Expression And Function Of Chromatin Remodeling Protein INO80 During Preimplantation Development Of Pig Embryos

Pigs are both important economic animals and ideal model animals.Pre-implantation embryo death is one of the main reasons leading to low fertility in pigs.Therefore,studying the mechanism of early embryonic development in pigs is of great significance to animal husbandry,medicine and basic research.The chromatin remodelling protein INO80 affects cell growth and development by regulating genomic stability and gene expression.At present,it has not been reported whether INO80 is involved in the regulation of early embryonic development in pigs.This article aims to use the pig's main organs and pre-implantation embryos as experimental materials to reveal the expression patterns and functions of INO80 in pig growth and development.Bioinformatics analysis of human,mouse,rat and pig INO80,INO80 B,INO80C,INO80 D,INO80E shows that pig INO80 was highly conserved with humans,mice and rats,and the amino acid sequence homology reach 97.51%,.96.42% and 96.42%,respectively;phylogenetic tree shows that the genetic relationship between pig INO80 and human INO80 is closely related;the space-time expression profile of pig INO80 is studied using qPCR,WB and IF methods and INO80 is expressed in the porcine major organs and PA embryos at the transcription level and the protein level;the RNAi method specifically knocks down the expression of INO80 in porcine PA embryos without affecting the expression of several subunits of IN080.The knockdown efficiency in 8 cells,morula and 5d-Blastocyst at the transcription level is 47.85%,80.68%,and 86.75%,respectively.At the protein level the knockdown efficiency of INO80 in 5d-Blastocyst is 69.03%.Phenotypic observation of knockdown pig PA and IVF embryos,it is found that compared with Uninjected and Sham-water control groups,the ratio of blastocyst in SiINO80 mixed-fragment group was significantly decreased.In the 5d-Blastocyst embryos,?-H2 A.X protein expression was not significantly affected with the knockdown of INO80,while SOX2 protein expression is significantly reduced by 48.77%.In conclusion,INO80 is widely involved in the growth and development of major organs of pigs and pre-implantation embryos;the pluripotency of the blastocyst and the separation of the cell lineage may be maintained through the pluripotent transcription factor SOX2,which in turn affects the formation of porcine blastocysts.