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Ras-dependent ERK activation by the human G(s)-coupled serotonin receptors 5-HT4(b) and 5-HT7(a)

Jens Henrik Norum1, Kent Hart, Finn Olav Levy

  • 1Merck Sharp and Dohme Cardiovascular Research Center, Institute for Surgical Research and Department of Pharmacology, Rikshospitalet University Hospital, University of Oslo, N-0316 Oslo, Norway.

The Journal of Biological Chemistry
|November 26, 2002
PubMed

Insights

Human serotonin receptors 5-HT(4(b)) and 5-HT(7(a)) activate extracellular signal-regulated kinases (ERK1/2) via a Ras-dependent pathway. This study reveals a novel signaling cascade in HEK293 cells, independent of Rap1.

Area of Science:

  • Cellular signaling pathways
  • Molecular biology
  • Neuroscience

Background:

  • Receptor tyrosine kinases (RTKs) activate mitogen-activated protein (MAP) kinases via Ras, Raf-1, and MEK.
  • G protein-coupled receptors (GPCRs) can transactivate RTKs, typically coupling to G(i) and G(q).
  • The human serotonin receptors 5-HT(4(b)) and 5-HT(7(a)) are G(s)-coupled and increase intracellular cyclic adenosine monophosphate (cAMP).

Purpose of the Study:

  • To investigate the activation of extracellular signal-regulated kinases (ERKs) 1 and 2 (p44 and p42 MAP kinase) by the human serotonin receptors 5-HT(4(b)) and 5-HT(7(a)).
  • To elucidate the signaling pathway involved in this activation, particularly the roles of protein kinase A (PKA), Ras, and Rap1.

Main Methods:

  • Transfection of COS-7 and human embryonic kidney (HEK293) cells with serotonin receptor constructs.
  • Stimulation with serotonin (5-HT) and assessment of ERK1/2 activation.
  • Use of inhibitors (H89, dominant-negative RasN17, dominant-negative Raf1, dominant-negative B-Raf) to probe signaling pathways.
  • Evaluation of Rap1 activation and the effect of Rap1GAP1.

Main Results:

  • 5-HT induced activation of ERK1/2 in transfected HEK293 cells.
  • This activation was sensitive to H89, indicating a role for PKA.
  • ERK1/2 activation was independent of epidermal growth factor receptor (EGFR) transactivation.
  • 5-HT stimulated both Ras and Rap1 activation.
  • ERK1/2 activation was abolished by dominant-negative Ras (RasN17) but not influenced by Rap1GAP1.
  • Dominant-negative Raf1 and B-Raf partially reduced ERK1/2 activation.

Conclusions:

  • The human G(s)-coupled serotonin receptors 5-HT(4(b)) and 5-HT(7(a)) activate ERK1/2 in HEK293 cells.
  • This signaling cascade is dependent on Ras and involves PKA.
  • The pathway is independent of Rap1 and EGFR transactivation, suggesting a distinct signaling mechanism.

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