Cell-Cell Communication and Gene Regulation by Quorum Sensing in the Symbiotic Nitrogen-Fixing Bacterium Rhizobium leguminosarum

9600766 Gray本研究的目的是表征共生固氮菌根瘤菌的群体感应调节系统。 群体感应是革兰氏阴性菌群体密度依赖性基因激活的保守机制。 在群体感应中，一种自我产生的细胞外信号分子（称为自诱导物）与转录激活蛋白相互作用以激活特定基因的表达。R.麦角菌产生一种独特的自诱导物，它与转录激活因子RhiR一起激活根际表达基因（rhiABC）的操纵子和一个导致细胞生长抑制的未识别位点。 rhiR和rhiABC都是由R的豌豆变种所特有的Sym质粒编码的。豆科植物 生长抑制功能似乎是Sym质粒pRL 1 JI的一种特定分离物所独有的，pRL 1 JI也编码阻断自诱导物正常产生的阻遏物功能。 我们最近发现了两个新的R。肠球菌自身诱导物，其产生不受该pRL 1 JI编码的阻遏物的影响。 新鉴定的R.麦角菌信号通过RhiR激活rhiABC，但不激活生长抑制。 我们还发现了一个独特的自诱导产生的相关物种R。草木樨，其功能目前尚不清楚。 本研究的具体目的是：1）对两个新发现的红曲霉的分子结构进行表征;同时，还研究了大豆根瘤菌产生的自诱导物。苜蓿草; 2）鉴定阻断自诱导物合成的pRL 1 JI编码的基因并阐明其作用机制;和3）研究群体感应在pRL 1 JI的接合转移中的作用。 这项研究解决了根瘤菌细胞间通讯，环境感知和基因调控的基本问题。 由于群体感应调节因子的极端保守性，本研究的结果可能对其他共生或致病细菌系统也有直接的应用。这项工作的结果将被整合到现有的微生物生理学遗传学课程中，并代表了细菌作为多细胞生物体的生物学新课程开发的重要组成部分。作为创新教学职业承诺的一部分，P.I.同时也在开发一项独立的研究活动，作为微生物生理学遗传学实验室课程的一部分。 根瘤菌属的细菌作为固定氮的生物来源具有相当大的生态和经济重要性。这些细菌通常与特定宿主植物（如豌豆和苜蓿）的根部形成复杂的共生关系。本研究旨在探讨一种根瘤菌R.豆科植物 将确定细菌产生的特定基因激活信号分子的分子结构，并研究调节这些不同信号产生的基因。这些信号分子在介导细菌种群间遗传交换中的可能作用也将被确定。这项工作的结果应该增加我们对这些重要细菌的调节生物学的理解，并为细胞外信号对其共生生活方式的影响提供新的见解。 ***

9600766 Gray The purpose of this research is to characterize the quorum sensing regulatory system of the symbiotic nitrogen-fixing bacterium Rhizobium leguminosarum. Quorum sensing is a conserved mechanism of population density-dependent gene activation in Gram-negative bacteria. In quorum sensing, a self-produced extracellular signal molecule called autoinducer interacts with a transcriptional activator protein to activate the expression of specific genes. R. Ieguminosarum produces a unique autoinducer that, together with the transcriptional activator RhiR, activates an operon of rhizosphere-expressed genes (rhiABC) and an unidentified locus that causes an inhibition of cell growth. Both rhiR and rhiABC are encoded by Sym plasmids unique to pea-nodulating biovars of R. Ieguminosarum. The growth-inhibiting function appears to be unique to one specific isolate of the Sym plasmid, pRLlJI, which also encodes a repressor function that blocks normal production of autoinducer. We have recently discovered two additional R. Ieguminosarum autoinducers, the production of which is unaffected by this pRLlJI-encoded repressor. The newly-identified R. Ieguminosarum signals activate rhiABC with RhiR, but do not activate growth inhibition. We have also discovered a unique autoinducer produced by the related species R. meliloti, for which no function is currently known. The specific objectives of this research are: 1) to characterize the molecular structures of the two newly-identified R. Ieguminosarum autoinducers, as well as the autoinducer produced by R. meliloti; 2) to identify the pRLlJI-encoded gene that blocks autoinducer synthesis and elucidate its mechanism of action; and 3) to investigate the role of quorum sensing in the conjugal transfer of pRLlJI. This research addresses fundamental questions of intercellular communication, environmental sensing, and gene regulation in Rhizobium. Due to the extreme conservation of quorum sensing regulators, the results of this research may have direct ap plications to other symbiotic or pathogenic bacterial systems as well. The results of this work will be integrated into an existing course in Microbial Physiology & Genetics, and represent an essential component in the development of a new course on the biology of Bacteria as Multicellular Organisms. As part of a career commitment to innovative instruction, the P.I. is also developing an independent research exercise as part of the laboratory curriculum for Microbial Physiology & Genetics. %%% Bacteria of the genus Rhizobium are of considerable ecological and economic importance as a biological source of fixed nitrogen. These bacteria typically form complex symbiotic associations with the roots of specific host plants such as peas and alfalfa. This research project is an investigation into the molecular aspects of intercellular signaling, cell-cell communication, and gene activation in one species of Rhizobium, R. Ieguminosarum. The molecular structures of specific gene-activating signal molecules produced by the bacteria will be determined, and the genes that regulate production of these different signals will be studied. The possible role of these signal molecules in mediating genetic exchange among populations of bacteria will also be determined. The results of this work should increase our understanding of the regulatory biology of these important bacteria and provide new insights into the effects of extracellular signals on their symbiotic lifestyle. ***