Related Experiment Videos
Study of gene expression in thyrotropin-stimulated thyroid cells by cDNA expression array: ID3 transcription
S Deleu1, V Savonet, J Behrends
1Institute of Interdisciplinary Research, School of Medicine, Free University of Brussels, Campus Erasme, route de Lennik 808, B-1070, Brussels, Belgium.
Abstract:
Induction of cell proliferation by mitogen or growth factor stimulation leads to the specific induction or repression of a large number of genes. To identify genes differentially regulated by the cAMP-dependent transduction pathway, which is poorly characterized so far, we used the cDNA expression array technology. Hybridizations of Atlas human cDNA expression arrays with (32)P-labeled cDNA probes derived from control or thyrotropin (TSH)-stimulated dog thyrocytes in primary culture generated expression profiles of hundreds of genes simultaneously. Among the genes that displayed modified expression, we selected the transcription factor ID3, whose expression was increased by a cAMP-dependent stimulus. ID3 overexpression after TSH stimulation was first verified by Northern blotting analysis, and its mRNA regulation was then investigated in response to a variety of agents acting on thyrocyte proliferation and/or differentiation. We show that: (1) ID3 mRNA induction was stronger after stimulation of the cAMP cascade, but was not restricted to this signaling pathway, as phorbol myristate ester (TPA) and insulin also stimulated mRNA accumulation; (2) in contrast, powerful mitogens for thyroid cells, epidermal growth factor and hepatocyte growth factor, did not significantly modify ID3 mRNA levels; (3) ID3 protein levels closely parallelled mRNA levels, as revealed by immunofluorescence experiments showing a nuclear signal regulated by TSH; (4) in papillary thyroid carcinomas, ID3 mRNA was downregulated. Our results suggest that ID3 expression might be more related to the differentiating process induced by TSH than to the proliferative action of this hormone.
Insights
The study identified the ID3 gene as upregulated by thyrotropin (TSH) stimulation in thyroid cells, suggesting a role in differentiation rather than proliferation. ID3 gene expression was found to be downregulated in papillary thyroid carcinomas.
Area of Science:
- Molecular Biology
- Cell Biology
- Endocrinology
Background:
- Cell proliferation is regulated by mitogens and growth factors, inducing differential gene expression.
- The cAMP-dependent signal transduction pathway's role in gene regulation is not fully understood.
Purpose of the Study:
- To identify genes differentially regulated by the cAMP-dependent pathway.
- To investigate the role of the transcription factor ID3 in thyrocyte function.
Main Methods:
- Utilized cDNA expression array technology to analyze gene expression profiles.
- Employed Northern blotting and immunofluorescence to validate and study ID3 mRNA and protein regulation.
- Investigated ID3 expression in response to various stimuli and in papillary thyroid carcinomas.
Main Results:
- TSH stimulation significantly increased ID3 gene expression in dog thyrocytes.
- ID3 mRNA accumulation was also induced by phorbol myristate ester (TPA) and insulin, but not by epidermal growth factor or hepatocyte growth factor.
- ID3 protein levels paralleled mRNA levels, with TSH regulating its nuclear localization.
- ID3 mRNA was found to be downregulated in papillary thyroid carcinomas.
Conclusions:
- ID3 expression is regulated by multiple signaling pathways, including the cAMP cascade.
- ID3's role appears more closely linked to thyroid cell differentiation induced by TSH than to proliferation.
- Downregulation of ID3 in papillary thyroid carcinomas may have implications for thyroid cancer development.