Cloning、tissue Expression And Bioinformatics Analysis Of CDNA Encoding FoxO Gene In Chinese Giant Salamander(Andrias Davidianus)

FoxO(Forkhead box O)gene is a subfamily member of Fox gene family and generally exists in both vertebrates and non-vertebrates.In this study,FoxO1,FoxO3 and FoxO3 gene c DNA complete sequence and FoxO6 gene partial c DNA sequences of Chinese giant salamander are cloned by reverse transcription polymerase chain reaction(RT-PCR)and rapid amplification of c DNA ends(RACE),and their biological characteristics are also analyzed.Analysis of Real-time PCR is performed to examine the expression in various tissues and development stage of the Chinese giant salamander.The following results have been obtained:1.Chinese giant salamander FoxO1,FoxO3 and FoxO4 gene c DNA sequences contain 2663 bp,2107bp and 2599 bp and encode 615 aa,640 aa and 548 aa,respectively.FoxO6 gene partial c DNA sequence of Chinese giant salamander contains 1894 bp and encodes 583 aa.Chinese giant salamander FoxO protein contains a FH domain with 91 aa and the tertiary structure prediction shows they are composed of threeα-helix,threeβsheets and a minor coilη1.There are multiple phosphorylation sites in Chinese giant salamander that are identical to mammals,and several large segments missing compared with bony fish,which may be the deletion of useless sequences in evolutionary process.cytoplasmic localization regulates and controls of Chinese giant salamander FoxO protein plays an important role.Highly conserved regions of Chinese giant salamander Fox transcription family FH domain is α3,and FoxO protein FH domain inserts 5 aa comparing with all FH domain Chinese giant salamander.2.FoxO1 gene is the ancestor sequence of FoxO genes in vertebrates,and FoxO has gone through positive selection in process of evolution,which may be of great importance to produce new genes and function in process of evolution.The phylogenetic tree shows that Chinese giant salamander FoxO gene is in a unique evolutionary position,that FoxO1 is in lower similarities with fish,but that FoxO3 is gathered together with fish and that FoxO4 has greater similarities with amphibian.All FH domain of Chinese giant salamander is highly homologous with other species,and Fox N/R may be ancestral gene.3.The expression of FoxO gene in China is characterized by tissue specificity,spatio-temporal specificity and male and female differences.In most cases,the expression of female is higher than that of male in deferent tissues,especially in ovary,and it is speculated that FoxO1 and FoxO6 gene may play an important role in early ovarian development and FoxO3 gene may be involved in regulating the closure and activation of follicles.FoxO4 gene may be the main functional gene of FoxO gene in Chinese giant salamander liver tissue.FoxO1 and FoxO4 genes are involved in regulating the main genes of oxidative stress in the kidney.FoxO3 gene is the main functional gene involved in immune regulation in the spleen of Chinese giant salamander.