Effect And Mechanism Of PSIP1 On Proliferation And Differentiation Of Skeletal Muscle Satellite Cells Of Pig

In the pork consumption market in our country,consumers’ demand for the pork quality is growing increasingly.In order to achieve the purpose of high quality development of animal husbandry constantly explores methods and ways to improve meat quality.Skeletal muscle fiber type is an important factor affecting pork tenderness,meat color and fat content,but the molecular network regulating skeletal muscle fiber type remains to be improved.Therefore,this study focused on the regulation of muscle fiber types,in order to find the key regulatory genes in this process,so as to dig out a new regulation mechanism of muscle fiber types.In this study,differentially expressed genes were screened from 5 different skeletal muscle model databases of pig,mouse and human,and conserved analysis was performed to identify their expression patterns in skeletal muscle of pig,analyze biological characteristics of PSIP1 and construct overexpression vector.Through overexpression(pc DNA3.1-PSIP1)and interference(si-PSIP1)tests,to explore whether PSIP1 gene plays a key role in proliferation and differentiation and muscle tube formation of skeletal muscle satellite cells of pig,and explore the mechanism of action of PSIP1-mediated ERK pathway on porcine skeletal muscle fiber types by inhibiting or activating ERK pathway.The main results are as follows:1.PSIP1 promotes proliferation of porcine skeletal muscle satellite cellsThe EDU staining results after overexpression of PSIP1 showed that the proliferative ability of skeletal muscle satellite cells of pig was significantly increased(P < 0.05),the m RNA expression of p21,p27 was significantly increased(P < 0.05),and the protein expression level of Ki67 was also significantly increased(P < 0.05).These results indicated that overexpression of PSIP1 promoted the proliferation of porcine skeletal muscle satellite cells.Interference with PSIP1 inhibited cell proliferation2.PSIP1 promotes porcine skeletal muscle satellite cell differentiationAfter overexpression of PSIP1,the differentiation related genes(MyOG,MyHC,MyOD)were significantly increased(P < 0.01),the protein expression level of MyHC was significantly increased(P <0.05),and the MyHC-positive muscle tubes in immunofluorescence staining was significantly increased(P < 0.05).These results indicated that PSIP1 promoted the differentiation of porcine skeletal muscle satellite cells.RT-qPCR test showed that the expression of MyHC Ⅱ x was significantly increased after overexpression of PSIP1(P < 0.05),and si-PSIP1 also significantly increased the m RNA expression level of MyHCⅡx(P < 0.05),indicating that PSIP1 inhibited the formation of oxidized muscle fibers.3.PSIP1 mediate ERK pathway that promote the differentiation of colytic muscle fibers,inhibited oxidative muscle fiberAfter overexpression or interference with PSIP1,p-ERk /ERK also increased or decreased(P < 0.05).After inhibiting ERK pathway during overexpression of PSIP1,proliferating cell of skeletal muscle was significantly decreased(P < 0.05),and the expression level of Ki67 protein was decreased(P < 0.05).The number of MyHC positive muscle tubes increased,the m RNA expression of oxidized muscle fiber and Slow protein expression were significantly increased(P < 0.05),and significantly decreased Fast protein expression(P < 0.05).These results are contrary to the results of activating the ERK pathway when interfering with PSIP1,suggesting that the PSIP1-mediated ERK pathway promotes the differentiation of glycolic muscle fibers and inhibits the differentiation of oxidative muscle fibers in porcine skeletal muscle satellite cells.In conclusion,at the cellular level,this study found that PSIP1 can promote proliferation and differentiation of skeletal muscle satellite cells of pig by the ERK pathway,and promote the differentiation of glycolic muscle fibers.This study not only clarified the effect of PSIP1 on muscle fiber type transformation of porcine skeletal muscle,but also provided a theoretical basis for improving pork quality.