Abstract
Egg envelopes (chorions) in medaka, Oryzias latipes, are composed of three major glycoproteins: ZI-1, − 2, and − 3. These gene-encoded chorion glycoproteins are expressed in the liver and/or ovarian oocytes of sexually mature female fish. In medaka, the glycoproteins produced in the female liver are induced by estrogen as Choriogenin (Chg.) H and Chg. H minor (m), which correspond to the zona pellucida (ZP) B (ZPB) protein in mammals, and Chg. L, which corresponds to ZPC in mammals. Chg. H, Chg. Hm, and Chg. L, are then converted to ZI-1, − 2, and − 3, respectively, during oogenesis in medaka ovaries.In the present study, we established a medaka line in which the chg.l gene was inactivated using the transcription activator-like effector nuclease (TALEN) technique. Neither intact chg.l transcripts nor Chg. L proteins were detected in livers of sexually mature female homozygotes for the mutation (homozygous chg.l knockout: chg.l−/−). The chg.l−/− females spawned string-like materials containing “smashed eggs.” Closer examination revealed the oocytes in the ovaries of chg.l−/− females had thin chorions, particularly at the inner layer, despite a normal growth rate. In comparing chorions from normal (chg.l+/+) and chg.l−/− oocytes, the latter exhibited abnormal architecture in the chorion pore canals through which the oocyte microvilli pass. These microvilli mediate the nutritional exchange between the oocyte and surrounding spaces and promote sperm-egg interactions during fertilization. Thus, following in vitro fertilization, no embryos developed in the artificially inseminated oocytes isolated from chg.l−/− ovaries. These results demonstrated that medaka ZI-3 (Chg.L) is the major component of the inner layer of the chorion, as it supports and maintains the oocyte’s structural shape, enabling it to withstand the pressures exerted against the chorion during spawning, and is essential for successful fertilization. Therefore, gene products of oocyte-specific ZP genes that may be expressed in medaka oocytes cannot compensate for the loss Chg. L function to produce offspring for this species.
Highlights
In most animals, the egg envelope has several functions, including attraction and activation of spermatozoa, prevention of polyspermy at fertilization, and protection of the developing embryo [1, 2]
In 1991, using specific antibodies, Murata et al discovered, in medaka, other high molecular weight chorion glycoproteins were produced in the liver of spawning females [13]. These results suggested, in medaka, all major components of the chorion were produced in the liver of spawning females
Design and construction of transcription activator-like effector nuclease (TALEN) Using TALE-NT 2.0 software [42], the potential TALEN target sites were searched with parameters including (1) a spacer length of 14–17, (2) a repeat array length of 15–18, and (3) an upstream base of T only
Summary
The egg envelope has several functions, including attraction and activation of spermatozoa, prevention of polyspermy at fertilization, and protection of the developing embryo [1, 2]. Oryzias latipes, there are two structural parts to the layers of the egg envelope — a thin, high-density outer layer and a thick, low-density, multiple-component inner layer. The formation of the outer layer begins during the perinuclear oocyte stage. This step is followed by the formation of the inner layer during the previtellogenic oocyte stage [3]. The terminology of the extracellular matrix (egg envelope) surrounding oocytes differs among groups of animals. In fish, it is the chorion, and in mammals, it is called the zona pellucida (ZP [4];; Fig. 1)
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