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

Abstract

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.

Positive selection in gamete interaction proteins inCarnivora

“…These results are consistent with the previously reported pseudogenization events of Zp1 in some carnivores, even-toed ungulates and cetacean species (Moros-Nicolás et al, 2018a;Goudet et al, 2008), the pseudogenization of Zp4 in the house mouse (Mus musculus) (Lefièvre et al, 2004) and in two South American marsupials (common opossum and gray short-tailed opossum) (Moros-Nicolás et al, 2018b), the fact that Zp4 appears to have diverged among vertebrates (Nishimura et al, 2019) and, finally, that ZP4 is not responsible for gamete interaction (Izquierdo-Rico et al, 2021). As Zp4 and Zp1 are recent paralogs that keep the same structure and domains, their functions would at least partially overlap and in those species with just one of these cross-linking proteins, the other would fulfill its function.…”

“…Other events of Zp1 gene pseudogenization are reported in small primates lineages, such as marmosets and tarsiers (Stetson et al, 2012). On the other hand, pseudogenization of Zp4 gene was described in the house mouse ( Mus musculus ) (Lefièvre et al, 2004) and other members of the Mus genus (Izquierdo-Rico et al, 2021). However, ZP3 and ZP2 proteins have not been reported as lacking in any mammalian species.…”

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

“…mZP1, mZP2, and mZP3 genes are located on chromosomes 19 (7.36 cM), 7 (11.3 cM), and 5 (9.2 cM), respectively, vary in length from 6.5 (ZP1) to 18.5 (ZP2) kb, and contain 12 (ZP1), 18 (ZP2), and 8 (ZP3) exons. mZP4 is a pseudogene located on chromosome 13; pseudogenization of mZP4 is restricted to the subgenus Mus [35]. hZP1, hZP2, hZP3, and hZP4 genes are located on chromosomes 11 (11q12.2), 16 (16p12.3-p12.2), 7 (7q11.23), and 1 (1q43), respectively, vary in length from 8.1 (ZP1) to 18.3 (ZP3) kb, and contain 12 (hZP1), 19 (hZP2), 8 (hZP3), and 12 (hZP4) exons.…”

Positive selection in gamete interaction proteins in Carnivora

“…Furthermore, pseudogenisation events affecting Zp1 have been reported in certain lineages of small primates, such as marmosets and tarsiers (Stetson et al, 2012). In contrast, the pseudogenisation of Zp4 gene has been documented in the house mouse (Mus musculus) and other members of the Mus genus (Izquierdo-Rico et al, 2021;Lefièvre et al, 2004). However, it is important to note that ZP3 and ZP2 proteins have not been reported as lacking in any mammalian species, indicating their essential function in gamete interaction and fertilisation (Liu et al, 1996;Rankin et al, 1996Rankin et al, , 2001.…”