van Horssen Remco, Janssen Edwin, Peters Wilma, van de Pasch Loes, Lindert Mariska M Te, van Dommelen Michiel M T, Linssen Peter C, Hagen Timo L M Ten, Fransen Jack A M, Wieringa Bé
Department of Cell Biology, Nijmegen Center for Molecular Life Sciences and Department of Hematology, Radboud University Nijmegen Medical Center, 6500 HB Nijmegen, The Netherlands.
J Biol Chem. 2009 Jan 16;284(3):1620-7. doi: 10.1074/jbc.M806974200. Epub 2008 Nov 12.
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.
ATP是新陈代谢中的“主要能量货币”，也是细胞活动中最为通用的小分子调节因子。尽管人们已经对ATP在生命系统基本过程中的作用有了很多了解，但关于其区域化的数据却相当稀少，而且我们对于细胞内ATP浓度分布模式（“ATP不均匀性”）是否存在及其是否具有调节作用的理解仍然非常有限。在此，我们报告了关于局部ATP供应与肌动球蛋白行为调节之间耦合关系的分析，肌动球蛋白行为是一个广泛且动态的过程，对ATP具有明显的高度依赖性，它对于细胞形状变化和细胞运动至关重要。作为实验模型，我们使用了来自敲除小鼠的胚胎成纤维细胞，这些小鼠缺乏主要的ATP-ADP交换酶，我们在其中（重新）引入了ATP/ADP交换酶腺苷酸激酶-1（AK1），并通过与不同的人工定位标签偶联来刻意操纵其空间定位。通过对AK1变体进行转染互补并与黄色荧光蛋白对照进行比较，我们发现带有粘着斑引导标签的AK1的细胞中，运动性和铺展性增强。在带有膜靶向或胞质AK1的细胞中观察到了中等程度的增强。在细胞中全局AK1活性恒定的条件下，使用异二聚体诱导方法使AK1瞬时转位至粘着斑，证实了这些结果。基于我们在这些模型系统中的发现，我们提出，当通过参与磷酸转移的酶来维持时，细胞周边的局部ATP供应以及肌动球蛋白机制的“现场”供能，是控制细胞运动的共同决定因素。
