Gene Cloning Of PGC-1α And Expression Research In Grass Carp, Ctenopharyngodon Idellus

Peroxisome proliferator-activated receptor-γcoactivator-1α(PGC-1α) plays a very important role in regulating energy metabolism. It involved in various metabolic process,such as adaptive heat production, glucose uptake, hepatic gluconeogenesis, hepatic fatty acid beta oxidation, adipocyte differentiation, muscle type conversion etc.It's well known that adipose tissue development in grass carp abdominal cavity is excessive. It would affect the growth of grass carp when adipose tissue overdevelopment in abdominal cavity. Some researchers found that high unsaturated fatty acid (HUFA) can inhibit lipid accumulation. This work studied the mRNA changes expression of PGC-1α, PPARs, and LPL after treated with HUFA in grass carp both in vivo and in vitro.In this experiment, grass carp were used as the experimental animal. PGC-1αgene was cloned in grass carp, and the pattern of tissue expression was investigated in grass carp. The influences of HUFA on genes expression of PGC-1αwere also determined. The main resultswere as follows: 1. The length of PGC-1αcDNA sequence in grass carp cloned by PCR technology was 612bp (GenBank number: HM015283). Homologous alignment analysis using Clustal W software showed that PGC-1αhad 75%-97% homology to that of other animals.2. The expression of PGC-1αmRNA in ten tissues was investigated by Real Time PCR. The results showed that PGC-1αgene was widely expressed in ten tissues of grass carp. The expression level of PGC-1αwas significantly highest in kidney and heart compared with others, and the expression level was the lowest in adipose tissue, spermary and spleen.3. After six days'feeding with diet contained HUFA, the PGC-1αexpression level in grass carp was significantly increased, then decreased quickly in the following days. The variation tendency of lipid content in hepatopancreas is droped at the beginning then bounces back; which is opposite to the gene expression. Our results indicated that PGC-1αexpression level could be regulated by HUFA. The expression pattern of PGC-1αinduced by dietary HUFA could be rising at first then going down back to the normal level. 4. Preadipocytes were treated with 0μmol/L (control group), 50μmol/L, 100μmol/L and 200μmol/L EPA or DHA. Cell proliferation was analyzed by MTT assay. The degree of adipogenesis and differentiation were measured by Oil Red O staining extraction assay and the expression of PGC-1αmRNA were detected by Real Time PCR. The results showed that: EPA can promote cell proliferation before 48h, then cell differentiation of adipocytes were inhibited at the beginning (24h). EPA and DHA can markedly decrease the expression of PGC-1αafter 48h treated with HUFA, which may be in part associated with its effect on decreasing the differentiation of adipocytes.In conclusion, our dates suggested that HUFA may have effect on grass carp lipid accumulation through PGC - 1αregulates the mitochondrial generation.