PI3K regulates endocytosis after insulin secretion via signaling crosstalk between Arf6 and Rab27a

Abstract

In secretory cells, endocytosis is coupled to exocytosis to enable proper secretion. Although endocytosis is crucial to maintain cellular homeostasis before and after secretion, knowledge about secretagogue-induced endocytosis in secretory cells is still limited. Here, we searched for Rab27a GTPase-activating protein (GAP) EPI64-interacting proteins and identified the Arf6 guanine-nucleotide-exchange factor (GEF) ARNO in pancreatic beta-cells. We found that the insulin secretagogue glucose promotes phosphatidylinositol-(3,4,5)-trisphosphate (PIP3) generation through phosphoinositide 3-kinase (PI3K), thereby recruiting ARNO to the intracellular side of the plasma membrane. Peripheral ARNO promotes clathrin assembly through its GEF activity for Arf6 and regulates the early stage of endocytosis. We also found that peripheral ARNO recruits EPI64 to the same area and that the interaction requires glucose-induced endocytosis in pancreatic beta-cells. Since GTP- and GDP-bound Rab27a regulate exocytosis and the late stage of endocytosis, our results indicate that the glucose-induced activation of PI3K plays a pivotal role in exocytosis-endocytosis coupling, and that ARNO and EPI64 regulate endocytosis at distinct stages.