Quorum sensing is a process that enables a group of single-celled bacteria to count their numbers and, based on cell density, behave in a coordinated manner through cell-cell communication. One bacterium that communicates through a quorum-sensing response is the plant pathogen Pantoea stewartii subsp. stewartii. This organism causes Stewart's wilt disease in sweet corn and maize, and thus has a negative impact on production of these important US agricultural commodities. The wilt is a consequence of the extracellular capsule or slime layer produced by the bacteria that blocks the water transport vessels of the plant. Production of this capsular material is controlled by quorum sensing. The quorum-sensing response involves extracellular signaling molecules made by the bacteria that are called acyl homoserine lactones (AHL) and a cellular receptor protein for the AHL called EsaR. EsaR represses production of the extracellular capsule at low cell density, but is inactivated by AHL at high cell density resulting in derepression of capsule synthesis. This mechanism of AHL control is atypical amongst bacterial quorum sensing regulatory proteins and is poorly understood. Most of the over fifty known bacterial quorum-sensing systems function only at high cell density, when the AHL signaling molecule is also present at high concentration. Recently the PI demonstrated that AHL-free EsaR can also activate gene expression at low cell density which is also a unique aspect of quorum sensing in P. stewartii in comparison to other systems. The project here is to test the hypothesis that EsaR acts both as an activator and as a repressor in P. stewartii. The specific aims of the project are to (1) elucidate the mechanism whereby EsaR responds to its ligand AHL in a manner opposite to that of the majority of quorum-sensing regulators and (2) determine the targets and the manner whereby EsaR controls not one, but two signal transduction pathways, one at low cell density and another at high cell density. Results from these studies will transform some accepted paradigms of quorum sensing regulators and the manner in which they coordinate changes in bacterial gene expression. Studies of EsaR and the genes it controls will also lead to a better understanding of the stages of pathogenic infections in plants, and are also likely to reveal possible avenues for disease control in the future.
Broader Impact Research training and mentoring of graduate and undergraduate students are top priorities of this project. Students will be trained in the methods of science as well as in scientific ethics. Students will be nurtured in other aspects of their professional development, such as giving presentations at scientific conferences and learning to work in a synergistic manner with each other and with collaborators. Ultimately, these junior scientists will graduate and move on to become productive members of the scientific community. To bring more diversity to this community, efforts will be made to recruit, mentor and retain women and underrepresented minorities to the research group and to maintain a supportive working environment for them. As an individual, the PI is actively engaged in improving the microbiology/immunology curriculum at Virginia Tech and serving as an academic advisor to individual students as well as to the undergraduate microbiology club. The club performs educational outreach at regional daycares, elementary schools and alumni events, teaching parents and children about the importance of microorganisms in their daily lives.
Quorum-sensing is a mechanism whereby bacterial cells can communicate with one another and coordinate their activities. The quorum-sensing system of the bacterial corn pathogen Pantoea stewartii was studied under this project award. We performed structure/function analysis on the key master protein regulator used during quorum sensing in P. stewartii, EsaR. Understanding how this protein recognizes its quorum-sensing signal may assist in the future development of synthetic compounds that will modulate bacterial quorum-sensing outputs to the benefit of society. We also investigated the genes whose expression is controlled by quorum-sensing in P. stewartii (the EsaR regulon) that contribute to the virulence of this plant pathogen. Better understanding the molecular basis of bacterial plant wilt disease might enable the development of novel mitigation strategies that will protect important agricultural commodities. In total eleven graduate students and thirteen undergraduate students received scientific training. All of the individuals that earned degrees (B.S., M.S. or Ph.D.) to date have pursued careers in some field of the biosciences upon their graduation. The classroom and professional educational activities of the PI as well as the K-5 level outreach activities of the Microbiology Club of Virginia Tech have contributed to a broader exposure of the public to the field of microbiology, which impacts almost every aspect of daily life.
