Modulation of Cell Motility by Spatial Repositioning of Enzymatic ATP/ADP Exchange Capacity

Abstract

ATP is the "principal energy currency" in metabolism and the most versatile small molecular regulator of cellular activities. Although already much is known about the role of ATP in fundamental processes of living systems, data about its compartmentalization are rather scarce, and we still have only very limited understanding of whether patterns in the distribution of intracellular ATP concentration ("ATP inhomogeneity") do exist and have a regulatory role. Here we report on the analysis of coupling of local ATP supply to regulation of actomyosin behavior, a widespread and dynamic process with conspicuous high ATP dependence, which is central to cell shape changes and cell motility. As an experimental model, we use embryonic fibroblasts from knock-out mice without major ATP-ADP exchange enzymes, in which we (re)introduce the ATP/ADP exchange enzyme adenylate kinase-1 (AK1) and deliberately manipulate its spatial positioning by coupling to different artificial location tags. By transfection-complementation of AK1 variants and comparison with yellow fluorescent protein controls, we found that motility and spreading were enhanced in cells with AK1 with a focal contact guidance tag. Intermediary enhancement was observed in cells with membrane-targeted or cytosolic AK1. Use of a heterodimer-inducing approach for transient translocation of AK1 to focal contacts under conditions of constant global AK1 activity in the cell corroborated these results. Based on our findings with these model systems, we propose that local ATP supply in the cell periphery and "on site" fuelling of the actomyosin machinery, when maintained via enzymes involved in phosphoryl transfer, are codetermining factors in the control of cell motility.Maintenance of adequate ATP supply is of crucial importance for the mechanisms of structural remodeling in cells with high shape plasticity, especially under high energy-demanding circumstances (1). During processes like cell motility or phagocytosis, the cell movement and cellular shape changes require active restructuring of the actin cytoskeleton. By spatially controlled polymerization of ATP-bound G-actin monomers to the plus end of growing actin filaments and formation of multiple branches, a dense network is built, which is called the actin cortex, based on its specific localization within the cell (2). In this network, ATP hydrolysis and release of P i drive actin filament dynamics by modulating filament stability and determine nucleotide-dependent filament conformation and interaction(s) with regulatory actin-binding proteins (3). Furthermore, force generation needed for contraction and cell movement is controlled by myosin and nonmuscle myosin ATPases (4), and also upstream signaling, involving small GTPases, is contingent upon nucleotide exchange (5). Taken together, actomyosin dynamics is overall an energy-demanding process, directly coupled to ATP availability.Indeed, actomyosin-based processes may consume a major fraction of cellular energy (6). Moreover, the coupling between global ATP suppl...

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