More importantly, both Ras inhibitors regulate the sensitivity of the cells. discoideum and human neutrophils, respectively. Recently, it has been revealed that C2GAP1 and CAPRI control the GPCR-mediated adaptation in D. Ras activation is the first step in the chemosensing GPCR signaling pathways that displays a transient activation behavior in both model organism Dictyostelium discoideum and mammalian neutrophils. Thus, adaptation provides a fundamental strategy for eukaryotic cells to chemotax through a gradient. Cells in adaptation no longer respond to the present stimulus but remain sensitive to stronger stimuli. Eukaryotic cells sense chemoattractant with G protein-coupled receptors (GPCRs) and chemotax toward gradients with an enormous concentration range through adaptation. We propose that key signalling network components act on, and are in turn acted upon, by surface charge, closing feedback loops, which bring about the global-scale molecular self-organization required for spontaneous protrusion formation, cell migration and polarity establishment.Ĭhemotaxis plays an essential role in recruitment of leukocytes to sites of inflammation. Computational simulations involving excitable biochemical networks demonstrated that slight changes in feedback loops, induced by recruitment of the charged actuators, could lead to outsized effects on system activation. Conversely, increasing the negative surface charge deactivated the network and locally suppressed chemoattractant-induced protrusions or subverted EGF-induced ERK activation. These effects were blocked by genetic or pharmacological inhibition of key signalling components such as AKT and PI3K/TORC2. Abruptly reducing the surface charge by recruiting positively charged optogenetic actuators was sufficient to trigger the entire biochemical network, initiate de novo protrusions and abrogate pre-existing polarity. Rapid alterations of inner leaflet anionic phospholipids-such as PI(4,5)P2, PI(3,4)P2, phosphatidylserine and phosphatidic acid-collectively contribute to the surface charge changes. Surface charge, or zeta potential, is transiently lowered at new protrusions and within cortical waves of Ras/PI3K/TORC2/F-actin network activation. Here we show that the regulation of negative surface charge on the inner leaflet of the plasma membrane plays an integrative role in the molecular interactions. Although biochemical and genetic analyses have delineated many specific interactions, how the activation and localization of so many different molecules are spatiotemporally orchestrated at the subcellular level has remained unclear. Scale bars: 10 mm.ĭuring cell migration and polarization, numerous signal transduction and cytoskeletal components self-organize to generate localized protrusions. GFP and RFP/TRITC channels are shown in the rows beneath. Composite images are shown in the top rows. Cells were plated on coverslips in buffer and behaviour recorded. Pten 2 cells were rescued with PTEN-RFP and coexpress PhdA-GFP as a marker for PtdIns(3,4,5)P 3 and possibly other lipid products of PI3-kinases. The green structures in the lower right are macropinosomes formed in a cell not expressing PTENRFP. An asterisk serves as a spatial landmark. PhdA-GFP stays associated with the forming vesicle, but is lost from the leading edge. (B) Macropinocytosis/motility: PhdA-GFP forms patches, and even those aligned with the direction of general movement, develop into macropinocytic vesicles. At 0 seconds contact with a yeast cell (marked with an asterisk) is established and as it is subsequently taken up, PhdA-GFP extends from the base of the cup all around the particle. Some yeast cells (red particles) have already been taken up.
(A) Phagocytosis: a Dictyostelium cell takes up TRITC-labelled yeast. PtdIns(3,4,5)P 3 (PIP3)-labelled structures in growing cells.
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