Effects of Pax8 and TTF-1 thyroid transcription factor gene transfer in hepatoma cells: imaging of functional protein-protein interaction and iodide uptake.
The thyroid transcription factors TTF-1 and Pax8 cooperate in the transcriptional activation of thyroid-specific genes such as thyroglobulin (Tg), thyroperoxidase (TPO), and sodium/iodide symporter (NIS).


METHODS
Dog TTF-1 (dTTF-1) and the human Pax8 (hPax8) gene were transfected in Morris hepatoma (MH3924A) cells to investigate (a) the possible visualization of functional protein-protein interaction and (b) the induction of thyroid-specific gene expression. In MH3924A cell lines expressing dTTF-1, hPax8, or both, the activation of human Tg (hTg), human TPO (hTPO), or rat NIS (rNIS) promoter/enhancer was measured using firefly luciferase reporter constructs. Furthermore, the possible induction of thyroid-specific genes was investigated in iodide uptake and reverse transcription polymerase chain reaction (RT-PCR) experiments.


RESULTS
Low transcriptional activation of these constructs was observed in cells expressing either hPax8 or dTTF-1 alone. In contrast, the hTg and hTPO and, to a lesser extent, the rNIS regulatory region were significantly activated in cell lines expressing both transcription factors. Imaging the transcriptional activation of the thyroid-specific regulatory regions by Pax8 and TTF-1 was possible in nude mice implanted with MHhPax8dTTF-1 cells using a cooled charge-coupled device camera. Na(125)I uptake experiments and RT-PCR showed no effect of hPax8 and dTTF-1 on endogenous thyroid-specific gene expression in genetically modified cells.


CONCLUSIONS
The activation of thyroid-specific promoter/enhancer elements in Morris hepatoma cells depends on the functional interaction of hPax8 and dTTF-1. The cooperation of these 2 transcription factors can be visualized in vitro as well as in vivo. With regard to a possible application for radioiodine therapy, further modifications are required.
Altmann
Annette
A
Clinical Cooperation Unit of Nuclear Medicine, German Cancer Research Center, Im Neuenheimer Feld 280, D-69120 Heidelberg, Germany. a.altmann@dkfz.de
Schulz
Ralf B
RB
Glensch
Gabriela
G
Eskerski
Helmut
H
Zitzmann
Sabine
S
Eisenhut
Michael
M
Haberkorn
Uwe
U
D000818:Animals; D045744:Cell Line, Tumor; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D020869:Gene Expression Profiling / Q000379:methods*; D015972:Gene Expression Regulation, Neoplastic / Q000235:genetics; D018014:Gene Transfer Techniques; D006801:Humans; D007457:Iodine Radioisotopes / Q000493:pharmacokinetics*; D008114:Liver Neoplasms, Experimental / Q000235:genetics* / Q000378:metabolism*; D008163:Luminescent Measurements / Q000379:methods; D051379:Mice; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D000072636:PAX8 Transcription Factor; D051761:Paired Box Transcription Factors; D025941:Protein Interaction Mapping / Q000379:methods*; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D000074482:Thyroid Nuclear Factor 1; D014018:Tissue Distribution; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*
D004268:DNA-Binding Proteins; D007457:Iodine Radioisotopes; C000614412:NKX2-1 protein, human; C000614414:Nkx2-1 protein, mouse; C000614416:Nkx2-1 protein, rat; D009687:Nuclear Proteins; D000072636:PAX8 Transcription Factor; C083325:PAX8 protein, human; D051761:Paired Box Transcription Factors; C499495:Pax8 protein, Canis familiaris; C499496:Pax8 protein, mouse; C499497:Pax8 protein, rat; D011994:Recombinant Proteins; D000074482:Thyroid Nuclear Factor 1; D015534:Trans-Activators; D014157:Transcription Factors