Turnover of FlhD and FlhC, master regulator proteins for Salmonella flagellum biogenesis, by the ATP‐dependent ClpXP protease
Turnover of FlhD and FlhC, master regulator proteins for Salmonella flagellum biogenesis, by the ATP‐dependent ClpXP protease
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DOI：
10.1046/j.1365-2958.2003.03437.x
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发表时间：
2003-04
影响因子：
3.6
通讯作者：
T. Tomoyasu;A. Takaya;E. Isogai;Tomoko Yamamoto
中科院分区：
文献类型：
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作者：
T. Tomoyasu;A. Takaya;E. Isogai;Tomoko Yamamoto
在肠细菌如沙门氏菌中，鞭毛生物发生依赖于鞭毛基因层级顶端的主操纵子flhDC，其分为三类1、2和3。先前我们报道了ClpXP ATP依赖性蛋白酶的耗尽导致2类和3类基因转录的显著增强，而flhDC操纵子的转录水平在沙门氏菌鼠伤寒血清型中保持正常。这表明ClpXP蛋白酶可能负责FlhD和FlhC主调节子的周转（Tomoyasu，T.，Ohkishi，T.，Ukyo，Y.，Tokumitsu，A.，Takaya，A.，铃木，M。例如，2002，J Bacteriol 184：645-653）。在本研究中，为了确定ClpXP蛋白酶在FlhD和FlhC蛋白周转中的作用，我们使用抗FlhD和抗FlhC抗体分析了野生型和ClpXP突变细胞中FlhD和FlhC蛋白的水平。结果表明，FlhD和FlhC蛋白在ClpXP突变体细胞中显著积累，并且ClpXP突变体中FlhC的半衰期约为5倍，表明ClpXP蛋白酶负责FlhD和FlhC的降解。结果还显示ClpXP蛋白酶降解FlhD 2FlhC 2复合物中的两种蛋白质，但似乎不识别单独合成的相应亚基。综上所述，表明FlhD 2FlhC 2主调节因子的细胞浓度在翻译后水平由ClpXP蛋白酶严格控制。我们还检查了ATP依赖性蛋白酶家族的其他成员在鞭毛生物发生调节中的作用，并得出结论，该家族中只有ClpXP作为沙门氏菌鞭毛生物发生的负调节剂发挥作用。
In enterobacteria such as Salmonella, flagellar biogenesis is dependent upon the master operon flhDC at the apex of the flagellar gene hierarchy, which is divided into three classes 1, 2 and 3. Previously we reported that depletion of the ClpXP ATP‐dependent protease results in dramatic enhancement of class 2 and class 3 gene transcription, whereas the transcription level of the flhDC operon remains normal in Salmonella enterica serovar Typhimurium. This suggests that the ClpXP protease may be responsible for the turnover of the FlhD and FlhC master regulators (Tomoyasu, T., Ohkishi, T., Ukyo, Y., Tokumitsu, A., Takaya, A., Suzuki, M. et al., 2002, J Bacteriol 184:645–653). In this study, to establish the role of the ClpXP protease in the turnover of FlhD and FlhC proteins, we analysed levels of the FlhD and FlhC proteins in wild‐type and ClpXP mutant cells using anti‐FlhD and anti‐FlhC antibodies. The results show that both FlhD and FlhC proteins are markedly accumulated in ClpXP mutant cells and the half‐life of FlhC is approximately fivefold longer in the ClpXP mutant, suggesting that the ClpXP protease is responsible for the degradation of FlhD and FlhC. The results also show that the ClpXP protease degrades both proteins in FlhD2FlhC2 complex but does not seem to recognize the respective subunits synthesized individually. Taken together, it is suggested that the cellular concentration of the FlhD2FlhC2 master regulator is tightly controlled at the post‐translational level by the ClpXP protease. We also examined the role of other members of the ATP‐dependent protease family in the regulation of flagellar biogenesis and concluded that only ClpXP in this family functions as a negative regulator for flagellar biogenesis in Salmonella.