Cloning Of CDNA And Rt-pcr Expression Analysis Of PPARβ In Larimichthys Crocea

PPAR (Peroxisome proliferator-activated receptors), are ligand-activated transcription factors belonging to the nuclear hormone receptor superfamily that functions as critical regulators of lipid and energy homeostasis. There are three PPAR subtypes in mammals, named PPARα, PPARγand PPARβ. Intensively studied about PPARαand PPARγ, but few of reports are heared of PPARβ. The objective of this work was to clone and characterize PPARβsubtypes in the Larimichthys crocea and Plecoglossus altivelis, in order to address its functions and modulation mechanism in fishes.A full-length cDNA of PPARβin Larimichthys crocea was amplified by RT-PCR and SMART RACE method. The cDNA was 3408bp in size, with 146bp 5'-UTR, 1729bp 3'-UTR and 1533bp ORF, encoding a protein of 510 amino acids with a molecular weight of 57.2 kDa and pI 6.46. The sequence analysis indicated that the deduced amino acid sequence of the cDNA shared high identity with other eighteen species, such as Sparus aurata, Dentex dentex, Rachycentron canadum, Dicentrarchus labrax, Lateolabrax japonicus, Pleuronectes platessa, Salmo salar, Takifugu rubripes, Danio rerio, Gallus gallus, Monodelphis domestica, Homo sapiens, Pan troglodytes, Bos taurus, Oryctolagus cuniculus, Sus scrofa, Mus musculus and Xenopus laevis,which is from 68% to 92%, and the highest are 91% with Sparus aurata, Dentex dentex, Rachycentron canadum and Dicentrarchus labrax. The RT-PCR analysis revealed that PPARβmRNA was expressed in all tissues tested, including the muscle, eye, spleen, kidney, liver, heart, gill, intestine and brain. Furthermore, the expression level of PPARβwas the highest in liver than in others, whereas the lowest was in muscle.Partial Plecoglossus altivelis PPARβcDNA sequence, 219bp in size, was amplified by RT-PCR. Two 5'side PPARβsubtypes segments were isolated by 5'SMART RACE and two 3'side segments with whole polyA structure were amplified by 3'SMART RACE. It was distinct that there were two subtypes of PPARβin Plecoglossus altivelis, but the full-length cDNA amplifying was failed. The amino acid sequences analysis suggested that the DNA binding domain is the most conservative district whereas the A/B domain is the least one. The homology analysis shows the fact the predict PPARβin Plecoglossus altivelis, Lateolabrax japonicus PPARβand Larimichthys crocea PPARβwere same subtybes with Danio rerio PPARβ2, Salmo salar PPARβ2A and Salmo salar PPARβ2B, While Danio rerio PPARβ1, Salmo salar PPARβ1A and Salmo salar PPARβ1B belonged to the same subtype.