Sequential up‐regulation of thyroid hormone β receptor, ornithine transcarbamylase, and carbamyl phosphate synthetase mRNAs in the liver of Rana catesbeiana tadpoles during spontaneous and thyroid hormone‐induced metamorphosis

Abstract

During both spontaneous and thyroid hormone (TH)-induced metamorphosis, the Rana catesbeiana tadpole undergoes postembryonic developmental changes in its liver which are necessary for its transition from an ammonotelic larva to a ureotelic adult. Although this transition ultimately results from marked increases in the activities and/or de novo synthesis of the urea cycle enzymes, the precise molecular means by which TH exerts this tissue-specific response are presently unknown. Recent reports, using RNA from whole Xenopus laevis tadpole homogenates and indirect means of measuring TH receptor (TR) mRNAs, suggest a correlation between the up-regulation of TR beta-mRNAs and the general morphological changes occurring during amphibian metamorphosis. To assess whether or not this same relationship exists in a TH-responsive tissue, such as liver, we isolated and characterized a cDNA clone containing the complete nucleotide sequence for a R. catesbeiana urea cycle enzyme, ornithine transcarbamylase (OTC), as well as a genomic clone containing a portion of the hormone-binding domain of a R. catesbeiana TR beta gene. Through use of these homologous sequences and a heterologous cDNA fragment encoding rat carbamyl phosphate synthetase (CPS), we directly determined the relative levels of the TR beta, OTC, and CPS mRNAs in liver from spontaneous and TH-induced tadpoles. Our results establish that TH affects an up-regulation of mRNAs for its own receptor prior to up-regulating CPS and OTC mRNAs. Moreover, results with cultured tadpole liver demonstrate that TH, in the absence of any other hormonal influence, can affect an up-regulation of both the TR beta and OTC mRNAs.

Genistein prevents thyroid hormone‐dependent tail regression of Rana catesbeiana tadpoles by targetting protein kinase C and thyroid hormone receptor α

“…The procedure used for tail culture was adapted from that described previously (Helbing et al, 1992). Tadpoles were euthanized in 0.1% tricaine methane sulfonate (MS-222) (Syndel Laboratories, Vancouver, BC).…”

Thyroid hormone-dependent metamorphosis in a direct developing frog

“…TH, shown in other systems to signal through thyroid hormone receptors (TRs), controls the larval-to-adult transition in biphasic frogs (4). TR␤ expression in both Xenopus laevis and Rana catesbeiana is temporally correlated with the onset of metamorphosis, implicating this gene as a likely metamorphic mediator (5)(6)(7). The role of TH in direct development is unclear.…”

Amphibian cardiac troponin I gene's organization, developmental expression, and regulatory properties are different from its mammalian homologue

“…To ensure that comparisons were performed on samples within the linear range of amplification, an 8-l aliquot was removed from each tube after 16 cycles and at the completion of every second subsequent cycle, and subjected to agarose gel electrophoresis, stained with ethidium bromide, and visualized on a ultraviolet transilluminator. The primer pairs used were the following: a Rana TR ␣ (5Ј-GGTGAGATG-GCAGTGAAGCGAGAACA G-3Ј, 5Ј-AAGATG G GTAGAGAGGAA-AGGGAATT-3Ј; Schneider and Galton, 1991), a Rana TR ␤ (5Ј-ACAAAA ATAATCACCCCAGCA-AT-3Ј, 5Ј-ATGCGGGTACTCGTGAC-TATCGTGTT-3Ј; Helbing et al, 1992), a Rana TnIs (5Ј-ATGCCAGAACCA-GAGAGGAAGTC-3Ј, 5Ј-AGGAC-GAAATCT GGAAAATAATGT-3Ј), and a Rana TnIc (5Ј-AAGCTA-GAGCCCAAGAGAAAGAGAG-3Ј, 5Ј-CTGATCCATCTTCTGACCTTTT-TAG-3Ј).…”

“…A Rana catesbeiana genomic library (Helbing et al, 1992) was screened (approximately 1 ϫ 10 6 plaques) by using a random primed [␣-32 P]dCTP-labeled, fulllength Rana TnIc cDNA. Hybridization was preformed as described for cDNA library screening except that 20% deionized formamide was used in the hybridization solution.…”