Formation and closure of macropinocytic cups in Dictyostelium

Macropinocytosis is a conserved endocytic process by which cells engulf droplets of medium into micron-sized vesicles. We use light-sheet microscopy to define an underlying set of principles by which macropinocytic cups are shaped and closed in Dictyostelium amoebae. Cups form around domains of PIP3 stretching almost to their lip and are supported by a specialized F-actin scaffold from lip to base. They are shaped by a ring of actin poly-merization created by recruiting Scar/WAVE and Arp2/3 around PIP3 domains, but how cups evolve overtime to close and form a vesicle is unknown. Custom 3D analysis shows that PIP3 domains expand from small origins, capturing new membrane into the cup, and crucially, that cups close when domain expansion stalls. We show that cups can close in two ways: either at the lip, by inwardly directed actin polymerization, or the base, by stretching and delamination of the membrane. This provides the basis for a conceptual mechanism whereby closure is brought about by a combination of stalled cup expansion, continued actin polymerization at the lip, and membrane tension. We test this through the use of a biophysical model, which can recapitulate both forms of cup closure and explain how 3D cup structures evolve over time to mediate engulfment.

Automated summary

This study reveals the underlying principles of macropinocytic cup formation and closure in Dictyostelium amoebae using light-sheet microscopy. Cups are formed by recruiting PIP3 domains and a specialized F-actin scaffold, and they close through inward or outward actin polymerization and membrane stretching and delamination when cup expansion stalls. This research is important for understanding intracellular fluid engulfment processes.