Signalwege des TSH-Rezeptors - Gq/11-vermittelte Regulation von Metallothioninen in Schilddrüsenkarzinomzellen

Metallothionine (MT) sind cysteinreiche intrazelluläre Proteine, für die eine zytoprotektive Wirkung beschrieben wurde, da sie Zellen gegen Schwermetallionen und oxidativen Stress schützen. In früheren Untersuchungen wurde die Expression von Metallothioninen in normalen sowie neoplastischen Schilddr...

Full description

Saved in:
Bibliographic Details
Main Author: Bäck, Christer
Contributors: Gudermann, Thomas (Prof. Dr.) (Thesis advisor)
Format: Doctoral Thesis
Published: Philipps-Universität Marburg 2009
Online Access:PDF Full Text
Tags: Add Tag
No Tags, Be the first to tag this record!

MT are cystein-rich intracellular proteins which exert anti-apoptotic effects by protecting cells against oxidative stress and heavy metal ions. Previously, expression of MT in normal and neoplastic thyroid tissue has been demonstrated. However, the thyroidal regulation of MT expression is widely unsettled. Thus, we investigated the expression of MT isoform 1 in human thyroid carcinoma cells (FTC-133-TSHR) and in primary human thyrocytes (SD-191) upon stimulation with thyrotropin (TSH). Using quantitative RT-PCR we found that TSH led to a dose-dependent increase in MT1 mRNA levels in these cells. To further characterize the signaling pathway involved in MT1 induction we investigated thyroid carcinoma cells expressing a mutated TSH receptor incapable to couple to Gq/11 proteins (FTC-133 Y601H cells). In these cells, TSH still led to a marked increase in intracellular cAMP levels whereas an increase in inositol phosphates was completely absent. Interestingly, TSH did not induce MT1 in these cells, giving evidence that regulation of MT1 was cAMP-independent but dependent on Gq/11- coupling. This finding was further verified by the fact that TSH-promoted induction of MT1 in FTC-133-TSHR cells was blocked by inhibitors of proteinkinase C (PKC), whereas activation of PKC by treatment with phorbol esters mimicked the effect of TSH. Finally, we investigated changes in MT1 protein levels. Immunoblots and immunocytochemistry with MT1 specific antibodies revealed a TSH-induced up-regulation of MT1 in FTC-133-TSHR cells whereas no effect of TSH occurred in FTC-133 Y601H cells. In addition we turned our attention to the PKC-depended phosphorylation of the transcription factor STAT1. With the help of immunoblots we could find a correlation between the TSH-depending phosphorylation of STAT through the PKC and the induction of MT1. To summarize the induction of MT1 in human thyroid cells through the TSHR is shown in fig. 1. Thus we were able to describe a receptor-mediated induction of an antioxidant protein in a cell, which is physiologically under oxidative stress. Interestingly, the TSH-receptor seems to activate oxidative and antioxidative mechanism simultaneously via different G-proteins. The finding of Gq/11-dependent regulation of MT1 by TSH adds further complexity to possible cAMP-independent functions of the TSH receptor.