Molecular cloning and regulation of mRNA expression of the thyrotropin and glycoprotein hormone subunits in red drum, Sciaenops ocellatus.
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abstract
Full-length cDNAs for thyrotropin (TSH) and glycoprotein hormone (GSU) subunits were cloned and sequenced from the red drum (Sciaenops ocellatus). The cDNAs for TSH (877 bp) and GSU (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 GSU and TSH sequences from representative vertebrate species. Multiple protein sequence alignments show that each subunit shares highest identity (79% for the TSH and 86% for the GSU) with perciform fish. Furthermore, in a single joint phylogenetic analysis, each subunit segregates most closely with corresponding GSU and TSH subunit sequences from closely related fish. Tissue-specific expression assays using RT-PCR showed expression of the TSH subunit limited to the pituitary. GSU 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 TSH and GSU, 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 TSH and GSU subunits and that goitrogen treatment caused a significant induction of TSH mRNA only. Both TSH and GSU mRNA expression in the pituitary varied significantly in vivo over a 24-h period. Maximal expression for both TSH and GSU 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.
