Phosphate starvation-inducible proteins of Bacillus subtilis:: Proteomics and transcriptional analysis
Phosphate starvation-inducible proteins of Bacillus subtilis:: Proteomics and transcriptional analysis
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DOI：
10.1128/jb.182.16.4478-4490.2000
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发表时间：
2000-08-01
影响因子：
3.2
通讯作者：
Hecker, M
中科院分区：
文献类型：
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作者：
Antelmann, H;Scharf, C;Hecker, M
使用磷酸盐饥饿细胞的细胞提取物和上清液的二维（2D）聚丙烯酰胺凝胶电泳分析了枯草杆菌的磷酸盐饥饿反应。大多数磷酸盐饥饿诱导的蛋白质是在控制下的西格玛（B），其活性增加的能量消耗。为了确定属于Pho调节子的蛋白质，其由双组分调节蛋白PhoP和PhoR调节，将野生型的2D蛋白质模式与sigB突变体和phoR突变体的2D蛋白质模式进行比较。基质辅助激光解吸电离飞行时间质谱，两个碱性磷酸酶（APases）（PhoA和PhoB），APase碱性磷酸二酯酶（PhoD），甘油磷酸二酯磷酸二酯酶（GlpQ），和脂蛋白YdhF被确定为非常强烈的诱导PhoPR依赖性蛋白分泌到细胞外介质。在细胞质部分，PstB 1，PstB 2和TuaD被确定为已知的PhoPR依赖性蛋白，除了PhoB，PhoD，和先前描述的PstS。glpQ和ydhF的转录研究证实了强烈的PhoPR依赖性。北方杂交和引物延伸实验表明glpQ从sigma（A）启动子单顺反子转录，该启动子被四个推定的TT（A/T）ACA样PhoP结合位点重叠。此外，ydhF可能与phoB从phoB启动子开始共转录。在phoR和sigB突变体中，只有一小部分蛋白质保持磷酸饥饿诱导，并且没有形成独特的调节组。其中，YfhM和YjbC由sigma（B）依赖性和未知的PhoPR独立机制控制。此外，YtxH和YvyD似乎在磷酸盐饥饿后在野生型中以sigma（B）依赖性方式被诱导，而在sigB突变体中可能通过sigma（H）被诱导。YxiE由磷酸盐饥饿诱导，不依赖于sigma（B）和PhoPR。
The phosphate starvation response in Bacillus subtilis was analyzed using two-dimensional (2D) polyacrylamide gel electrophoresis of cell extracts and supernatants from phosphate-starved cells. Most of the phosphate starvation-induced proteins are under the control of sigma(B), the activity of which is increased by energy depletion. In order to define the proteins belonging to the Pho regulon, which is regulated by the two-component regulatory proteins PhoP and PhoR, the 2D protein pattern of the wild type was compared with those of a sigB mutant and a phoR mutant. By matrix-assisted laser desorption ionization-time of flight mass spectrometry, two alkaline phosphatases (APases) (PhoA and PhoB), an APase-alkaline phosphodiesterase (PhoD), a glycerophosphoryl diester phosphodiesterase (GlpQ), and the lipoprotein YdhF were identified as very strongly induced PhoPR-dependent proteins secreted into the extracellular medium. In the cytoplasmic fraction, PstB1, PstB2, and TuaD were identified as already known PhoPR-dependent proteins, in addition to PhoB, PhoD, and the previously described PstS. Transcriptional studies of glpQ and ydhF confirmed the strong PhoPR dependence. Northern hybridization and primer extension experiments showed that glpQ is transcribed monocistronically from a sigma(A) promoter which is overlapped by four putative TT(A/T)ACA-like PhoP binding sites. Furthermore, ydhF might be cotranscribed with phoB initiating from the phoB promoter. Only a small group of proteins remained phosphate starvation inducible in both phoR and sigB mutant and did not form a unique regulation group. Among these, YfhM and YjbC were controlled by sigma(B)-dependent and unknown PhoPR-independent mechanisms. Furthermore, YtxH and YvyD seemed to be induced after phosphate starvation in the wild type in a sigma(B)-dependent manner and in the sigB mutant probably via sigma(H). YxiE was induced by phosphate starvation independently of sigma(B) and PhoPR.