Transcriptiome Analysis During Skeletal Muscle Injury Repair And Sdc4 Preliminary Functional Studies In Danio Rerio

Skeletal muscle is a regenerable tissue in animals after a certain degree of injury.Satellite cells are irreplaceable for the growth,development and injury repair of muscle.Injury repair of skeletal muscle is a complex and efficient cascade of interlocking processes.In order to understand the regulatory mechanism of fish after muscle injury,in this study,Danio rerio was used to construct a muscle injury model.Some differentially expressed genes（DEGs）were screened and identified during muscle regeneration based on transcriptome sequencing,and syndecan-4（sdc4）was selected for preliminary functional analysis.The main findings are as follows:（1）The muscle injury model of Danio rerio with three months was constructed using cardiotoxin,and the transcriptome as well as lnc RNA sequencing was performed.RNA-Seq analysis revealed that 2007 DEGs were found,including 1115up-regulated and 892 down-regulated genes.GO-enriched analysis revealed that they were enriched in the regeneration of cellular components,protein binding,enzyme activity and signal transduction.KEGG analysis showed that cytoskeleton remodeling and fatty acid metabolism were the most active in the repair processes.These genes involved in muscle fiber development（myog,tnnc2 and myo19,etc）and cytoskeleton remodeling（tiam1,sdc4 and tuba,etc）may play an important role during the repair process after muscle injury.（2）Lnc RNAs sequencing showed that 901 differentially expressed lnc RNAs was found.The 103 differentially expressed lnc RNAs targeting candidate genes were identified,including 50 up-regulated and 53 down-regulated.Enrichment analysis of target genes of top20 differentially expressed lnc RNAs showed that these target genes associated with myofiber development（foxo1a,tns1b,etc）and cytoskeleton remodeling（pnck,mical3a and nfixa）were significantly enriched,indicating that these lnc RNAs are also involved in the process of muscle injury repair by regulating the target genes.（3）Syndecan-4（sdc4）is the main surface marker gene of satellite cells.The sdc4^-/-mutants were constructed based on CRISPR/Cas9.Two types of mutation were obtained with deletion of 8 bp and 10 bp.The length and weight of mutants were shorter and smaller than the wild type,indicating that sdc4 knockout would have some effects on the growth and development of zebrafish.In addition,bioinformatics-based analysis revealed a binding site for dre-mi R-141 in the 3’UTR region of the sdc4 gene,further study by luciferase reporter system confirmed that dre-mi R-141 could bind to the sdc4 3’UTR to regulate its transcription.