Molecular cloning and regulation of mRNA expression of the thyrotropin β and glycoprotein hormone α subunits in red drum, Sciaenops ocellatus

Abstract

Full-length cDNAs for thyrotropin β (TSHβ) and glycoprotein hormone α (GSUα) subunits were cloned and sequenced from the red drum (Sciaenops ocellatus). The cDNAs for TSHβ (877 bp) and GSUα (661 bp) yielded predicted coding regions of 126 and 94 amino acid proteins, respectively. Both sequences contain all invariant cysteine and putative glycosylated asparagines characteristic of each as deduced by comparison with other GSUα and TSHβ sequences from representative vertebrate species. Multiple protein sequence alignments show that each subunit shares highest identity (79% for the TSHβ and 86% for the GSUα) with perciform fish. Furthermore, in a single joint phylogenetic analysis, each subunit segregates most closely with corresponding GSUα and TSHβ subunit sequences from closely related fish. Tissue-specific expression assays using RT-PCR showed expression of the TSHβ subunit limited to the pituitary. GSUα mRNA was predominantly expressed in the pituitary but was also detected in the testis and ovary of adult animals. Northern hybridization revealed the presence of a single transcript for both TSHβ and GSUα, each close in size to mRNA transcripts from other species. Dot blot assays from total RNA isolated from S. ocellatus pituitaries showed that in vivo T3 administration significantly diminished mRNA expression of both the TSHβ and GSUα subunits and that goitrogen treatment caused a significant induction of TSHβ mRNA only. Both TSHβ and GSUα mRNA expression in the pituitary varied significantly in vivo over a 24-h period. Maximal expression for both TSHβ and GSUα occurred during the early scotophase in relation to a peak in T4 blood levels previously documented. These results suggest the production of TSH in this species which may serve to drive daily cycles of thyroid activity. Readily quantifiable, variable, and thyroid hormone-responsive pituitary TSH expression, coupled with previously described dynamic daily cycles of circulating T4 and extensive background on the growth, nutrition, and laboratory culture of red drum, suggests that this species will serve as a useful model for experimental studies of the physiological regulation of TSH production.