Abstract
P21-activated kinase 1 (PAK1) is activated by binding to GTP-bound Rho GTPases Cdc42 and Rac via its CRIB domain. Here, we provide evidence that S79 in the CRIB domain of PAK1 is not directly involved in this binding but is crucial for PAK1 activation. S79A mutation reduces the binding affinity of PAK1 for the GTPases and inhibits autophosphorylation and kinase activity of PAK1. Thus, this mutation abrogates the ability of PAK1 to induce changes in cell morphology and motility and to promote malignant transformation of prostate epithelial cells. We also show that growth of the prostate cancer cell line PC3 is inhibited by the treatment of a PAK1-inhibiting peptide comprising 19 amino acids centered on S79, but not by the PAK1 peptide containing the S79A mutation, and that this growth inhibition is correlated with reduced autophosphorylation activity of PAK1. Together, these findings demonstrate a significant role of S79 in PAK1 activation and provide evidence for a novel mechanism of the CRIB-mediated interaction of PAK1 with Cdc42 and Rac.
Highlights
P21-activated kinase 1 (PAK1) is a major downstream effector of the Rho-GTPases Cdc42 and Rac, which act as molecular switches that transduce various extracellular signals into intracellular responses [1]
PAK1 interacts with Cdc42 and Rac via the CRIB domain [1]
Our results show that S79A mutation significantly decreases the phosphorylation of the three residues (Fig. 1C) and kinase activity of PAK1 toward the PAK1 substrates MBP and DLC1 peptide (Fig. 1D)
Summary
PAK1 is a major downstream effector of the Rho-GTPases Cdc and Rac, which act as molecular switches that transduce various extracellular signals into intracellular responses [1]. PAK1, the best-characterized member of the PAK family, forms a transinhibited dimer in its inactive state, in which the catalytic domain of one PAK1 monomer is blocked by the autoinhibitory domain (AID) of the other [2,3]. PAK1 is recruited to the plasma membrane via the SH3-containing proteins Nck and Grb2 [9,10,11], where it may be activated by signaling molecules such as PDK1 kinase [12], sphingosine [13,14] and PIP2 [15] in a manner independent of the GTPases. There has been a rapid expansion in the development of peptides as potential drugs for cancer therapy over the last decade [23]
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