Molecular basis for Gßγ-mediated activation of phosphoinositide 3-kinase γ.

ABSTRACT

The conversion of PIP2 to PIP3 by phosphoinositide 3-kinase γ (PI3Kγ) is a critical step in neutrophil chemotaxis and is essential for metastasis in many types of cancer. PI3Kγ is activated via directed interaction with Gßγ heterodimers released from cell-surface G protein-coupled receptors (GPCRs) responding to extracellular signals. To resolve how Gßγ activates PI3Kγ, we determined cryo-EM reconstructions of PI3Kγ-Gßγ complexes in the presence of various substrates/analogs, revealing two distinct Gßγ binding sites, one on the p110γ helical domain and one on the C-terminal domain of the p101 subunit. Comparison of these complexes with structures of PI3Kγ alone demonstrates conformational changes in the kinase domain upon Gßγ binding similar to those induced by Ras·GTP. Assays of variants perturbing the two Gßγ binding sites and interdomain contacts that change upon Gßγ binding suggest that Gßγ not only recruits the enzyme to membranes but also allosterically controls activity via both sites. Studies in a zebrafish model examining neutrophil migration are consistent with these results. These findings set the stage for future detailed investigation of Gßγ-mediated activation mechanisms in this enzyme family and will aid in developing drugs selective for PI3Kγ.