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Regulation of Raf by Akt controls growth and differentiation in vascular smooth muscle cells

H P Reusch1, S Zimmermann, M Schaefer

  • 1Institut für Klinische Pharmakologie und Toxikologie, Freie Universität Berlin, Garystr. 5, 14195 Berlin, Germany. reusch@medizin.fu-berlin.de

The Journal of Biological Chemistry
|July 10, 2001
PubMed

Insights

Platelet-derived growth factor (PDGF) and thrombin differentially regulate vascular smooth muscle cell (VSM) phenotype. The balance between PI 3-kinase/Akt and Ras/Raf/MEK pathways dictates ERK1/2 phosphorylation and VSM cell differentiation.

Area of Science:

  • Vascular Biology
  • Cell Signaling
  • Molecular Medicine

Background:

  • Vascular smooth muscle (VSM) cells undergo phenotypic modulation in response to growth factors and signaling molecules.
  • Platelet-derived growth factor (PDGF) is a known mitogen for VSM cells, while thrombin also activates similar signaling pathways.
  • Understanding the differential effects of these stimuli on VSM cell phenotype is crucial for cardiovascular research.

Purpose of the Study:

  • To investigate the distinct signaling mechanisms by which PDGF and thrombin influence VSM cell phenotype.
  • To elucidate the role of the Ras/Raf/MEK/ERK and PI 3-kinase/Akt pathways in mediating these cellular responses.
  • To determine how the interplay between these pathways regulates VSM cell differentiation markers like SM-MHC.

Main Methods:

  • Stimulation of VSM cells with PDGF and thrombin.
  • Analysis of signaling pathway activation, including ERK1/2 and Akt phosphorylation.
  • Measurement of smooth muscle-specific myosin heavy chain (SM-MHC) promoter activity.
  • Use of dominant-negative Akt and constitutively active Akt constructs.

Main Results:

  • Thrombin induced SM-MHC expression (re-differentiation) via biphasic ERK1/2 phosphorylation, with limited Akt activation.
  • PDGF, a mitogen, caused transient ERK1/2 phosphorylation but sustained Akt activation.
  • Akt directly inhibited Raf kinase activity, limiting ERK1/2 phosphorylation; disruption of PI 3-kinase signaling reversed this effect for PDGF.
  • Akt activation blocked thrombin-induced SM-MHC promoter activity, while inhibiting Akt enhanced PDGF-induced SM-MHC activity.

Conclusions:

  • The cross-regulation between PI 3-kinase/Akt and Ras/Raf/MEK pathways is critical for determining the temporal pattern of ERK1/2 phosphorylation.
  • This signaling balance dictates the phenotypic outcome of VSM cells, distinguishing between proliferation and differentiation.
  • Differential activation and cross-talk between these pathways provide a mechanism for controlling VSM cell fate in response to various stimuli.

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