Molecular Cloning And Nucleotide Sequence Analysis Of The Yak Melatonin Synthetase Genes: AA-NAT And HIOMT

Melatonin (MLT) is the nerve hormone that plays a major role in regulating the animal seasonal breeding. MLT is synthesized through four-step enzymatic pathway. Arylalkylamine N -acetyltransferase (AA-NAT) and hydroxyindole-O-methyltransfer ase (HIOMT) are critical enzymes in MLT biosynthesis. AA-NAT controls the circadian rhythm speed of MLT secretion, then, HIOMT controls the fluctuation amplitude of MLT secretion.It has been confirmed that the daily rhythmic synthesis of MLT in vertebrate species is controlled by the rhythmic enzymatic activity and mRNA levels of both AA-NAT and HIOMT. Molecular cloning of the AA-NAT gene and HIOMT gene in many species has been reported. While, the yak living in qingzang plateau is kinds of the animal with typical seasonality, the sequences of AA-NAT gene and HIOMOT gene in yak have not been reported now.Studying on it may contribute to explaining the mechanism of molecular regulation in seasonality empathema of yak, which could supply theory evidence. This study apply the mRNA sequences of Bos taurus AA-NAT gene and HIOMT gene by comparative genomics theory, we obtained the corresponding complete CDS sequences in yak by RT-PCR method. The results show that:(1) the homology of nucleotide sequences between Bos taurus and yak is 98.72% and 89.19%, which encoding 207 and 304 amino acid residues respectively, the homology of amino acid sequences between Bos taurus and yak is 99.04 % and 86.96%.(2) Comparison the AA-NAT gene nucleotide sequence between the yak and Bos taurus, we detected 8 variations, which resulted in 2 variations in amino acid sequence in yak, one is in Casein KinaseⅡbinding sites, the 43rd amino acid Lys(K)→Glu (E); The other is in putative acetyl CoA binding motif A site, the 128th amino acid, His(H)→Arg(R). While in these two sites are consistent with human, sheep, rat, chicken amino acids.(3) Comparison the HIOMT gene nucleotide sequence between the yak and Bos taurus, we detected 16 variations, while 11 variations occurred in coding region, which resulted in 4 variations in amino acid sequence in yak. In the meanwhile, we detected 123bp deletion in coding region, subsequently, we confirmed it was alternative splicing variant by genomic DNA PCR method, this deletion was exon 7 in yak by comparison analysis, and it retains the last codon.