Two-component response regulators of Vibrio fischeri:: Identification, mutagenesis, and characterization
Two-component response regulators of Vibrio fischeri:: Identification, mutagenesis, and characterization
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DOI：
10.1128/jb.00242-07
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发表时间：
2007-08-01
影响因子：
3.2
通讯作者：
Visickl, Karen L.
中科院分区：
文献类型：
--
作者：
Hussa, Elizabeth A.;O'Shea, Therese M.;Visickl, Karen L.
原核和真核细胞利用双组分信号转导系统来感知和响应环境刺激，以维持体内平衡并快速适应不断变化的条件。已经开始出现利用大规模诱变方法来分析原核生物中这些系统的研究。由于最近获得了其基因组序列，对于海洋生物发光细菌费氏弧菌来说，这种全球性的方法现在是可能的，这种细菌要么以自由生活的状态存在，要么以共生体的形式存在于宿主生物体中，例如夏威夷鱿鱼种Euprymna scolopes。在这项工作中，我们鉴定了费氏弧菌基因组中编码的 40 个假定的双组分反应调节因子。根据存在的效应结构域的类型，我们将 6 个分类为 NarL 型，13 个为 OmpR 型，6 个为 NtrC 型；其余 15 个缺乏预测的 DNA 结合域。随后，我们通过载体整合方法突变了其中 35 个基因，并分析了所得突变体在鱿鱼的生物发光、运动性和竞争性定殖中的作用。通过这些测定，我们鉴定了 V fischeri 发光的三个新调节因子和七个改变运动性的调节因子。此外，我们发现 11 个调节因子对宿主鱿鱼的竞争性定殖具有先前未描述的影响。有趣的是，五个新表征的调节因子各自影响两个或多个所检查的表型，强烈表明系统之间存在互连性。这项工作代表了首次使用基因组方法对费氏弧菌中的一类基因进行大规模诱变，并强调了双组分信号转导在细菌-宿主相互作用中的重要性。
Two-component signal transduction systems are utilized by prokaryotic and eukaryotic cells to sense and respond to environmental stimuli, both to maintain homeostasis and to rapidly adapt to changing conditions. Studies have begun to emerge that utilize a large-scale mutagenesis approach to analyzing these systems in prokaryotic organisms. Due to the recent availability of its genome sequence, such a global approach is now possible for the marine bioluminescent bacterium Vibrio fischeri, which exists either in a free-living state or as a mutualistic symbiont within a host organism such as the Hawaiian squid species Euprymna scolopes. In this work, we identified 40 putative two-component response regulators encoded within the V. fischeri genome. Based on the type of effector domain present, we classified six as NarL type, 13 as OmpR type, and six as NtrC type; the remaining 15 lacked a predicted DNA-binding domain. We subsequently mutated 35 of these genes via a vector integration approach and analyzed the resulting mutants for roles in bioluminescence, motility, and competitive colonization of squid. Through these assays, we identified three novel regulators of V fischeri luminescence and seven regulators that altered motility. Furthermore, we found 11 regulators with a previously undescribed effect on competitive colonization of the host squid. Interestingly, five of the newly characterized regulators each affected two or more of the phenotypes examined, strongly suggesting interconnectivity among systems. This work represents the first large-scale mutagenesis of a class of genes in V fischeri using a genomic approach and emphasizes the importance of two-component signal transduction in bacterium-host interactions.