Intellectual Merit: Myxococcus xanthus is a bacterial model for differentiation, intercellular communication, surface motility and bio-complexity. It integrates complex stimuli from its environment to regulate a decision-making process which culminates in formation of stressresistant spores. One critical class of signaling systems is comprised by "two components" (a sensor and a regulator) that directly link environmental stimuli to control of behavior and gene expression. Typically, the sensor and regulator are encoded by genes adjacent to one another on the chromosome and are thus predicted to comprise specific "two-componentsystem" (TCS) to regulate the biology of the cell. M.xanthus encodes at least 144 sensor histidine kinases (HK) and 150 response regulator (RR) proteins, many of which appear to be randomly positioned on the chromosome. As such, M.xanthus possesses one of the most complex and largest repertoires for signal transduction capacity in the bacterial world. Due to significant similarity between these evolutionarily related two-component proteins, prevention of undesired "cross-talk" (aberrant signaling or miscommunication) while also managing bona fide cross-regulation between systems is a critical feature for regulation of M.xanthus community structure. To probe features of TCS that maintain specificity or impart cross-regulation, the proposed study will focus on a subset of highly similar systems in M. xanthus delineated by their presence within gene clusters on the chromosome. Previous work demonstrated complex interactions between two such systems in M.xanthus that affects spore formation. Further dissection of the those pathways led to the finding that the sensor HK functions both as a kinase and as a phosphatase towards its target regulator. The current study will test the hypothesis that a single amino acid residue (in the appropriate context) is exclusively required for phosphatase activity for the majority of sensor kinase proteins in M.xanthus. Because similar sequences exist in nearly all bacterial sensor kinases, the proposed studies will impact understanding of similar systems in nearly all bacteria. The plan is to identify those residues exclusively required for kinase and phosphatase activity in a family of highly related TCS signaling proteins in M.xanthus. Biochemical studies will determine if these systems remain insulated or display cross-regulation. Network analysis of the interacting partners will be assessed using both genetics and biochemistry. A Systems Biology modeling effort will also be utilized to predict control and functionality for identified networks and can be extended to the remainder of the signaling systems in M.xanthus. Broader Impacts. Research: Results from this study will broadly impact the signal transduction field, understanding multicellular interactions in complex environments, interspecies interactions, and facilitate identification of signaling pathway interactions via systems biology in other organisms. Education: The laboratory places a strong emphasis on mentoring postdocs, graduate students, and undergraduates and has a consistent record of placing individuals in strong research environments following their training. Postdocs and students have received fellowships and national recognition for their accomplishments. The PI has participated in multiple NSF-funded educational programs over the past decade and taught in internationally recognized programs.
