Utilization of temperature-mediated activation of thyroid hormone-induced molecular memory to evaluate early signaling events in the olfactory epithelium of Rana [Lithobates] catesbeiana tadpoles.

The amphibian olfactory system is highly distinct between aquatic tadpole and terrestrial frog life stages and therefore must remodel extensively during thyroid hormone (TH)-dependent metamorphosis. Developmentally appropriate functioning of the olfactory epithelium is critical for survival. Previous studies in other Rana [Lithobates] catesbeiana premetamorphic tadpole tissues showed that initiation of TH-induced metamorphosis can be uncoupled from execution of TH-dependent programs by holding tadpoles in the cold rather than at warmer permissive temperatures. TH-exposed tadpoles at the nonpermissive (5 °C) temperature do not undergo metamorphosis but retain a "molecular memory" of TH exposure that is activated upon shift to a permissive warm temperature. Herein, premetamorphic tadpoles were held at permissive (24 °C) or nonpermissive (5 °C) temperatures and injected with 10 pmoles/g body weight 3,5,3'-triiodothyronine (T3) or solvent control. Olfactory epithelium was collected at 48 h post-injection. RNA-sequencing (RNA-Seq) and reverse transcriptase quantitative real-time polymerase chain reaction (RT-qPCR) analyses generated differentially expressed transcript profiles of 4328 and 54 contigs for permissive and nonpermissive temperatures, respectively. Translation, rRNA, spliceosome, and proteolytic processes gene ontologies were enriched by T3 treatment at 24 °C while negative regulation of cell proliferation was enriched by T3 at 5 °C. Of note, as found in other tissues, TH-induced basic leucine zipper-containing protein-encoding transcript, thibz, was significantly induced by T3 at both temperatures, suggesting a role in the establishment of molecular memory in the olfactory epithelium. The current study provides critical insights by deconstructing early TH-induced induction of postembryonic processes that may be targets for disruption by environmental contaminants.

Comparison of transcriptome responses of the liver, tail fin, and olfactory epithelium of Rana [Lithobates] catesbeiana tadpoles disrupted by thyroid hormones and estrogen.

Thyroid hormones (THs) are important developmental regulators in vertebrates, including during the metamorphosis of a tadpole into a frog. Metamorphosis is a post-embryonic developmental period initiated by TH production in the tadpole thyroid gland. The two main bioactive forms of TH are L-thyroxine (T4) and 3,5,3'-triiodothyronine (T3); these hormones have overlapping but distinct mechanisms of action. Premetamorphic tadpoles are highly responsive to TH and can be induced to metamorphose through exogenous TH exposure, making them an important model for both the study of vertebrate TH signaling and endocrine disrupting chemicals (EDCs). It is important to differentiate TH-mediated responses from estrogenic responses in premetamorphic tadpoles when assessing dysregulation by EDCs as crosstalk between the two endocrine systems is well-documented. Herein, we compare the RNA-sequencing-derived transcriptomic profiles of three TH-responsive tissues (liver, olfactory epithelium, and tail fin) in premetamorphic bullfrog (Rana [Lithobates] catesbeiana) tadpoles exposed to T3, T4, and estradiol (E2). These profiles were generated using the latest available genome assembly for the species. The data indicate that there is a clear distinction, and little overlap, between the transcriptomic responses elicited by E2 and the THs. In contrast, within the THs, the T3- and T4-induced transcriptomic profiles generally show considerable overlap; however, the degree of overlap is highly tissue-dependent, illustrating the importance of distinguishing the two THs and the affected signaling pathways within the target tissue type when evaluating hormone active agents. The data herein also show that E2 and TH treatment can uniquely induce significant changes in expression of their respective "classic" bioindicator transcripts vtg (E2) and thra, thrb, and thibz (THs). However, care must be taken in the interpretation of increased vep or esr1 transcripts as a change in transcript levels can be induced by THs rather than solely E2.