Calcium mobilization stimulatesDictyostelium discoideumshear-flow-induced cell motility

Abstract

Application of hydrodynamic mild shear stress to adherent Dictyostelium discoideum vegetative cells triggers active actin cytoskeleton remodeling resulting in net cell movement along the flow. The average cell speed is strongly stimulated by external calcium (Ca2+, K50%=22 μM), but the directionality of the movement is almost unaffected. This calcium concentration is ten times higher than the one promoting cell adhesion to glass surfaces (K50%=2 μM). Addition of the calcium chelator EGTA or the Ca2+-channel blocker gadolinium (Gd3+) transiently stops cell movement. Monitoring the evolution of cell-surface contact area with time reveals that calcium stimulates cell speed by increasing the amplitude of both protrusion and retraction events at the cell edge, but not the frequency. As a consequence, with saturating external calcium concentrations, cells are sensitive to very low shear forces (20 pN; σ=0.1 Pa). Moreover, a null-mutant lacking the unique Gβ subunit does not respond to external Ca2+ changes (K50%>1000 μM), although the directionality of the movement is comparable with that of wild-type cells. Furthermore, cells lacking the inositol 1,4,5-trisphosphate receptor (IP3-receptor) exhibit a markedly reduced Ca2+ sensitivity. Thus, calcium release from internal stores and calcium entry through the plasma membrane modulate cell speed in response to shear stress.

Chemical and mechanical stimuli act on common signal transduction and cytoskeletal networks

“…In fact, purified G proteins in phospholipid vesicles were activated in response to acute shear stress in the absence of other protein components (23). However, cells lacking β or γ subunits of G proteins migrate more slowly randomly, in chemotaxis, electrotaxis, and shear flow, and are deficient in phagocytosis, suggesting that the defect in mechanosensitivity might be related to the decreased baseline activity (13,(24)(25)(26).…”

“…Fache et al previously reported reduced shear-flow-induced motility for cells lacking G β (13). Thus, we examined whether heterotrimeric G proteins are involved in the response to acute stimulation with shear flow.…”

Computational modelling suggests dynamic interactions between Ca2+, IP3 and G protein-coupled modules are key to robust Dictyostelium aggregation

“…All Michaelis constants are also dimensionalized by d: (1) in the normalized form is transformed as follows:…”

“…Amongst many other factors, intracellular Ca 2+ has been shown to modulate both the speed and direction of Dictyostelium motility. 1 In an attempt to shed light on the above issues, we employed our model to develop the following testable hypothesis. We sought to investigate whether or not the Ca 2+ surges caused by the application of calmidazolium added at different time points of the oscillatory cycle would result in similar phase delays in optical density oscillations to those observed experimentally in ref.…”

“…It has been shown experimentally that the levels of intracellular Ca 2+ and cAMP in Dictyostelium are tightly interconnected [24][25][26][27] and a number of experiments involving the application of calmidazolium (R24571), a calmodulin (CaM) inhibitor, to Dictyostelium have demonstrated a dramatic impact on the light scattering oscillations 26,28 which are one of the major characteristics of aggregation. 26,29 Ca 2+ has also been shown to be directly involved in cell migration via the stretch-activated Ca 2+ channels (SAC)s. 1 Previously developed models for Dictyostelium signalling and aggregation do not allow these phenomena to be investigated, however, as they all omit one or more of the networks involved. To address this issue, we have employed a modular approach to develop a new model for cAMP oscillations in Dictyostelium that explicitly incorporates networks involving intracellular calcium (Ca 2+ ), inositol 1,4,5-triphosphate (IP 3 ) and the G protein coupled receptor cAR1.…”