Polyrhachis Ants (plyhachis Vicina Roher) The Dopr2 Cloning And Sequence Analysis

The dopaminergic system as the most importment substance in the central nervous system(CNS), may ploy roles in modulating locomotor activity,feelings,and the function of CNS.Dopamine receptors are related to many kinds of nervous or psychical disease.Dopamine receptor could as the best selected drug to treat the diseases.Dopamine receptors belong to the G protein-coupled receptor (GPCR) sub-family showing the highest degree of conservation.Based on amino acid sequence homology,signal transduction pathways,and sensitivity to class-specific agonist and antagonist drugs, the dopamine receptor family has been classified as two major subfamilies:the D1-like receptors(D₁ and D₅) and D2-like receptors(D₂,D₃,and D₄).D1-like receptors are coupled to stimulatory G proteins(Gs) thereby triggering activation of adenylate cyclase.D2-like receptors transduce second-messenger signaling via inhibitory G proteins(Gi),which lead to inhibition of adenylate cyclase and modulation of ion channels.Plyhachis vicina Roger belongs to the genus Polyrhachis(Hymenoptera Formicida).It is a typical eusocial insects characterized by common polymorphism and sophisticated behaviors.The group mostly consists of three castes:female(queen),male and worker.It exists widely in our country and is the only kind of ant for eating and medicinal herds,assigned by Ministry Health of the People's Republic of China since it is abundant in nutrition and officinal values.Consequently,P.vicina is a good material for the studies of the mechanisms of insect development and behavior.In this paper,the cloning and expression of PvDopR2 gene from P.vicina was investigated.The bioinformatics methods are used to analyze characteristics of cDNA and predict functional motifs of amino acids.The results of investigations are offered as follows:1.The total RNA was extracted from the P.vicina;degenerate primers are designed for highly conserved regions with a low level of degeneracy;and the cDNA of DopR2 is obtained by RT-PCR. The length of DopR2 gene is 1381bp,including an open reading frame(ORF) of 1329bp,code the protein that is constituted by 442 amino acids,the molecular weight is 50.3430KD,and theoretical pI is 9.21.2.Amino acids sequence analysis shows that the PvDopR2 belongs to the GPCR sub-family with a typical structure domain of seven-transmembrane helix.The prediction of the secondary structure of PvDopR2 using HNN method,show thatα-helix,irregular and extended coiled are mainly structural elements,only littleβ-sheet are found.The results of tertiary structure prediction using SwissModel First Approach show fiveα-helix domains and the domains of transmembrane helix domains TM Ⅰ-TMⅤare overlapped.A series of predicted function motifs are found in the PvDopR2 protein sequence,including N-glycosylation site,Protein kinase C phosphorylation site,Casein kinaseⅡphosphorylation site,N-myristoylation site and G-protein coupled receptors family 1 signature;most of these sites with functional information are located on these transmembrane helix domains.The result of amino acids sequence analysis shows that these domains are related the functions of the protein and were highly conserved.3.The analysis of molecular evolution indicats that this protein shares a higher lever identity within insects,and the degree of the identity beyond 60%in all known insect DopR2.The sequence identities of the PvDopR2 with the vertebrate and the human being are no more than 30%mostly.The phylogenetic tree based on NJ method show that the PvDopR2 clustered as expected with all insect DopR2.