PI(3,4)P2 and microtubule-dependent integrin endocytosis regulates invasive cell migration

Abstract

The dynamic assembly and disassembly of cell adhesions modulate cell movements. Adhesions are formed as integrin-mediated structures and include lamellipodia, focal adhesions, filopodia, and podosomes/invadopodia. Src-transformed mouse embryonic fibroblasts (MEFs) develop invadopodium arcs at the ventral membrane. Invadopodium turnover requires precise regulation of endocytosis and recycling of integrin receptors on the surface of the cell membrane. Endocytosis and exocytosis of integrins are coordinated by the active behaviour of actomyosin and microtubules. Recently, studies demonstrated that phosphoinositides such as PI(4,5)P2, PI(3,4,5)P3, PI(3,4)P2, and PI3P were also closely correlated with membrane receptor recycling. Researchers have revealed that the invadopodia formed by src-transformed fibroblasts and cancer cells exhibit abundant PI(3,4)P2; however, the specific functions of PI(3,4)P2 in adhesion dynamics are unclear. In this study, I observed repeatedly occurring PI(3,4)P2-enriched buds/tubules formation at the trailing edge of the classic invadopodium arc formed by src-transformed MEFs. PI(3,4)P2 is enriched at invadopodium arcs and locally generated by PIK3CA and SHIP2. Posterior to the invadopodium arc, the PI(3,4)P2 effector SNX9 triggers PI(3,4)P2-rich compartment formation of micrometre-long tubular membrane invaginations and recruitment of DNM2 to the base of membrane buds/tubules. The PI(3,4)P2-enriched membrane buds/tubules are organized by microtubules reaching into the inner part of the arc. The buds/tubules colocalize with dynein and microtubule adaptors. The microtubule-destabilizing agent nocodazole hinders PI(3,4)P2-enriched membrane buds/tubules formation behind the arcs. PI(3,4)P2 biogenesis and invaginations are associated with membrane receptor endocytosis and posterior invadopodium turnover. The receptor integrin β3 is enriched in PI(3,4)P2-containing membrane tubules and sorted in the endolysosomal network through PI(3,4)P2-enriched tubules. Suppression of PI(3,4)P2 enrichment inhibits integrin β3 internalization by the invadopodium and impedes invasive cell migration. Thus, microtubule-driven PI(3,4)P2-enriched membrane invaginations into the invadopodia promote integrin endocytosis and cell invasion. (281 words)