New model for the interaction of IQGAP1 with CDC42 and RAC1

Abstract

The specific and rapid formation of protein complexes, involving IQGAP family proteins, is essential for diverse cellular processes, such as adhesion, polarization, and directional migration. Although CDC42 and RAC1, prominent members of the RHO GTPase family, have been implicated in binding to and activating IQGAP1, the exact nature of this protein-protein recognition process has remained obscure. Here, we propose a mechanistic framework model that is based on a multiple-step binding process, which is a prerequisite for the dynamic functions of IQGAP1 as a scaffolding protein and a critical mechanism in temporal regulation and integration of cellular pathways.

Keywords: CDC42; IQGAP1; RAC1; RHO GTPases; RHO effectors.

Figure 1.

Domain organization of IQGAP1 and different constructs of C-terminal half (left panel) along with individual rate constants of interaction of corresponding proteins with active RAC1 and CDC42 (right panel). In the left panel IQGAP1 domain organization and White et al.⁶ and different constructs used in our previous study (ref. 42) is shown. Right panel shows the binding affinity of IQGAP1 proteins for RAC1/CDC42 proteins analyzed by stopped-flow fluorometry (kinetic measurements) and fluorescence polarization (equilibrium measurements) (adapted from Nouri et al.⁴²). Kinetic conditions provide individual association and dissociation rate constants (k_on and k_off) and determine the dissociation constants (K_d) which is obtained from the ratio k_off/k_on. Equilibrium conditions determine the equilibrium dissociation constants (K_d) directly.

Figure 2.

A proposed multiple-step mechanistic model of IQGAP1 interaction with CDC42 and RAC1. IQGAP1 harbors at least two distinct binding domains. The molecular nature of IQGAP1 interaction with CDC42 partially differs from that with RAC1 particularly with regard to the role of GRD. RGCT contributes with a high affinity binding to the switch regions of the GTP-bound, active (A) CDC42 and (B) RAC1. GRD more selectively recognizes active forms of CDC42 and RAC1 but also binds to other regions adjacent to the switch regions obviously in a nucleotide-independent manner. Equilibrium measurements using fluorescence polarization by Nouri et al.⁴² demonstrated that GRD undergoes a low-affinity interaction with CDC42 but its binding in contrast to RGCT is partially nucleotide dependent. The very C-terminal domain (CT) of IQGAP1 may potentiate the IQGAP1 interaction with RAC1 and CDC42 proteins by probably extending the resident time of the respective proteins complexes. Lack of a tight GRD interaction with RAC1 (B) may be compensated by calmodulin as an accessory protein,^46,47 which has been reported to bind the polybasic region of RAC1 and IQ motifs of IQGAP1.