Convergence of Ras- and Rac-regulated formin pathways is pivotal for phagosome formation and particle uptake in Dictyostelium

Abstract

Macroendocytosis comprising phagocytosis and macropinocytosis is an actin-driven process regulated by small GTPases that depend on the dynamic reorganization of the membrane that protrudes and internalizes extracellular material by cup-shaped structures. To effectively capture, enwrap, and internalize their targets, these cups are arranged into a peripheral ring or ruffle of protruding actin sheets emerging from an actin-rich, nonprotrusive zone at its base. Despite extensive knowledge of the mechanism driving actin assembly of the branched network at the protrusive cup edge, which is initiated by the actin-related protein (Arp) 2/3 complex downstream of Rac signaling, our understanding of actin assembly in the base is still incomplete. In the Dictyostelium model system, the Ras-regulated formin ForG was previously shown to specifically contribute to actin assembly at the cup base. Loss of ForG is associated with a strongly impaired macroendocytosis and a 50% reduction in F-actin content at the base of phagocytic cups, in turn indicating the presence of additional factors that specifically contribute to actin formation at the base. Here, we show that ForG synergizes with the Rac-regulated formin ForB to form the bulk of linear filaments at the cup base. Consistently, combined loss of both formins virtually abolishes cup formation and leads to severe defects of macroendocytosis, emphasizing the relevance of converging Ras- and Rac-regulated formin pathways in assembly of linear filaments in the cup base, which apparently provide mechanical support to the entire structure. Remarkably, we finally show that active ForB, unlike ForG, additionally drives phagosome rocketing to aid particle internalization.

Transmembrane formins as active cargoes of membrane trafficking

“…Formins also contribute to phagocytosis, not only in various metazoan cell types and diverse contexts ( Colucci-Guyon et al , 2005 ; Brandt et al , 2007 ; Lewkowicz et al , 2008 ; Junemann et al , 2016 ), but also in Entamoeba ( Bharadwaj et al ., 2018 , 2021 ) and Dictyostelium ( Körber et al ., 2023 ). Formin-dependent phagosome formation may facilitate microbial pathogen internalization by host cells or spreading across tissues ( Naj et al , 2013 ; Hoffman et al 2014 ; Rengarajan et al , 2016 ; Williams et al , 2016 ; Dhanda et al ., 2021 ).…”

IqgC is a potent regulator of macropinocytosis in the presence of NF1 and its loading to macropinosomes is dependent on RasG

“…Such extensive remodelling of the plasma membrane is achieved by Arp2/3 complex-nucleated dendritic F-actin network that pushes the membrane outwards [ 9 ]. At the same time, the protrusion is suppressed at the base of the cup where actin elongation factors, formins G and B in Dictyostelium , polymerize linear filaments that provide support [ 6 , 10 , 11 ]. Signals that guide cup formation and subsequent maturation of the nascent endosome are provided by the members of the Rho, Ras, Arf, and Rab families of small GTPases [ 6 , 12 ].…”