Models of the cooperative mechanism for Rho effector recognition: implications for RhoA-mediated effector activation - PubMed (original) (raw)

. 2004 Dec 17;279(51):53419-26.

doi: 10.1074/jbc.M409551200. Epub 2004 Oct 8.

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Models of the cooperative mechanism for Rho effector recognition: implications for RhoA-mediated effector activation

Lars Blumenstein et al. J Biol Chem. 2004.

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Abstract

Activated GTPases of the Rho family regulate a spectrum of functionally diverse downstream effectors, initiating a network of signal transduction pathways by interaction and activation of effector proteins. Although effectors are defined as proteins that selectively bind the GTP-bound state of the small GTPases, there have been also several indications for a nucleotide-independent binding mode. By characterizing the molecular mechanism of RhoA interaction with its effectors, we have determined the equilibrium dissociation constants of several Rho-binding domains of three different effector proteins (Rhotekin, ROCKI/ROK beta/p160ROCK, PRK1/PKNalpha where ROK is RhoA-binding kinase) for both RhoA.GDP and RhoA.GTP using fluorescence spectroscopy. In addition, we have identified two novel Rho-interacting domains in ROCKI, which bind RhoA with high affinity but not Cdc42 or Rac1. Our results, together with recent structural data, support the notion of multiple effector-binding sites in RhoA and strongly indicate a cooperative binding mechanism for PRK1 and ROCKI that may be the molecular basis of Rho-mediated effector activation.

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