Bacterial infections and surface colonization of implanted medical devices are and will continue to be major health threats especially with the development of multiple drug resistant pathogens. Bacterial pathogens rely on their membrane sensors to detect the presence of host cells in order to produce virulence factors at the right time to invade host cells or colonize the surface of medical devices. Blocking bacterial sensing of hosts could be a new approach to stop bacterial infection. As part of our long term effort to understand bacterial sensing and signaling during host cell invasion, we are studying the molecular mechanism of Sinorhizobium meliloti ExoR protein in controlling the switch from free living cells to invasion ready cells. We have shown previously that both the ExoR protein and the ExoS/ChvI two-component regulatory system regulates inversely the production of a polysaccharide (succinoglycan) required for host cell invasion, and flagella, required for bacterial motility. Our current hypothesis is that the active form of the ExoR protein (ExoRm) binds ExoS and keeps it in the OFF state, altering the expression of the genes regulated by ExoS/ChvI system. The amount of ExoRm in the periplasm is maintained through feedback regulation and the proteolysis of ExoRm to produce ExoRc20. The changes in ExoRm proteolysis rate is being used by the cells to sense environmental stimuli control the switch between free living and attached living. This model is based on the following key discoveries made in the last two years. First, ExoR functions upstream of ExoS/ChvI two-component regulatory system and negatively regulates the activity of ExoS; Second, ExoR autoregulates through feedback regulation; Third, ExoR is digested in periplasm to generate ExoRc20. We plan to 1) determine the function of different forms of ExoR proteins by expressing different forms of ExoR, ExoRp, ExoRm, and ExoRc20, individually; and by examining their biological functions and their ability to interact directly with ExoS protein; 2) determine the molecular mechanisms of ExoR proteolysis and its regulation by characterizing the molecular mechanism of ExoR proteolysis; identifying environmental stimuli modulating ExoR proteolysis, and identifying the S. meliloti ExoR protease. Our new discoveries can be applied immediately to the understanding of the pathogenicities of 56 animal and plant pathogens that have close homologs of S. meliloti ExoR. Our new discoveries will contribute to the development of new approaches to control bacterial infections. PUBLIC HEALTH RELEVANCE: After sensing the presence of their hosts, most pathogenic bacteria produce special products to enhance their abilities to infect host cells. Our study will lead to the development of new approaches to control bacterial infections by blocking their abilities to sense of the presence of their hosts.