrane, resulting in release of the acrosomal contents and exposure of the inner acrosomal membrane on the anterior head of the spermatozoon. Various physiological agents, such as progesterone, serum albumin, follicular fluid, hormones (including biogenic amines), hydrolytic enzymes (particularly proteases), hyaluronic acid and ZP glycoproteins, have been implicated in the induction of the AR. [1][2][3] In the present paper, we review the role of the ZP matrix and its constituents in AR induction. Various downstream signalling pathways involved in the ZP glycoprotein-induced AR will also be discussed. On the basis of the current literature and studies from our group, the salient differences in the ZP glycoprotein-mediated induction of the AR in mouse versus human will be highlighted.

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Sperm‐Binding Assay Using an In Vitro 3D Model of the Mammalian Cumulus‐Oocyte Complex

2020

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We have recently described a new model to study gamete interaction in mammalian species. The model recreates the spherical surface of the oocyte by using magnetic Sepharose beads coated with a layer of a recombinant protein involved in gamete interaction (such as ZP2, or the IZUMO1 receptor JUNO) and an external layer of cumulus oophorus cells, thus mimicking, to some extent, a native cumulus‐oocyte complex. Once generated, this 3D model can be used in a sperm‐binding assay to obtain valuable information about the molecular basis of gamete interaction, since different recombinant proteins can be used to coat the bead surface, thus generating a variety of models to be used for several species. Furthermore, thanks to the ability of the model to decoy sperm, the physiological status of the bound sperm can be studied, making this a powerful tool to select sperm with high fertilizing capacity, to unmask subfertile animals in livestock breeding centers, or for toxicological studies. Here, we describe how to generate and use this model for sperm‐binding assays, using porcine sperm as an example, and ZP2, a protein from zona pellucida, as the recombinant protein of interest. © 2020 Wiley Periodicals LLC. Basic Protocol 1: Generation of the in vitro 3D model Alternate Protocol 1: Binding cumulus oophorus cells to the model Basic Protocol 2: Quality control of the model by SDS‐PAGE electrophoresis and western blot Support Protocol 1: Immunochemistry to confirm proper protein distribution on surface of beads Support Protocol 2: Elution of recombinant conjugated proteins Basic Protocol 3: Sperm‐binding assay Alternate Protocol 2: Sperm preparation by the swim‐up method

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Biological Effects of Recombinant Human Zona Pellucida Proteins on Sperm Function1

Cited by 58 publications

References 63 publications

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

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

Acrosome reaction: relevance of zona pellucida glycoproteins

Sperm‐Binding Assay Using an In Vitro 3D Model of the Mammalian Cumulus‐Oocyte Complex

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