Dekkers L C, Bloemendaal C J, de Weger L A, Wijffelman C A, Spaink H P, Lugtenberg B J
Leiden University, Institute of Molecular Plant Sciences, Clusius Laboratory, The Netherlands.
Mol Plant Microbe Interact. 1998 Jan;11(1):45-56. doi: 10.1094/MPMI.1998.11.1.45.
We describe the characterization of a novel Tn5lacZ colonization mutant of the efficiently colonizing Pseudomonas fluorescens strain WCS365, mutant strain PCL1210, which is at least 300- to 1,000-fold impaired in colonization of the potato root tip after co-inoculation of potato stem cuttings with a 1:1 mixture of mutant and parental cells. Similarly, the mutant is also impaired in colonization of tomato, wheat, and radish, indicating that the gene involved plays a role in the ability of P. fluorescens WCS365 to colonize a wide range of plant species. A 3.1-kb DNA fragment was found to be able to complement the observed mutation. The nucleotide sequence of the region around the Tn5lacZ insertion showed three open reading frames (ORFs). The transcriptional start site was determined. The operon is preceded by an integration host factor (IHF) binding site consensus sequence whereas no clear -10 and -35 sequences are present. The deduced amino acid sequences of the first two genes of the operon, designated as colR and colS, show strong similarity with known members of two-component regulatory systems. ColR has homology with the response regulators of the OmpR-PhoB subclass whereas ColS, the product of the gene in which the mutation resides, shows similarity to the sensor kinase members of these two-component systems. Hydrophobicity plots show that this hypothetical sensor kinase has two transmembrane domains, as is also known for other sensor kinases. The product of the third ORF, Orf222, shows no homology with known proteins. Only part of the orf222 gene is present in the colonization-complementing, 3.1-kb region, and it therefore does not play a role in complementation. No experimental evidence for a role of the ColR/ColS two-component system in the suspected colonization traits chemotaxis and transport of exudate compounds could be obtained. The function of this novel two-component system therefore remains to be elucidated. We conclude that colonization is an active process in which an environmental stimulus, through this two-component system, activates a so far unknown trait that is crucial for colonization.
我们描述了高效定殖的荧光假单胞菌菌株WCS365的一种新型Tn5lacZ定殖突变体的特性，该突变体菌株为PCL1210，在用突变体和亲本细胞按1:1混合共同接种马铃薯茎切段后，其在马铃薯根尖定殖方面受损至少300至1000倍。同样，该突变体在番茄、小麦和萝卜的定殖中也受损，这表明所涉及的基因在荧光假单胞菌WCS365定殖多种植物物种的能力中发挥作用。发现一个3.1 kb的DNA片段能够弥补观察到的突变。Tn5lacZ插入位点周围区域的核苷酸序列显示有三个开放阅读框（ORF）。确定了转录起始位点。该操纵子之前有一个整合宿主因子（IHF）结合位点共有序列，而没有明显的-10和-35序列。该操纵子的前两个基因的推导氨基酸序列，命名为colR和colS，与双组分调节系统的已知成员有很强的相似性。ColR与OmpR-PhoB亚类的应答调节因子有同源性，而突变所在基因的产物ColS与这些双组分系统的传感激酶成员相似。疏水性图谱显示，这种假设的传感激酶有两个跨膜结构域，其他传感激酶也是如此。第三个ORF的产物Orf222与已知蛋白质没有同源性。在定殖互补的3.1 kb区域中仅存在orf222基因的一部分，因此它在互补中不起作用。没有获得关于ColR/ColS双组分系统在疑似定殖特性趋化性和渗出物化合物转运中作用的实验证据。因此，这种新型双组分系统的功能仍有待阐明。我们得出结论，定殖是一个活跃的过程，其中环境刺激通过这个双组分系统激活了一个迄今为止未知的、对定殖至关重要的特性。
