Identify M~6A-methylated Candidate Genes Impacting On Embryonic Skeletal Muscle Development In Yellow Chicken

To meet the demands of yellow chicken industry on improving our knowledge on the genetic mechanism on meat production and molecular breeding,in this study,we used m RNA and m~6A sequencing technology to detect the gene expression and m~6A modified profiles during the embryonic skeletal muscle development,resulting in more understand in the molecular regulatory mechanism on controlling skeletal muscle growth at the transcriptional and post-transcriptional level,and a number of major genes or molecular pathways identified to be related to meat production performance.The main findings are as following:(1)Based on the morphological characterization and dynamic gene expression patterns of the skeletal muscle tissues collected from yellow chicken embryo at their11,13,15,18 and 21 day old,two significantly development stages were divided,including the early stage of development(11-15 embryonic age)and late stage of development(18-21 embryonic age),which exhibited significant differences in the the muscle fiber formation and gene expression.(2)Through the analysis of transcription sequencing data,comparing to the expression data at the 11 embryonic age,a total of 1786 differentially expressed genes were found from the five timepoint,and 118,294,1157 and 1275differentially expressed genes for 13,15,18 and 21 embryonic age,respectively.Total 354 differentially expressed genes were identified in the early stage of development,and 763 were indentified in the late stage of development.(3)5392,3215,5756,3971 and 5,404 m~6A modified genes were detected at the different timepoints,respectively,and the modified genome regions were mostly nearby the transcription starting and terminal points.(4)Combining analysis on the gene expression changes and m~6A methylated regions,543 genes were identified to be regulated by m~6A epigenetic modification.Through the functional enrichment analysis via the DAVID system,total 11 signal pathways were highly enriched,including Wnt signaling pathway,Notch signaling pathway,Fox O signaling pathway and so on.Also,a number of candidate genes were identified to be regulated by m~6A methylation,including cell cycle factors(CCND1 and CDKN1A)and transcription regulatory factors(FOXO3 and FOXO6),key factors on insulin signaling pathways(PDPK1,IRS1,IRS4 and PIK3CA),and key regulatory factors on TGF signaling pathways(TGFB2,TGFB3 and TGFBR2).(5)Comparing the changes of gene expression and m~6A modification in the early stage and late stage,total 10 m~6A modified genes was identified in the late stage of development with significantly reduced in the gene expression,and 16 m~6A modified genes were identified in the late stage of development with significantly increased in the gene expression.In particular,the muscle-specific transcription regulatory factor,Myo G,were found to be modified by m~6A only in the early stage of development,which may be one of the key reason on the expression of Myo G with significantly higher level in the early stage than the late stage of development.This study firstly revealed the m~6A methylation profile in embryonic skeletal muscle tissue at the whole genome level,and several candidate genes identified to control the growth of skeletal muscle,which provided a novel insight on the genetic regulation mechanism of meat production performance,and would improve the development and application of molecular breeding technology for meat production performance in yellow chicken production.