Weinberg R A, Zusman D R
Department of Molecular and Cell Biology, University of California, Berkeley 94720.
J Bacteriol. 1990 May;172(5):2294-302. doi: 10.1128/jb.172.5.2294-2302.1990.
One of the signals that has been reported to be important in stimulating fruiting body formation of Myxococcus xanthus is starvation for phosphate. We therefore chose to study phosphatase activity during M. xanthus development. Many phosphatases can cleave the substrate p-nitrophenol phosphate. Using this substrate in buffers at various pHs, we obtained a profile of phosphatase activities during development and germination of M. xanthus. These experiments indicated that there are five patterns of phosphatase activity in M. xanthus: two vegetative and three developmental. The two uniquely vegetative activities have pH optima at 7.2 and 8.5. Both require magnesium and both are inhibited by the reducing agent dithiothreitol. The developmental (spores) patterns of activity have pH optima of 5.2, 7.2, and 8.5. All three activities are Mg independent. Only the alkaline phosphatase activity is inhibited by dithiothreitol. The acid phosphatase activity is induced very early in development, within the first 2 to 4 h. Both the neutral and alkaline phosphatase Mg-independent activities are induced much later, about the time that myxospores become evident (24 to 30 h). The three activities are greatly diminished upon germination; however, the kinetics of loss differ for all three. The acid phosphatase activity declines very rapidly, the neutral activity begins to decline only after spores begin to convert to rods, and the alkaline phosphatase activity remains high until the time the cells begin to divide. All three developmental activities were measured in the developmental signalling mutants carrying asg, csg, and dsg. The pattern of expression obtained in the mutants was consistent with that of other developmentally regulated genes which exhibit similar patterns of expression during development. The ease with which phosphatases can be assayed should make the activities described in this report useful biochemical markers of stages of both fruiting body formation and germination.
据报道,在刺激黄色粘球菌子实体形成过程中起重要作用的信号之一是磷酸盐饥饿。因此,我们选择研究黄色粘球菌发育过程中的磷酸酶活性。许多磷酸酶可以切割底物对硝基苯磷酸酯。在不同pH值的缓冲液中使用该底物,我们获得了黄色粘球菌发育和萌发过程中的磷酸酶活性图谱。这些实验表明,黄色粘球菌中有五种磷酸酶活性模式:两种营养型和三种发育型。两种独特的营养型活性在pH 7.2和8.5时具有最佳活性。两者都需要镁,并且都被还原剂二硫苏糖醇抑制。发育型(孢子)活性模式的最佳pH值为5.2、7.2和8.5。所有三种活性都不依赖镁。只有碱性磷酸酶活性被二硫苏糖醇抑制。酸性磷酸酶活性在发育早期,即最初的2至4小时内被诱导。中性和碱性磷酸酶的非镁依赖性活性诱导得晚得多,大约在粘孢子明显出现(24至30小时)的时候。这三种活性在萌发时大大降低;然而,三种活性丧失的动力学不同。酸性磷酸酶活性下降非常迅速,中性活性仅在孢子开始转化为杆状后才开始下降,而碱性磷酸酶活性在细胞开始分裂之前一直保持较高水平。在携带asg、csg和dsg的发育信号突变体中测量了所有三种发育活性。在突变体中获得的表达模式与其他发育调控基因的表达模式一致,这些基因在发育过程中表现出相似的表达模式。磷酸酶易于检测的特性应使本报告中描述的活性成为子实体形成和萌发阶段有用的生化标记。