Masato Nakagawa

Rac1 and Cdc42 Capture Microtubules through IQGAP1 and CLIP-170

Linkage of microtubules to special cortical regions is essential for cell polarization. CLIP-170 binds to the growing ends of microtubules and plays pivotal roles in orientation. We have found that IQGAP1, an effector of Rac1 and Cdc42, interacts with CLIP-170. In Vero fibroblasts, IQGAP1 localizes at the polarized leading edge. Expression of carboxy-terminal fragment of IQGAP1, which includes the CLIP-170 binding region, delocalizes GFP-CLIP-170 from the tips of microtubules and alters the microtubule array. Activated Rac1/Cdc42, IQGAP1, and CLIP-170 form a tripartite complex. Furthermore, expression of an IQGAP1 mutant defective in Rac1/Cdc42 binding induces multiple leading edges. These results indicate that Rac1/Cdc42 marks special cortical spots where the IQGAP1 and CLIP-170 complex is targeted, leading to a polarized microtubule array and cell polarization.

Roles of Rho-family GTPases in cell polarisation and directional migration.

Polarised cell migration is a tightly regulated process that occurs in tissue development, chemotaxis and wound healing. Rho-family GTPases, including Cdc42, Rac1 and RhoA, play a central role in establishing cell polarisation, which requires asymmetric and ordered distribution of the signalling molecules and the cytoskeleton. Recent advances reveal that Rho GTPases, together with phosphatidylinositol 3-kinase, contribute to asymmetric phosphatidylinositol 3,4,5-trisphosphate distribution via a positive-feedback loop. Phosphatidylinositol 3,4,5-trisphosphate thereby activates the signalling cascades to the cytoskeleton as a second messenger. Rho GTPases also capture and stabilise microtubules through their effectors (e.g. IQGAP1, mDia and Par6) near the cell cortex, leading to polarised cell morphology and directional cell migration. Thus, elucidation of the signal transduction cascades from receptors to Rho GTPases and, subsequently, from Rho GTPases to microtubules has begun.

Cdc42 and Rac1 Regulate the Interaction of IQGAP1 with β-Catenin

IQGAP1, a target of Cdc42 and Rac1 small GTPases, directly interacts with β-catenin and negatively regulates E-cadherin-mediated cell-cell adhesion by dissociating α-catenin from the cadherin-catenin complex in vivo (Kuroda, S., Fukata, M., Nakagawa, M., Fujii, K., Nakamura, T., Ookubo, T., Izawa, I., Nagase, T., Nomura, N., Tani, H., Shoji, I., Matsuura, Y., Yonehara, S., and Kaibuchi, K. (1998) Science 281, 832–835). Here we investigated how Cdc42 and Rac1 regulate the IQGAP1 function. IQGAP1 interacted with the amino-terminal region (amino acids 1–183) of β-catenin, which contains the α-catenin-binding domain. IQGAP1 dissociated α-catenin from the β-catenin-α-catenin complex in a dose-dependent manner in vitro. Guanosine 5′-(3-O-thio)triphosphate (GTPγS)·glutathioneS-transferase (GST)-Cdc42 and GTPγS·GST-Rac1 inhibited the binding of IQGAP1 to β-catenin in a dose-dependent manner in vitro, whereas neither GDP·GST-Cdc42, GDP·GST-Rac1, nor GTPγS·GST-RhoA did. The coexpression of dominant active Cdc42 with IQGAP1 suppressed the dissociation of α-catenin from the cadherin-catenin complex induced by the overexpression of IQGAP1 in L cells expressing E-cadherin (EL cells). Consistent with this, the overexpression of either dominant negative Cdc42 or Rac1 resulted in the reduction of E-cadherin-mediated cell adhesive activity in EL cells. These results indicate that Cdc42 and Rac1 negatively regulate the IQGAP1 function by inhibiting the interaction of IQGAP1 with β-catenin, leading to stabilization of the cadherin-catenin complex.

Regulation of cadherin-mediated cell-cell adhesion by the Rho family GTPases

Reports in the past two years have shown that Cdc42, Rac1, and Rho - belonging to the Rho small GTPase family - participate in the regulation of cadherin-mediated cell-cell adhesion. IQGAP1, an effector of Cdc42 and Rac1, interacts with cadherin and beta-catenin and induces the dissociation of alpha-catenin from the cadherin-catenins complex leading to disruption of cell-cell adhesion: activated Cdc42 and Rac1 counteract the effect of IQGAP1. Thus, Cdc42 and Rac1 appear to regulate cadherin-mediated cell-cell adhesion acting through IQGAP1.

Regulation of E-Cadherin-Mediated Cell-Cell Adhesion by Rho Family GTPases

Cell-cell adhesions are rearranged dynamically during tissue development and tumor metastasis. Recently, Rho-family GTPases, including RhoA, Rac1, and Cdc42, have emerged as key regulators of cadherin-mediated cell-cell adhesion. Following the identification and characterization of regulators and effectors of Rho GTPases, signal transduction pathways from cadherin to Rho GTPases and, in turn, from Rho GTPases to cadherin are beginning to be clarified.