Expression, Identification And Location Of GpHR Gene From Amphioxus Branchiostoma Belcheri Tsingtauense

The glycoprotein hormone receptors are classified as members of the G-coupled protein receptor (GPCR) super-family of membrane receptors. The importance of GpHRs in mammals is primarily related to control of the development and function of gonadal and thyroid glands via two main endocrine pathways:hypothalamus-pituitary-gonadal (H-P-G) axis and hypothalamus-pituitary-thyroid (H-P-T) axis. Amphioxus, a cephalochordate, has long been regarded as the extant invertebrate most closely related to the proximate ancestor of vertebrates. Studies on the gene in amphioxus will contribute to the understanding of the origin and evolution of vertebrates.We have cloned the GpHR gene of amphioxus (Branchiostoma belcheri tsingtaunese) by the methods of RT-PCR and RACE. The GpHR gene contains an open reading frame of 1377bp coding a polypeptide which molecular mass of approximately 50kDa and contains 459amino acid residues. It is between vertebrates and invertebrates in the phylogenetic tree, shows that Amphioxus GpHR is still in a primitive state. GpHR has been successfully expressed in Pichia pastoris and Escherichia coli. Western blot analysis confirmed that amphioxus has a native molecular mass of approximately 50 kDa. Immunohistochemical studies on adult amphioxus were performed by using rabbit anti-TSHR.The results showed that GpHR was localized in endostyle, ovary, testicular. And, Immunohistochemical studies on adult amphioxus were performed by using rabbit anti-FSHR and rabbit anti-LH/CG receptor. The results showed that GpHR was localized in ovary. In situ hybridization shows that GpHR gene has expression location at endostyle, ovary, testicular. Finally, the initial function of GpHR is discussed.