Related Experiment Videos
Transcriptional and post-transcriptional regulation of transforming growth factor beta type II receptor expression in
Weizhong Chang1, Macarena Parra, Changhua Ji
1Department of Surgery (Plastic Surgery Section), Yale University School of Medicine, 333 Cedar Street, New Haven, CT 06520-8041, USA.
Abstract:
Variations in transforming growth factor beta (TGF-beta) activity depend on the expression of specific receptors in normal as well as transformed cells. For example, in addition to mutations in TGF-beta type II receptor (TbetaRII) that abrogate normal TGF-beta function, its expression decreases during the transition from replication to extracellular matrix production, or in response to other growth regulators in bone. Therefore, to understand how TbetaRII expression is controlled, we cloned the rat TbetaRII gene promoter and defined basic aspects of its structure and activity. Among several cis-acting elements, mutations within an upstream E-box that specifically binds USF nuclear factors or a downstream Sp1 binding site significantly reduced TbetaRII promoter activity in primary cultures of fetal rat osteoblasts. Treatment with bone morphogenetic protein 2 (BMP-2), which induces further osteoblast differentiation, significantly reduced cell surface TbetaRII. However, BMP-2 did not alter TbetaRII promoter activity, steady state TbetaRII mRNA, or total TbetaRII protein, but caused an intracellular relocation of TbetaRII. Select transcriptional elements thus regulate TbetaRII gene expression, whereas post-translational events controlled by BMP-2 rapidly modify the amount of TbetaRII protein on the bone cell surface. Consequently, several processes can alter functional TbetaRII levels in order to regulate the biological effects of this important growth factor.
Insights
Transforming growth factor beta (TGF-beta) receptor II (TbetaRII) expression is regulated by specific DNA elements and post-translational modifications. Understanding these mechanisms is key to controlling TGF-beta signaling in bone cells.
Area of Science:
- Molecular Biology
- Cell Biology
- Biochemistry
Background:
- Transforming growth factor beta (TGF-beta) activity is modulated by its receptor (TbetaRII) expression levels.
- TbetaRII expression changes are observed in bone cells during differentiation and in response to growth regulators.
- Understanding TbetaRII gene regulation is crucial for comprehending TGF-beta signaling pathways.
Purpose of the Study:
- To investigate the transcriptional regulation of the rat TbetaRII gene.
- To identify key cis-acting elements and nuclear factors involved in TbetaRII promoter activity.
- To elucidate the effects of bone morphogenetic protein 2 (BMP-2) on TbetaRII expression and localization.
Main Methods:
- Cloning and characterization of the rat TbetaRII gene promoter.
- Site-directed mutagenesis of putative cis-acting elements (E-box, Sp1 site).
- Reporter gene assays in primary fetal rat osteoblast cultures.
- Analysis of TbetaRII mRNA and protein levels following BMP-2 treatment.
- Immunofluorescence to assess TbetaRII intracellular localization.
Main Results:
- Mutations in the upstream E-box (USF binding) or downstream Sp1 binding site significantly reduced TbetaRII promoter activity.
- BMP-2 treatment decreased cell surface TbetaRII but did not affect TbetaRII promoter activity, mRNA, or total protein.
- BMP-2 induced an intracellular relocation of TbetaRII in osteoblasts.
Conclusions:
- Specific transcriptional elements, including E-box and Sp1 sites, are critical for regulating TbetaRII gene expression.
- BMP-2 influences functional TbetaRII levels through post-translational mechanisms, specifically by altering its cell surface localization.
- Both transcriptional and post-translational regulation contribute to the control of TbetaRII levels, impacting TGF-beta signaling in bone cells.