Thewes Sascha, Schubert Sebastian K, Park Kyuhyeon, Mutzel Rupert
Department of Biology, Chemistry, Pharmacy, Institute for Biology - Microbiology, Freie Universität Berlin, Berlin, Germany.
J Basic Microbiol. 2014 Jun;54(6):607-13. doi: 10.1002/jobm.201200574. Epub 2013 May 20.
Calcium signaling is one of the most important signaling-pathways in all eukaryotes. One important target activated by an increased intracellular calcium concentration via calmodulin is the protein phosphatase calcineurin, which is composed of a catalytic subunit (calcineurin A) and a regulatory subunit (calcineurin B). The importance of calcium and calcineurin for the differentiation and development of the social amoeba Dictyostelium discoideum has already been shown by pharmacological approaches. However, so far only a RNAi-silenced calcineurin B mutant has been investigated on a molecular level. Here, we describe the construction and phenotypic investigation of a RNAi-silenced calcineurin A mutant. Phenotypic aberrations during development resemble those produced by silencing of calcineurin B with ectopic tip formation of the fruiting bodies. Additionally, we tested the response of the mutants under various stress conditions in liquid culture as well as during development. Both, calcineurin A and B RNAi-mutants, are hypersensitive during development towards cation stress. Besides its role in development, calcineurin is thus also involved in the stress response in D. discoideum. Further, our data imply that many functions of calcineurin are conserved among the eukaryotes.
钙信号传导是所有真核生物中最重要的信号通路之一。通过钙调蛋白使细胞内钙浓度升高而激活的一个重要靶点是蛋白磷酸酶钙调神经磷酸酶,它由一个催化亚基(钙调神经磷酸酶A)和一个调节亚基(钙调神经磷酸酶B)组成。药理学方法已表明钙和钙调神经磷酸酶对群居变形虫盘基网柄菌的分化和发育具有重要性。然而,到目前为止,在分子水平上仅对RNA干扰沉默的钙调神经磷酸酶B突变体进行了研究。在此,我们描述了RNA干扰沉默的钙调神经磷酸酶A突变体的构建及表型研究。发育过程中的表型异常类似于通过使钙调神经磷酸酶B沉默并使子实体异位形成顶端而产生的异常。此外,我们测试了突变体在液体培养的各种应激条件下以及发育过程中的反应。钙调神经磷酸酶A和B的RNA干扰突变体在发育过程中对阳离子应激均高度敏感。因此,除了其在发育中的作用外,钙调神经磷酸酶还参与盘基网柄菌的应激反应。此外,我们的数据表明钙调神经磷酸酶的许多功能在真核生物中是保守的。