Genome-wide Remodeling Of H3K27me3 Landscape During Primary Myofibers And Secondary Myofibers Formation In Porcine Skeletal Muscle And The Differentially Expressed Gene Analysis

Pork,which is the main constitution of Chinese food,is in a great increasing requirement because of the great improvement of national living standards.Therefore,it’s of great significance to increase pork production for our life.The pork is mainly composed by muscle,which then can be divided into different types,such as skeletal muscle,smooth muscle and myocardium according to its structure and function.Research on the regulation mechanism of skeletal muscle will help to increase the production of pork meat.The development of porcine skeletal muscle can be divided into two stages according to the formation of myofibers.The primary myofibers begin to form at around 35 days of gestation,and this process lasts until the 60^th day of gestation.The secondary myofibers begin to form at 50^th-60^th day of gestation,and the number fo myofibers stop increasing until the 90^th day after pregnancy while the proliferation peaks at about 75^th day of gestation.The histone modification H3K27me3 has been reported to be involved in th regulation of skeletal muscle,but the particular mechanism in skeletal muscle especially in porcine skeletal muscle is still not clear.In this study,we collected the longissimus dorsi of 33^th days duroc embryos,65^th days duroc embryos and90^th days duroc embryos.We test the H3K27me3 expression in longissimus dorsi of different embryos by Western Blotting,and we test the genome-wide modification of H3K27me3 in longissimus dorsi from different embryos through Ch IP-seq.RNA-seq was used to detect the transcription level of genes in longissimus dorsi.Furthermore,Ch IP-seq and RNA-seq were combined to screen out the genes,which were modified by H3K27me3,that may make important roles during the formation of primary myofibers and secondary myofibers.The results are as follows:（1）The result of Western Blotting showed that H3K27me3 was founded in the longissimus dorsi of 33^th days,65^th days and 90^th days duroc embryos.The H3K27me3 modifications in the longissimus dorsi of 33^th days duroc embryos are lower than that of 65^th days,which are lower than that of90^th days.The result indicates that H3K27me3 may play an important role in the formation of primary myofibers and secondary myofibers.（2）RNA-seq shows that there were total 540 genes significantly express in differences in the longissimus dorsi of 33^th days,65^th days and 90^thdays duroc embryos.By analyzing with Ch IP-seq and RNA-seq collectively,we had found total 329 H3K27me3 modified genes,some of which were involved in PI3K/Akt signaling pathway and MAPK signaling pathway,express differently in the longissimus dorsi of 33^th days,65^th days and 90^thdays duroc embryos.（3）By selecting the H3K27me3 modified genes that decreasing in the formation of primary myofibers and secondary myofibers while the transcription expression of these genes increasing in the same time,we found total 110 genes.Based on published research results,we finally selected 3genes from PI3K/Akt signaling pathway and MAPK signaling pathway:MAP3K14,RASGRP1 and ITGB6,and we test these genes with Ch IP-q PCR and RT-q PCR.As the result of RT-q PCR,the transcription expression of these genes increased significantly during the formation of primary myofibers and secondary myofibers.The Ch IP-q PCR showed that the expressions of H3K27me3 modified RASGRP1 and ITGB6 in skeletal muscle of 90^th days duroc embryos lower than that of 65^th days,which were lower than that of 33^thdays.However,the expressions of H3K27me3 modified RASGRP1 and ITGB6in skeletal muscle of 90^th days duroc embryos lower than that of 65^th days,which were lower than that of 33^th days.This study was a preliminary study about H3K27me3 on regulation of skeletal muscle,and the results of this study provide new concept and candidate genes to make a further research on the regulation of skeletal muscle.