Molecular cloning, gene structure, molecular evolution and expression analyses of thyrotropin-releasing hormone receptors from medaka (Oryzias latipes).

Molecular cloning of thyrotropin-releasing hormone receptors (TRHR) was performed in a model teleost fish, medaka (Oryzias latipes). Four subtypes of TRHR were cloned and named them as TRHR1a, TRHR1b, TRHR2 and TRHR3 based on their similarity to known TRHR subtypes in vertebrates. TRHR1a, TRHR1b, TRHR2, and TRHR3 of medaka encode 416, 398, 451, and 386 amino acid residues, respectively. Comparison of cDNA sequences of medaka TRHR subtypes with respective genomic DNA sequences revealed gene structures: TRHR1a, TRHR1b and TRHR3genes consist of two exons while the TRH2 gene consists of five exons. Molecular phylogenetic analyses depicted the molecular evolution of TRHR in vertebrates: From the ancestral molecule, TRHR2 diverged first and then TRHR1 and TRHR3. Reverse transcription-polymerase chain reaction analyses revealed the sites of TRHR expression: Expression of TRHR1, TRHR1b and TRHR2 subtypes has been confirmed in the brain, pineal organ, retina and pituitary gland. In addition, TRHR1b is expressed in spleen, digestive tract and skin, and TRHR2 in testis, ovary and gill. TRHR3 is widely expressed in various tissues. These results indicate that in medaka, TRH might exert multiple functions mediated by different TRHR subtypes expressed in each tissue.