Mechanism of multi-site phosphorylation from a ROCK-I: RhoE complex structure

• David Komander ^[1] ; Ritu Garg ^[2] ; Paul T C Wan ^[1] ; Anne J Ridley ^[2] ; David Barford ^[1]
  1. [1] Institute of Cancer Research

     Institute of Cancer Research

     Reino Unido

     
  2. [2] King's College London

     King's College London

     Reino Unido

     
• Localización: EMBO journal: European Molecular Biology Organization, ISSN 0261-4189, Vol. 27, Nº. 23, 2008, págs. 3175-3185
• Idioma: inglés
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• Resumen

  • The ROCK-I serine/threonine protein kinase mediates the effects of RhoA to promote the formation of actin stress fibres and integrin-based focal adhesions. ROCK-I phosphorylates the unconventional G-protein RhoE on multiple N- and C-terminal sites. These phosphorylation events stabilise RhoE, which functions to antagonise RhoA-induced stress fibre assembly. Here, we provide a molecular explanation for multi-site phosphorylation of RhoE from the crystal structure of RhoE in complex with the ROCK-I kinase domain. RhoE interacts with the C-lobe αG helix of ROCK-I by means of a novel binding site remote from its effector region, positioning its N and C termini proximal to the ROCK-I catalytic site. Disruption of the ROCK-I:RhoE interface abolishes RhoE phosphorylation, but has no effect on the ability of RhoE to disassemble stress fibres. In contrast, mutation of the RhoE effector region attenuates RhoE-mediated disruption of the actin cytoskeleton, indicating that RhoE exerts its inhibitory effects on ROCK-I through protein(s) binding to its effector region. We propose that ROCK-I phosphorylation of RhoE forms part of a feedback loop to regulate RhoA signalling.