The action of guanine nucleotide exchange factors (GEFs) on the ADP-Ribosylation Factor (ARF) family of small GTPases initiates intracellular transport pathways. This role requires ARF GEFs to be recruited from the cytosol to intracellular membrane compartments. An ARF GEF known as General receptor for 3-phosphoinositides 1 (Grp1) is recruited to the plasma membrane through its pleckstrin homology (PH) domain that recognizes phosphatidylinositol 3,4,5-trisphosphate (PIP3). Here, we find that the phosphorylation of Grp1 induces its PH domain to recognize instead phosphatidylinositol 4-phosphate (PI4P). This phosphorylation also releases an auto-inhibitory mechanism that results in the coil-coil (CC) domain of Grp1 engaging two peripheral membrane proteins of the recycling endosome. Because the combination of these actions results in Grp1 being recruited preferentially to the recycling endosome rather than to the plasma membrane, our findings reveal the complexity of recruitment mechanisms that need to be coordinated in localizing an ARF GEF to an intracellular compartment to initiate a transport pathway. Our elucidation is also remarkable for having revealed that phosphoinositide recognition by a PH domain can be switched through its phosphorylation.

鸟嘌呤核苷酸交换因子（GEF）对小GTPase的ADP-核糖基化因子（ARF）家族的作用启动了细胞内转运途径。该角色要求将ARF GEF从细胞质中募集到细胞内膜区室中。通过识别磷脂酰肌醇3,4,5-三磷酸（PIP3）的pleckstrin同源性（PH）域，将称为3-磷酸肌醇1（Grp1）通用受体的ARF GEF募集到质膜。在这里，我们发现Grp1的磷酸化诱导其PH结构域，而不是识别磷脂酰肌醇4-磷酸（PI4P）。这种磷酸化作用还释放了一种自动抑制机制，该机制导致Grp1的卷曲螺旋（CC）域与回收内体的两个外围膜蛋白结合。因为这些动作的组合导致Grp1被优先招募到再循环内体而不是质膜，所以我们的发现揭示了招募机制的复杂性，在将ARF GEF定位到细胞内区室以启动转运途径时需要进行协调。我们的说明也很出色，因为它揭示了PH结构域对磷酸肌醇的识别可以通过其磷酸化来切换。