Directional Sensing Requires Gβγ-Mediated PAK1 and PIXα-Dependent Activation of Cdc42

* move toward higher concentrations of chemoattractants. Summary Cdc42 has been shown to interact with and/or activate proteins known to be involved in cytoskeleton reorgani-Efficient chemotaxis requires directional sensing and zation, including the PAK kinases and Wiscott Aldrich cell polarization. We describe a signaling mechanism syndrome protein (WASP) (Schmidt and Hall, 1998; Bo-involving G␤␥, PAK-associated guanine nucleotide ex-koch, 2000). However, the precise mechanisms by which change factor (PIX␣), Cdc42, and p21-activated kinase Cdc42 regulates cytoskeleton reorganization are not (PAK) 1. This pathway is utilized by chemoattractants clear. There are three highly homologous PAK isoforms to regulate directional sensing and directional migra-that contain a C-terminal kinase domain, a PIX binding tion of myeloid cells. Our results suggest that G␤␥ domain, several proline-rich domains, and an autoregu-binds PAK1 and, via PAK-associated PIX␣, activates latory segment. The autoregulatory segment contains a Cdc42, which in turn activates PAK1. Thus, in this path-CRIB (Cdc42/Rac interaction/binding) domain, dimer-way, PAK1 is not only an effector for Cdc42, but it ization sequences, an inhibitory switch domain, and a also functions as a scaffold protein required for Cdc42 kinase inhibition sequence (Lim et al. sential for the localization of F-actin formation to the domain binds with a very high affinity to PIX guanine leading edge, the exclusion of PTEN from the leading nucleotide exchange factors (GEFs), consisting of PIX␣ edge, directional sensing, and the persistent direc-and PIX␤ (also called Cool1 and Cool2, respectively). tional migration of chemotactic leukocytes. Although PAK and PIX proteins are readily coimmunoprecipitated ligand-induced production of PIP 3 is not required for from unstimulated cells (Manser et al., 1998), suggesting activation of this pathway, PIP 3 appears to localize the that these two proteins are normally associated. PIX␣ activation of Cdc42 by the pathway. has been shown to stimulate nucleotide exchange activity for Cdc42 and Rac in vivo and in vitro (Manser et al.,