Cryo‐EM structure of native human uromodulin, a zona pellucida module polymer

Stsiapanava, A.; Xu, Chenrui; Brunati, Martina; Zamora-Caballero, Sara; Schaeffer, Céline; Bokhove, Marcel; Han, Ling; Hebert, Hans; Carroni, Marta; Yasumasu, Shigeki; Rampoldi, Luca; Wu, Bin; Jovine, Luca

doi:10.15252/embj.2020106807

Cited by 36 publications

(63 citation statements)

References 109 publications

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“…Otherwise, the N-glycosylation at Asn-146 of ZP3 may be necessary for a formation of sperm-binding active site between interfaces of the ZP3 fragment and ZP4. Consistent with this hypothesis, a recent 3D model of pig ZP3/ZP4 complex based on the cryo-EM structure of the filaments of uromodulin pointed out the idea that the hinge region of ZP3 is important for its interaction with ZP4 and for the formation of sperm recognition surface [ 26 ].…”

Section: Discussionmentioning

confidence: 92%

Sperm-binding regions on bovine egg zona pellucida glycoprotein ZP4 studied in a solid supported form on plastic plate

et al. 2021

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The zona pellucida (ZP) is a transparent envelope that surrounds the mammalian oocyte and mediates species-selective sperm–oocyte interactions. The bovine ZP consists of the glycoproteins ZP2, ZP3, and ZP4. Sperm-binding mechanisms of the bovine ZP are not yet fully elucidated. In a previous report, we established the expression system of bovine ZP glycoproteins using Sf9 insect cells and found that the ZP3/ZP4 heterocomplex inhibits the binding of sperm to the ZP in a competitive inhibition assay, while ZP2, ZP3, ZP4, the ZP2/ZP3 complex, and the ZP2/ZP4 complex do not exhibit this activity. Here, we show that bovine sperm binds to plastic plates coated with ZP4 in the absence of ZP3. We made a series of ZP4 deletion mutants to study the sperm-binding sites. The N-terminal region, Lys-25 to Asp-136, and the middle region, Ser-290 to Lys-340, of ZP4 exhibit sperm-binding activity. These results suggest that among the three components of bovine ZP glycoproteins, ZP4 contains the major potential sperm-binding sites, and the formation of a multivalent complex is necessary for the sperm-binding activity of ZP4.

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Section: Discussionmentioning

confidence: 92%

Sperm-binding regions on bovine egg zona pellucida glycoprotein ZP4 studied in a solid supported form on plastic plate

et al. 2021

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“…These include the ZP-N subdomain of mouse ZP3 (2.3 Å) [85], full-length chicken ZP3 (2.0 Å) [86], ZP-C subdomain of mouse ZP2 (2.25 Å) [66,89], and homodimers of chicken ZP1 (2.7 Å resolution) [91]. Electron cryo-microscopy has also been used to solve the structure of fibrils of uromodulin, another ZPD protein [92].…”

Section: Zp Protein 3-dimensional Structurementioning

confidence: 99%

Zona Pellucida Genes and Proteins: Essential Players in Mammalian Oogenesis and Fertility

