Cloning, Sequence Analysis And Expression Of Musiclin CDNA In Pig

Accumulating evidences exist that skeletal muscle acts as an endocrine organ modulating hepatic glucose production and white adipose tissue lipolysis. such as production of interleukin-6 (IL-6) during exercise. Recently the people using an efficient signal sequence trap (SST) method from the 1812 cloning technology in almost a screened only in the expression of skeletal muscle in mice with a single signal sequence without cross-domain model of the unknown protein, was named "Musclin". Insulin can induce Musclin mRNA expression and the transfer Musclin Adenovirus can cause mice fat tissue volume decreased significantly, and lipid metabolism in an important transcription factor Foxo1 can significantly lower Musclin mRNA expression, to speculate Musclin gene in lipid metabolism also has important The regulation of, suggesting that skeletal muscle could be Musclin secretion of sugar and fat metabolism in the activity of another factor. Musclin for control of Health revealed the molecular mechanism of fat and lean pig breeding eventually provide a theoretical basis. The present study examined the distrution of pig,we cloned the pig Musclin gene and analyzed structure and characteristics of Musclin by bioinformatics. Levels of Musclin mRNA in different tissues, at different ages ,in adipose muscle tissues of different breeds, primary adipocytes and co-culture cells (adipocyte and muscle satellite cell), muscle satellite cell were measured by Semi Quantitative Reverse Transcription-Polymerase Chain Reaction ( SQ RT-PCR). Correlation were studied between mRNA level of musclin and that of lipogenetic genes including lipoprotein lipase(LPL), fatty acid synthetase (FAS), peroxisome proliferator-activated receptorγ(PPARγ), triacylglycerol hydrolase(TGH).and specificity genes of skeletal muscle including Myf5 and MyoG. It may plays an important role in lipid metabolism. The main results were summarized as following:1.Successfully cloned 374bp of the pig Musclin gene, Musclin pigs and mice muscle gene sequences were consistent Musclin cDNA structure has a serine protease cleavage point - KKKR, homologous sequences than on that pig Musclin gene sequence and rabbits, cattle, sheep and The identity of more than 85 percent, while the identity of chicken and only 59 percent of that in the course of evolution Musclin conservative poor.2.Musclin hydrophilic proteins are unstable protein, to the Rulesαspiral and non-curly-based structure, inclusion of some middle Extended strand of protein can be identified Musclin hybrid protein. Mainly concentrated in the secretion of ways, the mitochondria in the small, is secreted protein and 1-26 in the first amino acid sequence is a signal peptide, between 26 and 27 there is a restriction site. The N-terminal No. 3-44 , 81-93 and 111-131 maybe the predominant epitopes of the B cell. This study would be helpful for research of mmunological function of Musclin.3.The highest of Musclin expression levels were found in visceral adipose tissues and subcutaneous adipose tissues and the lowest expression in spleen among all other tissues(P<0.05); the expression of musclin mRNA in muscle and subcutaneous adipose tissue were decreased significantly during aging (P<0.05); in the same ages, the levels of Musclin mRNA in subcutaneous adipose and muscle of lean-type pig was significantly higher than that of obesity-type pig ( P<0.05); and the express of Musclin was negative correlated and positively correlated with FAS and PPARγrespectively (P<0.05). However,it had no significant correlation with TGH ( P>0.05). So, we concluded that pig Musclin mRNA was expressed in the muscle as well as other tissues such as adipose tissue. The express of porcine Musclin gene was related to age, breed and FAS, PPARγ. it may plays an important role in lipid metabolism4.Musclin in the primary fat cells of a time-dependent reduction in the cultured cells Musclin expression was significantly higher than the same period in a separate training of primary fat cells (P <0.01). But lower than the same period a separate culture of skeletal muscle satellite cells, the expression of 10 days to achieve significant levels (P> 0.01). Expression of Musclin mRNA in cell had significant correlation with and LPL, PPARγ, Myf5, MyoG. Musclin speculate may be involved in regulating fat synthesis, but we have to further study this mechanism.