Department of Biochemistry and Microbiology, University of Victoria, Victoria, BC V8W 2Y2, Canada.
Department of Chemistry and Biochemistry, University of Oregon, Eugene, OR 97403, USA; Institute of Molecular Biology, University of Oregon, Eugene, OR 97403, USA.
Cell Rep. 2023 Mar 28;42(3):112172. doi: 10.1016/j.celrep.2023.112172. Epub 2023 Feb 26.
Class IB phosphoinositide 3-kinase (PI3Kγ) is activated in immune cells and can form two distinct complexes (p110γ-p84 and p110γ-p101), which are differentially activated by G protein-coupled receptors (GPCRs) and Ras. Using a combination of X-ray crystallography, hydrogen deuterium exchange mass spectrometry (HDX-MS), electron microscopy, molecular modeling, single-molecule imaging, and activity assays, we identify molecular differences between p110γ-p84 and p110γ-p101 that explain their differential membrane recruitment and activation by Ras and GPCRs. The p110γ-p84 complex is dynamic compared with p110γ-p101. While p110γ-p101 is robustly recruited by Gβγ subunits, p110γ-p84 is weakly recruited to membranes by Gβγ subunits alone and requires recruitment by Ras to allow for Gβγ activation. We mapped two distinct Gβγ interfaces on p101 and the p110γ helical domain, with differences in the C-terminal domain of p84 and p101 conferring sensitivity of p110γ-p101 to Gβγ activation. Overall, our work provides key insight into the molecular basis for how PI3Kγ complexes are activated.
I 类磷酸肌醇 3-激酶（PI3Kγ）在免疫细胞中被激活，并且可以形成两种不同的复合物（p110γ-p84 和 p110γ-p101），这两种复合物分别被 G 蛋白偶联受体（GPCRs）和 Ras 激活。我们使用 X 射线晶体学、氘氢交换质谱（HDX-MS）、电子显微镜、分子建模、单分子成像和活性测定的组合，确定了 p110γ-p84 和 p110γ-p101 之间的分子差异，这些差异解释了它们通过 Ras 和 GPCR 进行的差异膜募集和激活。与 p110γ-p101 相比，p110γ-p84 复合物是动态的。虽然 p110γ-p101 被 Gβγ 亚基强烈募集，但 p110γ-p84 仅被 Gβγ 亚基弱募集到膜上，并且需要 Ras 的募集来允许 Gβγ 激活。我们在 p101 和 p110γ 螺旋域上绘制了两个不同的 Gβγ 界面，p84 和 p101 的 C 末端结构域的差异赋予了 p110γ-p101 对 Gβγ 激活的敏感性。总体而言，我们的工作提供了关键的见解，阐明了 PI3Kγ 复合物如何被激活的分子基础。
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