Oncogene. 2005 Oct 27;24(47):6993-7001.

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Retinoblastoma protein acts as Pax 8 transcriptional coactivator.

Miccadei S, Provenzano C, Mojzisek M, Natali PG, Civitareale D.

Molecular Pathology Laboratory, Regina Elena Cancer Institute, Via delle Messi d'Oro 156, 00158
Rome, Italy.

Control of cell proliferation and differentiation by the retinoblastoma protein (pRb) depends on its interactions with key cellular substrates. Available data indicate that pRb and the transcription factor Pax 8 play a crucial role in the differentiation of thyroid follicular cells. In this study, we show that pRb takes part in the complex assembled on the thyroperoxidase gene promoter acting as a transcriptional coactivator of Pax 8. Accordingly, pRb interacts with and potentiates Pax 8 transcriptional activity. In addition, we show that the downregulation of pRb gene expression, in thyrocytes, through RNA interference results in a reduction of the thyroperoxidase gene promoter activity mediated by the Pax 8-binding site. In agreement with these results and with the ability of the adenoviral protein E1A to bind pRb, we show that E1A downregulates Pax 8 activity and that such inhibition requires the E1A-Rb interaction. Furthermore, we show that the Pax 8/pRb synergy plays a role on the sodium/iodide symporter gene expression as well.

PMID: 16007137 [PubMed - indexed for MEDLINE]

Endocrinology. 2005 Sep;146(9):4009-15. Epub 2005 Jun 16.

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Expression, regulation, and function of paired-box gene 8 in the human placenta and placental cancer cell lines.

Ferretti E, Arturi F, Mattei T, Scipioni A, Tell G, Tosi E, Presta I, Morisi R, Lacroix L, Gulino A, Russo D, Damante G, Filetti S.

Department of Clinical Science,
University of Rome, La Sapienza, Rome, Italy.

Pax proteins are transcriptional regulators that control a variety of developmental decisions in vertebrates. During development, the paired-box gene 8 (PAX8) is expressed in the thyroid, kidney, and several areas of the central nervous system. It is also expressed in the adult thyroid gland, in which it mediates TSH-induced modulation of the expression of important genes, such as those encoding thyroglobulin, thyroperoxidase, and the sodium/iodide symporter (
NIS). Thus far, placental expression of PAX8 has been described only in mice. In the present study, we show that PAX8 is also expressed in the human placenta at term. In an in vitro model of placental cancer, the JAR choriocarcinoma cell line, human chorionic gonadotropin (hCG) increased levels of PAX8 mRNA and protein, and gel retardation assays indicated that the up-regulation of PAX8 protein expression is associated with an increase in its DNA-binding activity. The effects of hCG were mimicked by forskolin, indicating that they are cAMP dependent. Levels of mRNA for the Wilms' tumor 1 (WT1) and NIS genes were increased in JAR cells by hCG treatment, whereas overexpression of PAX8 increased only levels of WT1 mRNA. In cells transfected with PAX8-specific small interfering RNA, the stimulatory effects of hCG on WT1 mRNA levels were abolished, but hormonal enhancement of NIS mRNA levels was unchanged. These findings indicate that, in JAR cells, hCG activates a cAMP-dependent pathway that can up-regulate WT1 expression through PAX8.

Mech Dev. 2005 Jul;122(7-8):887-99.

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J Biol Chem. 2005 Jul 8;280(27):25901-6. Epub 2005 May 11.

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Glutathionylation of two cysteine residues in paired domain regulates DNA binding activity of Pax-8.

Cao X, Kambe F, Lu X, Kobayashi N, Ohmori S, Seo H.

Department of Endocrinology and Metabolism, Division of Molecular and Cellular Adaptation, Research Institute of Environmental Medicine,
Nagoya University, Nagoya 464-8601, Japan. kambe@riem.nagoya-u.ac.jp

We reported that the first two cysteine residues out of three present in paired domain (PD), a DNA-binding domain, are responsible for redox regulation of Pax-8 DNA binding activity. We show that glutathionylation of these cysteines has a regulatory role in PD binding. Wild-type PD and its mutants with substitution of cysteine to serine were synthesized and named CCC, CSS, SCS, SSC, and SSS according to the positions of substituted cysteines. They were incubated in a buffer containing various ratios of GSH/GSSG and subjected to gel shift assay. Binding of CCC, CSS, and SCS was impaired with decreasing GSH/GSSG ratio, whereas that of SSC and SSS was not affected. Because [3H]glutathione was incorporated into CCC, CSS, and SCS, but not into SSC and SSS, the binding impairment was ascribed to glutathionylation of the redox-reactive cysteines. This oxidative inactivation of PD binding was reversed by a reductant dithiothreitol and by redox factor (Ref)-1 in vitro. To explore the glutathionylation in cells, Chinese hamster ovary cells overexpressing CSS and SCS were labeled with [35S]cysteine in the presence of cycloheximide. Immunoprecipitation with an antibody against PD revealed that treatment of the cells with an oxidant diamide induced the 35S incorporation into both mutants, suggesting the PD glutathionylation in cells. Since the two cysteine residues in PD are conserved in all Pax members, this novel posttranslational modification of PD would provide a new insight into molecular basis for modulation of Pax function.

PMID: 15888455 [PubMed - indexed for MEDLINE]
















Flowchart: Preparation: Pax8












Text Box: E1AText Box: pRbText Box: Pax8Text Box: TSH









    Placental cancer