Thyroid diseases affect over 25 million Americans and constitute the second most common group of metabolic disorders. Thyroid hormone (TH) is critical for growth, development, and metabolism, and works by binding TH receptors in the cell nucleus and changing gene expression. Importantly, TH must cross the plasma membrane from the blood to bind TH receptors in the nucleus. One cell surface protein in this translocation process, the L-type amino acid transporter LAT1, is strongly up-regulated during TH-dependent development. Interestingly, LAT1 expression is also associated with rapidly dividing cells, such as activated lymphocytes and cancer. Even though many in vitro studies on LAT1 function have been carried out, the in-vivo developmental role of LAT1 is not well understood. The objective of the current project is to study how LAT1 regulates tissue responses to TH during development. This project complements the investigator's long-term goal of elucidating molecular mechanisms underlying control of TH signaling by cytoplasmic proteins. To achieve this objective, the plan is to, in Specific Aim 1, develop and characterize an improved tetracycline-inducible transgenic system. He will design, produce, and characterize transgenic inducer and reporter lines that, when crossed, will be capable of high levels of transgene induction. This improved system will enable us to minimize leaky transgene expression, reduce variability in transgene expression levels caused by insertion sites, and """"""""dial in"""""""" transgene expression levels with the inducing chemical doxycycline (Dox) administered at different doses. In addition, this system can expand to enable tissue-specific transgene induction. Preliminary data showed minimal leaky expression in the absence of Dox and high transgene induction in the presence of Dox.
Specific Aim 2 will then examine the in-vivo role of LAT1 in TH-dependent development. The investigator will use the transgenic technology from Aim 1 to test the hypothesis that LAT1 modulates TH signaling in vivo. He will compare rate and magnitude of TH-induced morphological and gene regulation changes in transgenic tadpoles treated with and without Dox to induce LAT1 overexpression. This project is expected to reveal fundamental insights about how LAT1 regulates TH signaling in vivo. In particular, it is expected that overexpression of the TH transporter LAT1 will increase availability of TH to nuclear TH receptors as measured by increased TH-dependent gene expression and morphological change. A significant positive impact of this project is that the investigator will extend the usefulness of a well-established system for studying TH-dependent development, frog metamorphosis, and provide new transgenic resources to the frog community. These resources will be useful for a wide variety of experimental approaches. In addition, knowledge of how LAT1 regulates tissue responses to TH in-vivo is significant when considering LAT1 as a target for drug development in the treatment of thyroid disorders and cancer. PROJECT NARRATIVE: The investigator is studying how the cell surface protein LAT1 affects thyroid hormone signaling during development. Importantly, LAT1 has been identified as a drug target in the treatment of thyroid diseases and related disorders and complications, such as obesity, cardiovascular disease, and cancer. Thus, knowledge of developmental roles of LAT1 is important when considering use of drugs targeting LAT1 to evaluate potential benefits and side-effects.
We are studying the how the cell surface protein LAT1 affects thyroid hormone signalling during development. Importantly, LAT1 has been identified as a drug target in the treatment of thyroid diseases and related disorders and complications, such as obesity, cardiovascular disease, and cancer. Thus, knowledge of developmental roles of LAT1 is important when considering use of drugs targeting LAT1 to evaluate potential benefits and side-effects.
| Buchholz, Daniel R (2012) Tet-On binary systems for tissue-specific and inducible transgene expression. Methods Mol Biol 917:265-75 |
| Kerney, Ryan R; Brittain, Alison L; Hall, Brian K et al. (2012) Cartilage on the move: cartilage lineage tracing during tadpole metamorphosis. Dev Growth Differ 54:739-52 |
| Hollar, Amy R; Choi, Jinyoung; Grimm, Adam T et al. (2011) Higher thyroid hormone receptor expression correlates with short larval periods in spadefoot toads and increases metamorphic rate. Gen Comp Endocrinol 173:190-8 |
| Rankin, Scott A; Zorn, Aaron M; Buchholz, Daniel R (2011) New doxycycline-inducible transgenic lines in Xenopus. Dev Dyn 240:1467-74 |
