Constitutive ERK1/2 activation by a chimeric neurokinin 1 receptor-beta-arrestin1 fusion protein. Probing the composition and function of the G protein-coupled receptor "signalsome" |
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Authors: | Jafri Farahdiba El-Shewy Hesham M Lee Mi-Hye Kelly Margaret Luttrell Deirdre K Luttrell Louis M |
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Institution: | Department of Medicine, Medical University of South Carolina, Charleston, South Carolina 29425, USA. |
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Abstract: | The beta-arrestins, a small family of G protein-coupled receptor (GPCR)-binding proteins involved in receptor desensitization, have been shown to bind extracellular signal-regulated kinases 1 and 2 (ERK1/2) and function as scaffolds for GPCR-stimulated ERK1/2 activation. To better understand the mechanism of beta-arrestin-mediated ERK1/2 activation, we compared ERK1/2 activation by the wild-type neurokinin 1 (NK1) receptor with a chimeric NK1 receptor having beta-arrestin1 fused to the receptor C terminus (NK1-betaArr1). The NK1 receptor couples to both G(s) and G(q/11), resides on the plasma membrane, and mediates rapid ERK1/2 activation and nuclear translocation in response to neurokinin A. In contrast, NK1-betaArr1 is a G protein-uncoupled "constitutively desensitized" receptor that resides almost entirely in an intracellular endosomal compartment. Despite its inability to respond to neurokinin A, we found that NK1-betaArr1 expression caused robust constitutive activation of cytosolic ERK1/2 and that endogenous Raf, MEK1/2, and ERK1/2 coprecipitated in a complex with NK1-betaArr1. While agonist-dependent ERK1/2 activation by the NK1 receptor was independent of protein kinase A (PKA) or PKC activity, NK1-betaArr1-mediated ERK1/2 activation was completely inhibited when basal PKA and PKC activity were blocked. In addition, the rate of ERK1/2 dephosphorylation was slowed in NK1-betaArr1-expressing cells, suggesting that beta-arrestin-bound ERK1/2 is protected from mitogen-activated protein kinase phosphatase activity. These data suggest that beta-arrestin binding to GPCRs nucleates the formation of a stable "signalsome" that functions as a passive scaffold for the ERK1/2 cascade while confining ERK1/2 activity to an extranuclear compartment. |
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