Auto-regulation of the thyroid gland by classical and non-classical pathways

甲状腺激素（TH）靶细胞需要采取机制来维持足够的TH水平，以确保正常的功能。对TH缺乏或过量的自动调节或自动保护机制的干扰是器官功能不足的第一可能原因。在甲状腺本身的特定情况下，自动保护对于依赖TH的所有其他器官也至关重要。因此，该项目的主要目标是更好地了解甲状腺功能自动调节的细胞和分子途径。为了这个目的，我们研究了经典的监管机制，通过比较野生型小鼠与组织蛋白酶缺陷的动物，其特点是改变TH代由于受损的甲状腺球蛋白加工。甲状腺自身调节的非经典途径进行了研究，通过分析TH转运体和甲状腺素。我们的研究结果表明甲状腺球蛋白加工和TH转运蛋白之间的相互作用。我们建议，TH传感机制，使不同的TH转运蛋白存在于特定的亚细胞位置在甲状腺上皮细胞选择性监测甲状腺球蛋白处理的各个步骤，因此，细胞TH状态。因此，我们建立的甲状腺结构和功能参数的自动化、无偏倚分析管道将用于进一步分析小鼠甲状腺球蛋白加工改变或TH易位对原位甲状腺功能的影响。此外，我们将使用一个建立在体外系统，FRT和FRTL-5细胞组成，调查运输TH转运蛋白，并研究其功能TH清洁和生理TH条件下。我们希望证实我们的建议，蛋白酶和TH转运功能连接。甲状腺胺是经典TH的衍生物，也可能参与甲状腺调节。已知甲状腺原氨酸3-T1 AM在体外触发G蛋白偶联痕量胺相关受体1（Taar 1）。我们在培养的甲状腺细胞的顶端质膜的纤毛和甲状腺组织中检测到Taar 1。这些结果表明，甲状腺滤泡腔作为甲状腺原氨酸生成的假定位置，并通过作用于甲状腺上皮细胞顶端纤毛的Taar 1对甲状腺的自动调节做出贡献。为了通过信号传导研究阐明这种途径，我们将在体外用3-T1 AM或其他已知激动剂孵育正常和Taar（过）表达的甲状腺细胞。此外，表型Taar 1缺陷小鼠将使我们能够研究Taar 1对甲状腺原位自动调节的具体贡献。最后，甲状腺组织和血清中经典和非经典TH（甲状腺素组）的测定是及时的，将有助于了解甲状腺球蛋白加工、TH易位和甲状腺素应答能力受到挑战的小鼠中甲状腺素的代谢可用性。

Thyroid hormone (TH) target cells need to adopt mechanisms to maintain sufficient levels of TH to ensure regular functions. Interference with auto-regulative or auto-protective mechanisms against a lack or an excess of TH is a first likely cause of inadequate organ function. In the specific case of the thyroid gland itself, auto-protection is also vital for all other organs that are dependent on TH. The main objective of this project is therefore to reach a better understanding of cellular and molecular pathways underlying auto-regulation of thyroid functions. To this aim, we investigated classical regulatory mechanisms by comparing wild type mice with cathepsin-deficient animals that are characterized by altered TH generation due to impaired thyroglobulin processing. Non-classical pathways of thyroid auto-regulation were studied by analyzing TH transporters and thyronamines. Our results suggest an interplay between thyroglobulin processing and TH transporters. We propose that a TH sensing mechanism is enabled such that different TH transporters are present in specific subcellular locations in thyroid epithelial cells selectively monitoring individual steps of thyroglobulin processing and thus, the cellular TH status. Therefore our established pipeline of automated, non-biased analyses of structural and functional parameters of the thyroid gland will be employed to further analyze mice for the effects of altered thyroglobulin processing or TH translocation on thyroid function in situ. Additionally we will use an established in vitro system, consisting of FRT and FRTL-5 cells, to investigate trafficking of TH transporter proteins and to study their function under TH-clean and physiological TH conditions. We expect to substantiate our proposal that proteases and TH transporters are functionally connected. Thyronamines are derivatives of the classical TH which might also engage in thyroid regulation. It is known that the thyronamine 3-T1AM triggers the G-protein coupled trace amine-associated receptor 1 (Taar1) in vitro. We detected Taar 1 at cilia of the apical plasma membrane of cultured thyrocytes and in thyroid tissue. These results suggest the thyroid follicle lumen as the putative location of thyronamine generation and its contribution to auto-regulation of the thyroid gland by acting on Taar1 at apical cilia of thyroid epithelial cells. To elucidate such pathways by signaling studies we will incubate normal and Taar (over)-expressing thyrocytes with 3-T1AM or other known agonists in vitro. In addition, phenotyping Taar1-deficient mice will allow us to investigate the specific contribution of Taar1 to auto-regulation of the thyroid gland in situ. Finally, determination of classical and non-classical TH (the thyronome) in thyroid tissue and blood serum is timely and will help to understand metabolic availabilities of thyronamines in mice with challenged thyroglobulin processing, TH translocation, and thyronamine-responsive capabilities.