The goal of this work is to understand the molecular mechanisms driving the cell cycle of the bacterium Caulobacter crescentus. Caulobacter controls its lifecycle by using a small number of master transcriptional regulators orchestrating in concert to drive the cell cycle. A phospho-signaling two-component pathway involving the histidine kinase CckA controls these regulatory mechanisms, but a fundamental question remains: how does the cell activate CckA at the appropriate time in the cell cycle? This work focuses on identifying the stimulus that turns on CckA and the pertinent partners mediating this signal. Since small molecule inhibitors will allow temporal dissection of the Caulobacter cell cycle, the experiments involve developing chemical inhibitors of signal transduction components and using these inhibitors to uncover the signals that trigger the cell cycle in space and time. A small molecule inhibitor against CckA will be found using a high-throughput screen in a Caulobacter strain engineered to report CckA inhibition. Signaling partners involved in activating CckA will be determined using a genetic screen in the presence of a CckA inhibitor: these signaling partners are expected to be transcripts that vary with the cell cycle, yet are unaffected by the CckA signaling pathway. Characterization of these leads will determine whether they control CckA localization and activity, a key component of this strategy, since CckA activation correlates with its spatial orientation at a specific point of the cell cycle. This project will reveal the principle signaling partners necessary to activate the cell cycle in Caulobacter, providing new insight into mechanisms that define how and when bacteria decide to divide. These discoveries into the fundamental aspects of bacterial physiology will open the door to potential new targets for the development of antibacterial agents.
Development of novel antibiotics will be essential to combat the emerging drug-resistant strains of bacteria that pose a serious threat to public health. Histidine kinases make ideal targets for the development of new antibiotics, since they play a central - and often essential - role in controlling bacterial physiology. By developing a new way to find histidine kinase inhibitors, this work will facilitate systematic development of histidine kinase inhibitors, paving the way to developing a new class of antibiotics.
