Recombinant Hamster Oviductin Is Biologically Active and Exerts Positive Effects on Sperm Functions and Sperm-Oocyte Binding

Abstract

Studies carried out in several mammalian species suggest that oviductin, also known as oviduct-specific glycoprotein or OVGP1, plays a key role in sperm capacitation, fertilization, and development of early embryos. In the present study, we used recombinant DNA technology to produce, for the first time, recombinant hamster OVGP1 (rHamOVGP1) in human embryonic kidney 293 (HEK293) cells. rHamOVGP1 secreted in the culture medium was purified by affinity chromatography. The resulting protein migrated as a poly-dispersed band of 160-350 kDa on SDS-PAGE corresponding to the molecular mass of the native HamOVGP1. Subsequent mass spectrometric analysis of the purified rHamOVGP1 confirmed its identity as HamOVGP1. Immunocytochemistry demonstrated binding of rHamOVGP1 to the mid-piece and head of hamster sperm and to the zona pellucida (ZP) of ovarian oocytes. In vitro functional experiments showed that addition of rHamOVGP1 in the capacitation medium further enhanced tyrosine phosphorylation of two sperm proteins of approximately 75 kDa and 83 kDa in a time-dependent manner. After 3 hours of incubation in the presence of rHamOVGP1, a significant increase in acrosome reaction was measured. Pretreatment of either sperm or oocyte with 20 μg/ml of rHamOVGP1 prior to sperm-egg binding assay significantly increased the number of sperm bound to the ZP. Addition of rHamOVGP1 in the medium during sperm-egg binding with either oocyte or sperm pretreated with rHamOVGP1 also saw an increase in the number of sperm bound to ZP. In all experimental conditions, the effect of rHamOVGP1 on sperm-oocyte binding was negated by the addition of monoclonal anti-HamOVGP1 antibody. The successful production and purification of a biologically active rHamOVGP1 will allow further exploration of the function of this glycoprotein in reproductive function.

The Trithorax group protein dMLL3/4 instructs the assembly of the zygotic genome at fertilization

“…We observed that the dMLL3/4-regulated glycosyl hydrolase IDGF4 has a mitogenic effect on the early embryo. Despite being important for fertilization and early embryogenesis across different mammalian species, the exact function of OVGP1 remains elusive [52]. OVGP1 localizes to the oviduct-the site of mammalian fertilization-where it has been shown to be a component of the extracellular matrix of ovulated oocytes and early embryos [50,51].…”

“…OVGP1 localizes to the oviduct-the site of mammalian fertilization-where it has been shown to be a component of the extracellular matrix of ovulated oocytes and early embryos [50,51]. Despite being important for fertilization and early embryogenesis across different mammalian species, the exact function of OVGP1 remains elusive [52]. Accordingly, OVGP1 has been associated with processes as wide-ranging as sperm capacitation and blastocyst formation [53][54][55].…”

Sperm selection in the female mammalian reproductive tract. Focus on the oviduct: Hypotheses, mechanisms, and new opportunities

“…Immediate and dramatic control of sperm physiology within the oviduct can be exerted by local concentrations of small molecules such as bicarbonate [103], calcium, [104,105] and nitric oxide [106] as well as by secreted proteins [107]. However, there is little to suggest at present that these effects are preferentially exerted on sperm subpopulations with specific genetic properties.…”

Identification of Rabbit Oviductal Fluid Proteins Involved in Pre‐Fertilization Processes by Quantitative Proteomics

“…However, only a few ODF proteins, in particular glycoproteins, such as oviduct‐specific glycoprotein (OVGP1), have been utilized as potential supplements to support and/or control the in vitro processes that precede fertilization . Supplementation of the media with recombinant OVGP1 modulates human and hamster spermatozoa capacitation in vitro. Likewise, lactoferrin (LTF) might be added to promote the capacitation of spermatozoa …”