Wassarman

Litscher

2021

Genes

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All mammalian oocytes and eggs are surrounded by a relatively thick extracellular matrix (ECM), the zona pellucida (ZP), that plays vital roles during oogenesis, fertilization, and preimplantation development. Unlike ECM surrounding somatic cells, the ZP is composed of only a few glycosylated proteins, ZP1–4, that are unique to oocytes and eggs. ZP1–4 have a large region of polypeptide, the ZP domain (ZPD), consisting of two subdomains, ZP-N and ZP-C, separated by a short linker region, that plays an essential role in polymerization of nascent ZP proteins into crosslinked fibrils. Both subdomains adopt immunoglobulin (Ig)-like folds for their 3-dimensional structure. Mouse and human ZP genes are encoded by single-copy genes located on different chromosomes and are highly expressed in the ovary by growing oocytes during late stages of oogenesis. Genes encoding ZP proteins are conserved among mammals, and their expression is regulated by cis-acting sequences located close to the transcription start-site and by the same/similar trans-acting factors. Nascent ZP proteins are synthesized, packaged into vesicles, secreted into the extracellular space, and assembled into long, crosslinked fibrils that have a structural repeat, a ZP2-ZP3 dimer, and constitute the ZP matrix. Fibrils are oriented differently with respect to the oolemma in the inner and outer layers of the ZP. Sequence elements in the ZPD and the carboxy-terminal propeptide of ZP1–4 regulate secretion and assembly of nascent ZP proteins. The presence of both ZP2 and ZP3 is required to assemble ZP fibrils and ZP1 and ZP4 are used to crosslink the fibrils. Inactivation of mouse ZP genes by gene targeting has a detrimental effect on ZP formation around growing oocytes and female fertility. Gene sequence variations in human ZP genes due to point, missense, or frameshift mutations also have a detrimental effect on ZP formation and female fertility. The latter mutations provide additional support for the role of ZPD subdomains and other regions of ZP polypeptide in polymerization of human ZP proteins into fibrils and matrix.

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“…The second one proposed by Dean in 2004, describes a ZP formed by repeats of Zp3-Zp2 and Zp3-Zp1 heterodimers that form the main fibrillar structure, being bound through the glycoproteins Zp1 and Zp2 (Dean, 2004 ). The third one, is a variation of the first model, so that the Zp1 glycoprotein is incorporated into the long filaments through its ZP domain; therefore in the mouse, the ZP would be formed by a fibrillar framework constituted by long polymers of Zp1-Zp2-Zp3 which are joined to each other by Zp1 homodimers through disulfide bonds forming a three-dimensional structure (Monné and Jovine, 2011 ; Stsiapanava et al, 2020 ). However, the ZP structure of the species with four proteins remains unproven.…”

Section: Introductionmentioning

confidence: 99%

ZP4 Is Present in Murine Zona Pellucida and Is Not Responsible for the Specific Gamete Interaction

Izquierdo-Rico

Moros-Nicolás

Pérez-Crespo

et al. 2021

Front. Cell Dev. Biol.

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Mammalian eggs are surrounded by an extracellular matrix called the zona pellucida (ZP). This envelope participates in processes such as acrosome reaction induction, sperm binding, protection of the oviductal embryo, and may be involved in speciation. In eutherian mammals, this coat is formed of three or four glycoproteins (ZP1–ZP4). While Mus musculus has been used as a model to study the ZP for more than 35 years, surprisingly, it is the only eutherian species in which the ZP is formed of three glycoproteins Zp1, Zp2, and Zp3, Zp4 being a pseudogene. Zp4 was lost in the Mus lineage after it diverged from Rattus, although it is not known when precisely this loss occurred. In this work, the status of Zp4 in several murine rodents was tested by phylogenetic, molecular, and proteomic analyses. Additionally, assays of cross in vitro fertilization between three and four ZP rodents were performed to test the effect of the presence of Zp4 in murine ZP and its possible involvement in reproductive isolation. Our results showed that Zp4 pseudogenization is restricted to the subgenus Mus, which diverged around 6 MYA. Heterologous in vitro fertilization assays demonstrate that a ZP formed of four glycoproteins is not a barrier for the spermatozoa of species with a ZP formed of three glycoproteins. This study identifies the existence of several mouse species with four ZPs that can be considered suitable for use as an experimental animal model to understand the structural and functional roles of the four ZP proteins in other species, including human.

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Cryo‐EM structure of native human uromodulin, a zona pellucida module polymer

Cited by 36 publications

References 109 publications

Sperm-binding regions on bovine egg zona pellucida glycoprotein ZP4 studied in a solid supported form on plastic plate

Sperm-binding regions on bovine egg zona pellucida glycoprotein ZP4 studied in a solid supported form on plastic plate

Zona Pellucida Genes and Proteins: Essential Players in Mammalian Oogenesis and Fertility

ZP4 Is Present in Murine Zona Pellucida and Is Not Responsible for the Specific Gamete Interaction

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