The Cold-adaptation Research Of Zona Pellucida Protein In Cold Water Fishes

Freezing avoidance is an essential trait for Antarctic notothenioids to survive subzero temperatures. The effect is known to be conferred by the Antifreeze glycoproteins(AFGPs). Being deficient of AFGP expression in situ, the mechanism by which notothenioid eggs hatching in an icy environment avoid freezing is unknown. The zona pellucida(ZP) protein gene family, encodes the egg chorion which isinvolved in sperm-egg recognition during fertilization, was previously found to be specifically amplified with regard to multiple ZP types in Antarctic notothenioids.Here, we report an unexpected ice-melting function of Antarctic notothenioid ZP proteins(Annoto ZPs) that promotes ice melt at temperatures lower than colligative melting points. All seven of the Annoto ZP types expressed in vitro exhibited ice-binding activity, with a type-specific non-colligative freezing and melting point depression(FMPD).Similar to the Antarctic notothenioids, Schizothorax prenanti(SP) belongs to cold water fishes that its living quarter has a long winter freeze time and summer temperature is very low. So, whether the ZP proteins of Schizothorax prenanti have the same function? The main content of this study is cloning of zona penucida 3 genes coding regions from the total RNA of Schizothorax prenanti(SP) by RT-PCR, and build the gene expression vector, then express in the Chinese hamster ovary cells(CHO-K1), after isolate and purify zona pellucida 3 protein, and then we do crystals experiments, the results show that SP_ZP3 ice crystal shape is six prism, prove SP_ZP3 protein can combine with ice crystals and change the growth form of ice crystals, this illustrate the protein have similar function with Antarctic notothenioids ZPs. But unlike Antarctic notothenioids ZPs, its melting point solution to SP_ZP3 protein has not been significantly decreased.ZP is the abbreviation of zona penucida. ZP protein is a kind of protein with the ZP domain, exists in outer membrane of all vertebrates that from the lower organisms(e.g. C.elegans) to higher mammals. The gene of zona penucida engraves the genetic evolutionary history. The protein called the chorionin in fish and the zona pellucida protein in higher mammal. ZP protein as an important composition of the oocyte plasma membrane involved in the process of fertilization and studies show that genes involved in this process evolve faster than other genes. Antarctic Notothenioids ZP gene in cold conditions of a large number of amplification, is the important mechanism of the Antarctic fish to adapt to the extreme cold environment at the antarctic waters, ZP gene amplification and accelerated the evolution of the ZP gene, make its evolved into a kind of cold stress proteins, adapt the Antarctic fish to the extreme cold of the water environment. But this research shows that SP_ZP3 protein have ice crystals binding activity, this activity may be a primitive function of ZP proteins in fish living in cold environment, in the Antarctic Notothenioids, this functionality in high strength cold pressure will be continued to enlarge, have acquired an ice-melting promotionactivity.