Effects Of MSTN Gene On Muscle Fiber Types In Sheep And Its Cellular And Molecular Mechanisms

Myostatin(MSTN)is a negative regulator of skelet al muscle growth,which specifically inhibits the proliferation and differentiation of skelet al muscle cells and plays an important role in skelet al muscle and fat development.Recent studies have shown that MSTN not only regulates muscle growth and development,but also plays an important role in muscle fiber types.Loss of MSTN can improve the formation of fast muscle(type II)fibers in skelet al muscle.However,the mechanism of MSTN in muscle fiber types remains unclear.In this study,two m,scle(MSTN)gene was introduced into txcel ram(AA)× Altai ewe(GG)cross,and 208 F2 sheep with different genotypes were selected as the experimental population through simultaneous estrus and artificial insemination.Body weight,body size and carcass traits of F2 sheep were measured,and correlation analysis was conducted.Analysis MSTN 3’UTR g+6723G>A;A mutation and the sheep muscle fiber types of relationships,and is verified by sheep myoblast MSTN gene of myoblast differentiation types of influence,to clarify MSTN influence on formation of muscle fiber,explore its action mechanism,revealing the MSTN gene in sheep muscle fiber types and mechanism of the influence of provide theoretical basis for improving the sheep production performance.The full results are as follows:(1)MSTN gene 3’UTR g+6723G>A of crossbred F2 sheep;There were three genotypes of SNP in A,including two alleles of A and G.AG type had the highest genotype frequency,AA type had the lowest genotype frequency,and G was the dominant allele.Association analysis showed that there were differences in body weight,carcass weight,left hip and leg weight and left 5 rib weight between RAMS and ewe(P<0.05);The body weight,carcass weight and left hip and leg weight of AA genotypes were significantly higher than those of GA and GG genotypes(P<0.05),the tail fat of GG genotype was significantly higher than that of AA genotype(P<0.05),there was no significant difference between GA and GG genotypes in tail fat(P>0.05),there were significant differences in birth weight,eye muscle length and hip width between AA and GG genotypes(P<0.05),there was no significant difference in other indexes(P>0.05).(2)The diameters of muscle fibers of longissimus dorsi muscle of F2 generation were significantly different among different genotypes(P<0.01),the cross-sectional area and number of muscle fibers were significantly different among different genotypes(P<0.05).In alkaline condition(p H 9.6),at PASE staining was used to detect type I muscle fibers in light color,type IIB fibers in dark brown,and type IIA fibers in intermediate color between light color and dark brown.AA genotype type I fiber was the least,type IIA fiber was the most,and type IIB fiber was more.GG genotype had the most type I fiber and the least type IIA fiber.AG genotype I and IIB fibers are in the center.The proportion of type I(slow type)muscle fiber was significantly different among different genotypes(P<0.01),the number of type I muscle fiber was significantly different among different genotypes(P<0.05),the number and proportion of type I muscle fiber of AA genotype was the least,and that of GG genotype was the most;The number and proportion of type II(fast muscle)muscle fibers were significantly different among different genotypes(P<0.05),in contrast with type I,type AA had the largest number and proportion of type II muscle fiber,while type GG had the least.(3)The number of My HC type I muscle fibers was significantly higher than that of the Mock control after induction and differentiation of MSTN overexpressed cells.When MSTN was disturbed(MSTN+sh R218),the number of My HC type I muscle tubes was significantly decreased,suggesting that MSTN promoted the generation of type I muscle fibers.After overexpression of MSTN,the expression level of Ca N gene was highly consistent with that of MSTN,while the expression level of My HC I gene was changed by the expression level of Ca N gene,but the expression of My HC II gene was not consistent with that of Ca N,indicating that MSTN did not regulate the generation of type II muscle fibers.MSTN and Si-CAN(interfered Ca N)co-existed in myoblasts,which inhibited differentiation more significantly.When MSTN was interfered,the inhibition of MSTN was relieved,and the fusion rate of differentiated myoduct was increased.This suggests that MSTN may inhibit myoblast differentiation types through Ca N gene signaling pathway,but not through MEF2 C pathway.