PI3Kα, consisting of the p110α isoform of the catalytic subunit of PI 3-kinase (encoded by PIK3CA) and the p85α regulatory subunit (encoded by PI3KR1) is activated by growth factor receptors. The identification of common oncogenic mutations in PIK3CA has driven the development of many inhibitors that bind to the ATP-binding site in the p110α subunit. Upon activation, PI3Kα undergoes conformational changes that promote its membrane interaction and catalytic activity, yet the effects of ATP-site directed inhibitors on the PI3Kα membrane interaction are unknown. Using FRET and Biolayer Interferometry assays, we show that a class of ATP-site directed inhibitors represented by GSK2126458 block the growth factor activated PI3KαWT membrane interaction, an activity dependent on the ligand forming specific ATP-site interactions. The membrane interaction for hot spot oncogenic mutations that bypass normal p85α regulatory mechanisms was insensitive to GSK2126458, while GSK2126458 could regulate mutations found outside of these hot spot regions. Our data show that the effect of GSK126458 on the membrane interaction requires the enzyme to revert from its growth factor activated state to a basal state. We find that an ATP substrate analogue can increase the wild type PI3Kα membrane interaction, uncovering a substrate based regulatory event that can be mimicked by different inhibitor chemotypes. Our findings, together with the discovery of small molecule allosteric activators of PI3Kα illustrate that PI3Kα membrane interactions can be modulated by factors related to ligand binding both within the ATP site and at allosteric sites.

中文翻译：

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PI3K 酶的 ATP 竞争性抑制剂通过控制膜结合表现出亚型选择性双重作用。


PI3Kα 由 PI 3 激酶催化亚基（由 PIK3CA 编码）的 p110α 亚型和 p85α 调节亚基（由 PI3KR1 编码）组成，被生长因子受体激活。PIK3CA 中常见致癌突变的鉴定推动了许多与 p110α 亚基中 ATP 结合位点结合的抑制剂的开发。激活后，PI3Kα 发生构象变化，促进其膜相互作用和催化活性，但 ATP 位点导向抑制剂对 PI3Kα 膜相互作用的影响尚不清楚。使用 FRET 和生物膜干涉测定法，我们表明以 GSK2126458 为代表的一类 ATP 位点定向抑制剂阻断生长因子激活的 PI3KαWT 膜相互作用，这种活性取决于配体形成特定的 ATP 位点相互作用。绕过正常 p85α 调节机制的热点致癌突变的膜相互作用对GSK2126458不敏感，而 GSK2126458 可以调节在这些热点区域之外发现的突变。我们的数据表明，GSK126458 对膜相互作用的影响要求酶从其生长因子激活状态恢复到基础状态。我们发现 ATP 底物类似物可以增加野生型 PI3Kα 膜相互作用，揭示了一种基于底物的调节事件，该事件可以通过不同的抑制剂化学型来模拟。我们的研究结果以及 PI3Kα 小分子变构激活剂的发现表明，PI3Kα 膜相互作用可以受到 ATP 位点和变构位点配体结合相关因素的调节。