Molecular cloning and sequence analysis of the cDNA encoding thyroid-stimulating hormone beta-subunit of common duck and mule duck pituitaries: in vitro regulation of steady-state TSHbeta mRNA level.

For better understanding of phylogenetic diversity and evolution of pituitary thyroid-stimulating hormone (TSH) in birds, we have cloned the cDNAs encoding TSH beta subunit (TSHbeta), by reverse transcription-polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE) from two species of domestic ducks, common duck (Tsaiya duck and Pekin duck) (Anas platyrhynchos domesticus) and mule duck (hybrid of male muscovy duck Cairina moschata and female A. platyrhynchos domesticus). The nucleotide sequences of isolated TSHbeta cDNAs of the two species of ducks are identical, with each including 66 bp of 5'-untranslated region (UTR), 402 bp of coding region, and 82 bp 3'-UTR followed by 18 bp poly A tract. The deduced TSHbeta subunit of the ducks contains 134 amino acids consisting of a putative signal peptide of 19 amino acids and a putative mature protein of 115 amino acids. However, the TSHbetas of common duck and mule duck differ from the TSHbeta of muscovy duck in one amino acid at position 97 of the mature protein: isoleucine for common duck and mule duck, and valine for muscovy duck. Our findings thus demonstrate that inter-genus variation of TSHbeta exists in Family Anatidae, and that TSHbeta gene in the mule duck is preferentially transcribed from the maternal genome rather than from the paternal genome. TSHbeta mRNA expressions were investigated by culturing common duck pituitaries with various doses of hormones. Thyrotropin-releasing hormone (TRH) stimulated, while thyroid hormones, triiodothyronine (T(3)) and thyroxine (T(4)), inhibited the TSHbeta mRNA levels, in dose-related manners. The findings thus support that the mode of regulation of TSH gene expression in hypothalamo-pituitary-thyroid axis in birds is similar to that in mammals. Cortisol and corticosterone decreased the steady-state TSHbeta mRNA levels at the pituitary level, in a dose-related manner, the first-time demonstration in vertebrates. The results may suggest that glucocorticoids exert suppressive action directly at pituitary level in modulation of steady-state TSHbeta mRNA level.