The Pseudomonas aeruginosa global regulator MvaT specifically binds to the ptxS upstream region and enhances ptxS expression
The Pseudomonas aeruginosa global regulator MvaT specifically binds to the ptxS upstream region and enhances ptxS expression
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DOI：
10.1099/mic.0.27270-0
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发表时间：
2004-11-01
期刊：
影响因子：
2.8
通讯作者：
Hamood, AN
中科院分区：
文献类型：
--
作者：
Westfall, LW;Luna, AM;Hamood, AN
铜绿假单胞菌外毒素A的产生受几个基因的正调控或负调控。两个这样的调控基因，ptxR和ptxS，它们是从彼此分化转录，先前已经描述。虽然计算机分析表明ptxR-ptxS基因间区域含有几种调节蛋白的潜在结合位点，但调节铜绿假单胞菌中ptxR或ptxS表达的机制尚不清楚。确定了特异性结合该区域内的区段的铜绿假单胞菌蛋白复合物的存在。在该研究中，结合区域定位于基因间区域的150 Iop片段，并且构成结合复合物的蛋白质被表征为铜绿假单胞菌HU和MvaT。重组MvaT被纯化为融合蛋白（MAL-MvaT），并显示特异性结合到ptxR-ptxS基因间区。构建并鉴定了mvaT缺陷的PAO 1同基因突变体PAODeltamvaT。PAODeltamvaT的裂解物不能与150 bp探针结合。使用实时PCR实验检测mvaT对ptxS和ptXR表达的影响。PTxS在PAODeltamvaT中的表达低于PAO 1，而PTxR在PAODeltamvaT中的表达无差异。这些结果表明，MvaT正调控ptxS的表达特异性结合到ptxS的上游区域。
Exotoxin A production in Pseudomonas aeruginosa is regulated positively or negatively by several genes. Two such regulatory genes, ptxR and ptxS, which are divergently transcribed from each other, have been described previously. While computer analysis suggested that the ptxR-ptxS intergenic region contains potential binding sites for several regulatory proteins, the mechanism that regulates the expression of either ptxR or ptxS in P. aeruginosa is not known. The presence of a P. aeruginosa protein complex that specifically binds to a segment within this region was determined, In this study the binding region was localized to a 150 lop fragment of the intergenic region and the proteins that constitute the binding complex were characterized as P. aeruginosa HU and MvaT. Recombinant MvaT was purified as a fusion protein (MAL-MvaT) and shown to specifically bind to the ptxR-ptxS intergenic region. A PAO1 isogenic mutant defective in mvaT, PAODeltamvaT, was constructed and characterized. The lysate of PAODeltamvaT failed to bind to the 150 bp probe. The effect of mvaT on ptxS and ptXR expression was examined using real-time PCR experiments. The expression of ptxS was lower in PAODeltamvaT than in PAO1, but no difference was detected in ptxR expression. These results suggest that MvaT positively regulates ptxS expression by binding specifically to the ptxS upstream region.