Characterization of a Gene Family of Outer Membrane Proteins (ropB) in Rhizobium leguminosarum bv. viciae VF39SM and the Role of the Sensor Kinase ChvG in Their Regulation
Characterization of a Gene Family of Outer Membrane Proteins (ropB) in Rhizobium leguminosarum bv. viciae VF39SM and the Role of the Sensor Kinase ChvG in Their Regulation
复制标题
DOI：
10.1128/jb.01140-09
复制
发表时间：
2010-02-01
影响因子：
3.2
通讯作者：
Yost, Christopher K.
中科院分区：
文献类型：
--
作者：
Foreman, Dallas L.;Vanderlinde, Elizabeth M.;Yost, Christopher K.
革兰氏阴性菌的外膜代表细菌与其外部环境之间的界面。它作为有害物质的保护屏障具有关键作用，并且在宿主-细菌相互作用中也很重要，代表宿主细胞和细菌细胞之间的初始物理接触点。RopB是一种来自豆科根瘤菌Rhizobium leguminosarum bv. viciae，存在于自由生活的细胞中，但在拟杆菌中不存在（H.罗斯特岛H.穆德斯角A. Wijffelman和B. J. Lugtenberg，Mol.植物微生物相互作用8：576-583，1995）。RopB的功能和RopB基因调控的分子机制仍不清楚。我们鉴定并克隆了来自R.豆科牧草VF 39 SM基因组。报告基因融合表明，ropB的表达是8倍高，当细胞生长在复杂的媒体比当他们生长在基本培养基中，而ropB 3的表达组成型表达在复杂和基本培养基中的低水平。在所有测试条件下，ropB 2的表达可忽略不计。使用补充有各种来源的肽的基本培养基导致ropB表达增加5倍。在chvG突变体背景中没有观察到在肽存在下ropB表达的增加，表明传感器激酶在调节ropB表达中的作用。ropB基因家族的每个成员使用插入诱变进行突变，并测定突变体对抗菌剂和共生表型的敏感性。所有突变体在豌豆植株上形成有效的根瘤，并且类杆菌中每个rop基因的基因表达可以忽略不计。ropB 2和ropB 3的功能仍然是神秘的，而ropB突变体对洗涤剂，疏水性抗生素和弱有机酸的敏感性增加，这表明RopB在外膜稳定性中的作用。
The outer membrane of Gram-negative bacteria represents the interface between the bacterium and its external environment. It has a critical role as a protective barrier against harmful substances and is also important in host-bacteria interactions representing the initial physical point of contact between the host cell and bacterial cell. RopB is a previously identified outer membrane protein from Rhizobium leguminosarum bv. viciae that is present in free-living cells but absent in bacteroids (H. P. Roest, I. H. Mulders, C. A. Wijffelman, and B. J. Lugtenberg, Mol. Plant Microbe Interact. 8: 576-583, 1995). The functions of RopB and the molecular mechanisms of ropB gene regulation have remained unknown. We identified and cloned ropB and two homologs (ropB2 and ropB3) from the R. leguminosarum VF39SM genome. Reporter gene fusions indicated that the expression of ropB was 8-fold higher when cells were grown in complex media than when they were grown in minimal media, while ropB3 expression was constitutively expressed at low levels in both complex and minimal media. Expression of ropB2 was negligible under all conditions tested. The use of minimal media supplemented with various sources of peptides resulted in a 5-fold increase in ropB expression. An increase in ropB expression in the presence of peptides was not observed in a chvG mutant background, indicating a role for the sensor kinase in regulating ropB expression. Each member of the ropB gene family was mutated using insertional mutagenesis, and the mutants were assayed for susceptibility to antimicrobial agents and symbiotic phenotypes. All mutants formed effective nodules on pea plants, and gene expression for each rop gene in bacteroids was negligible. The functions of ropB2 and ropB3 remain cryptic, while the ropB mutant had an increased sensitivity to detergents, hydrophobic antibiotics, and weak organic acids, suggesting a role for RopB in outer membrane stability.