Thyroglobulin repression of thyroid transcription factor 1 (TTF-1) gene expression is mediated by decreased DNA binding of nuclear factor I proteins which control constitutive TTF-1 expression.
Follicular thyroglobulin (TG) selectively suppresses the expression of thyroid-restricted transcription factors, thereby altering the expression of thyroid-specific proteins. In this study, we investigated the molecular mechanism by which TG suppresses the prototypic thyroid-restricted transcription factor, thyroid transcription factor 1 (TTF-1), in rat FRTL-5 thyrocytes. We show that the region between bp -264 and -153 on the TTF-1 promoter contains two nuclear factor I (NFI) elements whose function is involved in TG-mediated suppression. Thus, NFI binding to these elements is critical for constitutive expression of TTF-1; TG decreases NFI binding to the NFI elements in association with TG repression. NFI is a family of transcription factors that is ubiquitously expressed and contributes to constitutive and cell-specific gene expression. In contrast to the contribution of NFI proteins to constitutive gene expression in other systems, we demonstrate that follicular TG transcriptionally represses all NFI RNAs (NFI-A, -B, -C, and -X) in association with decreased NFI binding and that the RNA levels decrease as early as 4 h after TG treatment. Although TG treatment for 48 h results in a decrease in NFI protein-DNA complexes measured in DNA mobility shift assays, NFI proteins are still detectable by Western analysis. We show, however, that the binding of all NFI proteins is redox regulated. Thus, diamide treatment of nuclear extracts strongly reduces the binding of NFI proteins, and the addition of higher concentrations of dithiothreitol to nuclear extracts from TG-treated cells restores NFI-DNA binding to levels in extracts from untreated cells. We conclude that NFI binding to two NFI elements, at bp -264 to -153, positively regulates TTF-1 expression and controls constitutive TTF-1 levels. TG mediates the repression of TTF-1 gene expression by decreasing NFI RNA and protein levels, as well as by altering the binding activity of NFI, which is redox controlled.
Nakazato
M
M
Cell Regulation Section, Metabolic Diseases Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892, USA.
Chung
H K
HK
Ulianich
L
L
Grassadonia
A
A
Suzuki
K
K
Kohn
L D
LD
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D022762:CCAAT-Enhancer-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D002417:Cattle; D002478:Cells, Cultured; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins; D003958:Diamide / Q000737:chemistry / Q000494:pharmacology; D005786:Gene Expression Regulation; D008969:Molecular Sequence Data; D051839:NFI Transcription Factors; D009687:Nuclear Proteins / Q000235:genetics* / Q000378:metabolism; D009841:Oligonucleotides / Q000378:metabolism; D011401:Promoter Regions, Genetic; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D051381:Rats; D012045:Regulatory Sequences, Nucleic Acid; D013954:Thyroglobulin / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D013961:Thyroid Gland / Q000166:cytology / Q000187:drug effects / Q000502:physiology; D000074482:Thyroid Nuclear Factor 1; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism; D051840:Y-Box-Binding Protein 1
D022762:CCAAT-Enhancer-Binding Proteins; D004268:DNA-Binding Proteins; D051839:NFI Transcription Factors; C000614412:NKX2-1 protein, human; C000614416:Nkx2-1 protein, rat; D009687:Nuclear Proteins; D009841:Oligonucleotides; D020033:Protein Isoforms; D000074482:Thyroid Nuclear Factor 1; D014157:Transcription Factors; D051840:Y-Box-Binding Protein 1; C070856:YBX1 protein, human; D003958:Diamide; D004247:DNA; D013954:Thyroglobulin