Sequence Characteristics and Tissue Expression and Its Effect on Gene Expression of BCAAs Metabolic Enzymes of KLF6 Gene in Siniperca chuatsi
In order to study the sequence characteristics and tissue expression of KLF6 gene in Siniperca chuatsi and its influence on the expression of branchable amino acid metabolizing enzyme gene,bioinformatics analysis of KLF6 gene was conducted through online tools and software.Real-time fluorescence quantitative PCR(RT-qPCR)was used to detect the expression abundance of KLF6 gene in 7 different tissues of the Sinfinfish and the mRNA expression levels of KLF6 gene and BCAAs metabolizing enzyme related genes after siRNA interference.The results showed that the total length of the coding region of the KLF6 gene of S.chuatsi was 3 791 bp(entry number:XM_044176561.1),in which the 5'end and 3'end were 271 bp and 2 659 bp,respectively,and the open reading frame was 860 bp,encoding a total of 286 amino acids.It was most closely related to the KLF6 gene(98.6％)and more distant to the KLF6 gene(82.3％)of carp.KLF6 has one KLF6-N type domain(2-204 aa),one COG5048 domain(194-268 aa)and three Znf-C2H2 domains,which were located between 203-227,233-257 and 263-285 amino acids,respectively.The structure of the S.chuatsi KLF6 protein was composed of random curl(56.99％),α-helix(26.22％),extension chain(10.84％)and β-angle(5.94％).The results of RT-qPCR showed that the relative expression level of KLF6 gene in S.chuatsi tissues was in the following order from high to low:heart,spleen,brain,kidney,intestine,muscle and liver,and the expression level in the center was significantly higher than that in liver(P＜0.01)and muscle(P＜0.05),while the expression level in spleen,brain,kidney and intestine was higher than that in liver(P＜0.05).The mRNA expression of BCAAs metabolizing enzymes(BCKDHb,ACADM,MCCC1,MCCC2,MUT,IVD)was significantly increased after the mRNA expression of KLF6 gene was inhibited(P＜0.05).This study provides a basis for the study of the regulatory mechanism of KLF6 gene in muscle BCAAs metabolism of S.chuatsi.
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