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p21 Ras as a governor of global gene expression
M Abdellatif1, W R MacLellan, M D Schneider
1Department of Medicine, Baylor College of Medicine, Houston, Texas 77030.
Abstract:
The molecular mechanisms for signaling by receptor serine/threonine kinases are incompletely understood. To test the potential involvement of p21 H-Ras proteins in signal transduction for type beta transforming growth factors (TGF beta), TGF beta-responsive and constitutive reporter genes were cotransfected into cardiac myocytes and mink lung epithelial cells, with dominant inhibitory (Asn-17) or activated (Arg-12) Ras expression vectors. Asn-17 Ras inhibited both TGF beta-dependent and basal expression of inducible promoters (skeletal alpha-actin and plasminogen activator inhibitor-1), with equivalent dose-response relations. All seven reporter constructs were comparably sensitive to down-regulation by Asn-17 Ras, including those driven by nominally constitutive viral control regions or a TATA-less initiator element. All constructs were up-regulated by Arg-12 Ras more variably. Wild-type Ras had intermediate effects and could rescue a minimal thymidine kinase promoter from inhibition by dominant negative Ras. Thus, a Ras-dependent event is required for efficient expression of an unexpectedly global or inclusive set of genes.
Insights
Ras proteins are essential for transforming growth factor beta (TGF-β) signaling. This study shows Ras-dependent events regulate a broad range of genes, impacting TGF-β signal transduction.
Area of Science:
- Cell Biology
- Molecular Biology
- Biochemistry
Background:
- The molecular mechanisms of receptor serine/threonine kinase signaling, particularly for transforming growth factor beta (TGF-β), are not fully elucidated.
- Ras proteins are known regulators of cellular signaling pathways.
Purpose of the Study:
- To investigate the role of p21 H-Ras proteins in TGF-β signal transduction.
- To determine if Ras proteins are involved in the regulation of TGF-β-responsive genes.
Main Methods:
- Cotransfection of TGF-β-responsive and constitutive reporter genes into cardiac myocytes and mink lung epithelial cells.
- Utilized dominant inhibitory (Asn-17) and activated (Arg-12) Ras expression vectors.
- Assessed gene expression changes using reporter constructs driven by various promoters.
Main Results:
- Dominant-negative Asn-17 Ras significantly inhibited both TGF-β-dependent and basal gene expression across multiple reporter constructs.
- Activated Arg-12 Ras showed variable up-regulation of reporter gene expression.
- Wild-type Ras exhibited intermediate effects and could overcome inhibition by dominant-negative Ras.
Conclusions:
- Ras-dependent signaling events are required for the efficient expression of a wide array of genes.
- These findings suggest Ras proteins play a critical and global role in TGF-β-mediated gene regulation.