Abstract
The P-Rex (phosphatidylinositol (3,4,5)-trisphosphate (PIP3)-dependent Rac exchanger) family (P-Rex1 and P-Rex2) of the Rho guanine nucleotide exchange factors (Rho GEFs) activate Rac GTPases to regulate cell migration, invasion, and metastasis in several human cancers. The family is unique among Rho GEFs, as their activity is regulated by the synergistic binding of PIP3 and Gβγ at the plasma membrane. However, the molecular mechanism of this family of multi-domain proteins remains unclear. We report the 1.95 Å crystal structure of the catalytic P-Rex1 DH-PH tandem domain in complex with its cognate GTPase, Rac1 (Ras-related C3 botulinum toxin substrate-1). Mutations in the P-Rex1·Rac1 interface revealed a critical role for this complex in signaling downstream of receptor tyrosine kinases and G protein-coupled receptors. The structural data indicated that the PIP3/Gβγ binding sites are on the opposite surface and markedly removed from the Rac1 interface, supporting a model whereby P-Rex1 binding to PIP3 and/or Gβγ releases inhibitory C-terminal domains to expose the Rac1 binding site.
Highlights
P-Rex1 activity is correlated with tumorigenesis in cancer
The P-Rex11⁄7Rac1 interface is critical for Rac1 activation in breast cancer cell lines
Using structure-guided mutagenesis we have dissected the central role of the P-Rex1 DH domainRac1 interface in promoting Rac1 activation both in vitro and downstream of G protein-coupled receptors (GPCRs) and receptor tyrosine kinases (RTKs) signaling in cancer cell lines
Summary
P-Rex (phosphatidylinositol [3,4,5]-trisphosphate-dependent Rac exchanger-1) activity is correlated with tumorigenesis in cancer. We have limited molecular insight into how Rho GTPases are activated by P-Rex and how P-Rex activity is regulated by the coordinated actions of PIP3, the G␥ subunits, and protein phosphorylation. Using structure-guided mutagenesis we have dissected the central role of the P-Rex DH domainRac interface in promoting Rac activation both in vitro and downstream of GPCR and RTK signaling in cancer cell lines. Our structural data indicate that the PIP3/G␥ binding sites in P-Rex are markedly removed and on the opposite surface from the Rac interface. Together, these data provide insight into the future therapeutic targeting of P-Rex in the treatment of a number of cancers
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