Neurofibromin controls macropinocytosis and phagocytosis in Dictyostelium

Abstract

Cells use phagocytosis and macropinocytosis to internalise bulk material, which in phagotrophic organisms supplies the nutrients necessary for growth. Wildtype Dictyostelium amoebae feed on bacteria, but for decades laboratory work has relied on axenic mutants that can also grow on liquid media. We used forward genetics to identify the causative gene underlying this phenotype. This gene encodes the RasGAP Neurofibromin (NF1). Loss of NF1 enables axenic growth by increasing fluid uptake. Mutants form outsized macropinosomes which are promoted by greater Ras and PI3K activity at sites of endocytosis. Relatedly, NF1 mutants can ingest larger-than-normal particles using phagocytosis. An NF1 reporter is recruited to nascent macropinosomes, suggesting that NF1 limits their size by locally inhibiting Ras signalling. Our results link NF1 with macropinocytosis and phagocytosis for the first time, and we propose that NF1 evolved in early phagotrophs to spatially modulate Ras activity, thereby constraining and shaping their feeding structures.DOI: http://dx.doi.org/10.7554/eLife.04940.001

WASH drives early recycling from macropinosomes and phagosomes to maintain surface phagocytic receptors

“…As a genetically tractable professional phagocyte, Dictyostelium has been well used for studies on phagosome maturation, and the pathways involved are highly conserved with those in mammalian macrophages (33)(34)(35). Much less is known about macropinosome maturation, but the large and frequent macropinosomes formed by laboratory strains of Dictyostelium make it ideal for investigations (2). Here we show that without WASH, cells are unable to prevent the degradation of the plasma membrane proteins captured by macropinocytosis.…”

“…This process most likely originally evolved in unicellular protists such as the amoeba Dictyostelium discoideum as a means to obtain nutrients from the environment (1)(2)(3). Although this process is not necessary in most cells in multicellular organisms, macropinocytosis can still be induced in many mammalian cell lines (4)(5)(6) and is constitutively up-regulated in Ras-transformed cancer cells where it performs its original role in capturing nutrients (7).…”

Uses and abuses of macropinocytosis

“…However, most of the widely used strains carry mutations that promote growth in these media. When the key mutation was finally identified, it was found to reside in the RasGAP and tumour suppressor gene NF1 (Bloomfield et al, 2015). Loss of NF1 results in increased Ras activity at sites of largescale endocytosis, resulting in enhanced ruffling, increased rates of macropinocytosis and larger macropinosomes (Fig.…”

“…The injection of activated (oncogenic) Ras protein into fibroblasts causes ruffling and macropinocytosis (Bar-Sagi and Feramisco, 1986; see also below). Genetic ablation of certain Ras proteins inhibits macropinocytosis in Dictyostelium amoebae (Chubb et al, 2000;Hoeller et al, 2013), and loss of the Ras GTPase-activating protein (RasGAP) NF1 increases the frequency of macropinocytosis and size of macropinosomes in these cells (Bloomfield et al, 2015; see also below). Ras activates class-I phosphatidylinositol 3-kinases (PI3Ks) through their Rasbinding domain, and these enzymes produce the membrane lipid PIP3 (Rodriguez-Viciana et al, 1994), which is an ether-linked plasmanyl inositol in Dictyostelium (Clark et al, 2014).…”

Phagocytosis-dependent and independent mechanisms underlie the microglial cell damage caused by carbon nanotube agglomerates

“…Cells internalize larger particles by phagocytosis or macropinocytosis (> 1 μm), and smaller particles by endocytosis (60-120 nm) (May et al, 2000;Conner and Schmid, 2003). Phagocytosis and macropinocytosis are closely related processes (Swanson, 2008(Swanson, , 2014, however, the regulatory architecture of these processes are not fully understood (Bloomfield et al, 2015). These processes are triggered by the interaction between the particles and various kinds of receptors (Jang et al, 2009;Ramirez-Ortiz et al, 2013) As for CNTs, phagocytosis of SWCNTs by macrophages (Dong et al, 2013) and macropinocytosis of MWCNTs through macrophage receptor with collagenous structure (MARCO) (Hirano et al, 2012) have been reported, while the interaction of CNTs with scavenger receptors (ScRs), which are important for both phagocytosis and micropinocytosis, has also been reported (Wang et al, 2012).…”