Glycosylated Chicken ZP2 Accumulates in the Egg Coat of Immature Oocytes and Remains Localized to the Germinal Disc Region of Mature Eggs1

Abstract

Vertebrate eggs are surrounded by an egg coat, which is a specific extracellular egg matrix consisting of several glycoproteins with a conserved zona pellucida (ZP) domain. Two mammalian egg coat subunits, ZP2 and ZP3, have been suggested to act as sperm receptors. In bird eggs, however, ZP2 has never been identified in the egg coat of mature oocytes and ovulated eggs. Here we report that chicken ZP2 is expressed in immature small follicles and remains as an egg-coat component locally in the germinal disc region of mature eggs. RT-PCR analysis indicated marked expression of the ZP2 and ZP4 genes in the granulosa cells of immature white follicles, whereas the ZP3 and ZPD genes showed marked expression in the cells of maturing yellow follicles. ZP2 was identified in the egg coat isolated from immature follicles as a heavily N-glycosylated glycoprotein of ∼200 kDa, which was enzymatically converted to a 70-kDa deglycosylated form. Immunoblotting and immunohistological analyses showed that ZP2 was localized around the germinal disc region of mature follicles. ZP2 was accumulated in the egg coat of immature white follicles at the earlier stages of oocyte development and became a minor component in the egg coat of maturing yellow follicles, except for the germinal disc region. Localization of ZP2 in the germinal disc region of mature eggs, where sperm bind to the egg coat at high density, suggests some role for ZP2 in the preferential binding and penetration of sperm in the germinal disc region of bird eggs.

Molecular basis of egg coat cross-linking sheds light on ZP1-associated female infertility

“…9a, c). This observation supports the recent suggestion that cZP4 functionally substitutes cZP1 in the VE of the immature oocyte, which is assembled before cZP1 starts being expressed 30, 34 . Consistent with the role of cZP1 as a major component of the VE of mature oocytes 30,34 , the P/Q-rich repeated region of this subunit (absent in cZP4) may act as a molecular spring that stretches while the egg coat dramatically expands over the course of oocyte maturation.…”

“…Interestingly, the soluble precursor of ZP1 is monomeric, suggesting that the protein only forms intermolecular disulphides upon incorporation into the growing VE 33 . On the other hand, avian ZP4 is synthesized by the ovary and only expressed in limited amounts during the early stage of folliculogenesis, so that it remains largely localized within the germinal disc region of the eggs where ZP2 is also found 34 .…”

Mammalian egg coat modifications and the block to polyspermy

“…This can contain additional subunits that are structurally related to ZP1–4, such as for example ZPY in Xenopus , ZPD in chicken, and ZPAX in amphibian and bird (Frankenberg & Renfree, 2018; Goudet et al, 2008; Lindsay, Wallace, & Hedrick, 2001; Nishio, Okumura, & Matsuda, 2018); on the contrary, in most species of fish, the VE lacks ZP2 (Litscher & Wassarman, 2018). In the case of chicken, the VE subunit composition also varies during oocyte development, so that ZP2 remains concentrated around the small germinal disk region of the mature egg (Nishio et al, 2014, 2018). Moreover, an apparently functional ZPAX orthologue was recently detected in the genome of a marsupial species (Frankenberg & Renfree, 2018).…”

Egg Envelope Glycoproteins ZP1 and ZP3 Mediate Sperm-Egg Interaction in the Japanese Quail

“…ZP2 is expressed in the oocyte of immature follicles and exists in the pvm as a minor component in Japanese quail (Kinoshita et al, 2010). ZP2 in the chicken also accumulates in the egg envelope of immature oocytes but it stays behind in the germinal disk region even after the follicle grows to the largest (Nishio et al, 2014). These authors emphasized that this restricted localization of ZP2 on the germinal disk may link to the sperm perforation preference around the germinal disk region.…”