Unusual mechanisms of metal regulation down to single-cell single-molecule level

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Enter the text here that is the new abstract information for your application. This section must be no longer than 30 lines of text. Defining how cells regulate the uptake and efflux of transition metals such as Zn and Cu is a key component in elucidating cellular mechanisms of metal homeostasis. Bacterial model systems provide paradigms for understanding metal-responsive gene regulation. In E. coli, the metalloregulator ZntR senses Zn excess and activates Zn efflux, while Zur senses Zn sufficiency and represses Zn uptake, to keep this essential metal at appropriate physiological levels in the cell. CueR, a homolog of ZntR, senses intracellular Cu to activate Cu efflux/detoxification genes to keep this toxic metal minimal. The long-term goal here is to understand how metal regulation in the cell can be manipulated for preventive and therapeutic purposes. Toward this goal, the PI has established an internationally unique research program that applies and develops advanced single- molecule single-cell approaches to interrogate and understand the mechanisms of bacterial metal regulation both in vitro and in live cells, which are further enhanced by bulk biochemical/biophysical and protein/genetic engineering approaches and established collaborations with biologists. The research has led to discoveries of first-of-their-kind mechanisms of Cu/Zn-responsive transcriptional regulation, but new questions also emerged. The objective of this renewal is to continue this program in studying the mechanism of metal regulation in bacteria. The premise of this research comprises the importance of (bacterial) metal regulation in biology, the discovered novel and broadly relevant regulation mechanisms, and the power of combining single- molecule/cell and bulk measurements. The proposed research contains a few specific aims, each with sub- aims, to study the mechanism of Zn-sensing metelloregulator Zur and the Cu-sensing two component system CusRS in E. coli, as well as the cell-cell interactions on the Zu regulations of individual cells. The research is significant because it will elucidate novel molecular mechanisms of metalloregulators in regulating metal efflux and uptake, as well as provide fundamental knowledge about cell biology of metals in general, for identifying causes or developing preventions of diseases that involve similar regulation processes, and for helping the development of (bio)chemical strategies to manipulate bacterial Zn/Cu regulation to impair pathogen growth. The research is innovative because it applies/develops novel single-molecule manipulation, imaging, and analysis methods, and introduce new mechanistic concepts in transcription regulation.