Quorum-Sensing and Growth Control in Rhizobium sp. NGR234

变形杆菌类群中的革兰氏阴性细菌通常使用酰化高丝氨酸内酯(acyl-HSLs)作为群体感应(QS)过程中的分子信号。群体感应细菌释放可扩散的信号分子，随着细胞数量的增加而积累，最终触发适应性反应。Luxi和LuxR家族的调节蛋白通常分别是合成和响应酰基-HSL所必需的。扩散的细胞群体通常会产生恒定的低水平的酰基-HSL，这些细胞会沿着浓度梯度迅速扩散出去。种群密度的增加增加了酰基-HSL的相对浓度，最终促进了信号与LuxR-型蛋白的相互作用，进而控制目标基因的转录。尽管这一基本机制是保守的，但QS调控的背景及其控制下的细胞功能在不同的细菌中是高度不同的。本项目的重点是固氮植物共生菌根瘤菌所使用的Luxi-LuxR型QS系统的调控背景。NGR234的作用机制及其对细胞生长速度的影响。NGR234能促进多种豆科植物根部形成固氮共生根瘤。这种混杂宿主相互作用的分子基础已被广泛研究。许多协调植物相互作用的功能都在536kb的pNGR234a质粒上进行。PNGR234a还携带一大簇与其他细菌的质粒复制(Rep)和结合转移(trb/tra)基因同源的基因。Rep/trb/tra簇包括Luxi-LuxR型调控对Trai和Trar，以及额外的QS调节器Tram。TRAI合成3-氧辛酰基-L-高丝氨酸内酯，TRAR与此酰基HSL相互作用，调节trA/trb和rep操纵子的表达。TRAM通过形成抗激活复合体来抑制TRAR。用分子遗传学的方法研究NGR234的QS机制。控制TRAR表达的调控信号和途径将被调查，以确定促进QS的条件。在类似系统的基础上，寄主植物很可能在这一调控中发挥作用。TRAR调控下受QS调控的pNGR234a基因将被鉴定，并阐明其调控表达的机制。最后，将研究QS对细胞生长速度的控制，QS是几种根瘤菌中常见的QS调控靶标。该研究项目产生的结果将提供关于一种重要的植物共生体的细胞间通信的基本信息，以及这种通信在其高度可塑性的宿主相互作用中的作用。更广泛地说，这些研究将增加对细胞间通信如何直接和间接影响与宿主生物的相互作用的理解，并使群体感应微生物能够平衡它们的生理活动与宿主环境。NGR234在与高等植物的共生关系中利用细胞间的通讯。该研究项目检查了这种细胞间通讯的生化和遗传机制，以及它对这种细菌和寄主植物相互作用的影响。这项工作产生的发现将提供利用这些细菌通信系统、抗击动植物传染病以及促进有益的微生物相互作用所需的基本知识。

Gram-negative bacteria within the Proteobacteria group commonly use acylated homoserine lactones (acyl-HSLs) as molecular signals in the process of quorum sensing (QS). Quorum-sensing bacteria release diffusible signal molecules that accumulate with increasing cell number, eventually triggering adaptive responses. Regulatory proteins of the LuxI and LuxR families are usually required for synthesis of, and response to acyl-HSLs, respectively. Diffuse populations of cells generally produce a constant, low level of acyl-HSLs, and these rapidly diffuse out of cells down their concentration gradient. Elevated population density increases the relative acyl-HSL concentration, eventually fostering interaction of the signals with LuxR-type proteins, which in turn, control the transcription of target genes. Although this basic mechanism is well conserved, the context of QS regulation and the cellular functions under its control are highly variable among different bacteria. This project focuses on the regulatory context of a LuxI-LuxR-type QS system employed by the nitrogen-fixing plant symbiont Rhizobium sp. NGR234, its mechanism of action, and its effect on cellular growth rate. NGR234 incites the formation of nitrogen-fixing, symbiotic nodules on the roots of a wide range of leguminous plants. The molecular basis of this promiscuous host interaction has been extensively studied. Many of the functions that orchestrate the plant interaction are carried on the 536 kb pNGR234a plasmid. The pNGR234a plasmid also carries a large cluster of genes homologous to plasmid replication (rep) and conjugal transfer (trb/tra) genes from other bacteria. The rep/trb/tra cluster includes a LuxI-LuxR-type regulatory pair, TraI and TraR, and the additional QS regulator TraM. TraI synthesizes 3-oxo-octanoyl-L-homoserine lactone and TraR interacts with this acyl HSL to regulate tra/trb and rep operon expression. TraM acts to inhibit TraR through formation of an anti-activation complex. A molecular genetic approach is being employed to study the QS mechanism in NGR234. The regulatory signals and pathways that control traR expression will be investigated to determine the conditions that foster QS. Based on analogous systems, the host plant is likely to play a role in this regulation. The QS-regulated pNGR234a genes under TraR control will be identified and the mechanism by which TraR controls their expression elucidated. Lastly, control of cellular growth rate by QS, a common QS regulatory target among several Rhizobium species, will be examined. Findings generated from the research project will provide fundamental information on cell-to-cell communication in an important plant symbiont, and the role of this communication in its highly plastic host interactions. More generally, these studies will add to the understanding of how cell-to-cell communication directly and indirectly influences interactions with host organisms, and enables quorum-sensing microbes to balance their physiological activity with the host environment.The bacterium Rhizobium sp. NGR234 uses cell-to-cell communication during its symbiotic relationships with higher plants. The research project examines the biochemical and genetic mechanisms underlying this intercellular communication, and its influence on the interaction of this bacterium and host plants. The findings generated from this work will provide fundamental knowledge required to take advantage of these bacterial communication systems, for combating infectious disease in plants and animals, as well as promoting beneficial microbial interactions.