Spatiotemporal biology of E. coli

The cytoplasm of E. coli exhibits a remarkable degree of spatial and temporal organization in spite of the absence of organelles. Single-cell based widefield fluorescence microscopy methods have enabled direct study of organization and motion of bacterial components involved in translation and transcription. However, fluorescence has long been hampered by the diffraction limit of resolution (~250nm). While cryo-EM and especially the new cryo-tomography techniques provide extraordinarily detailed views of ribosomes and proteins within cells, they are limited to fixed cells and cannot distinguish different proteins from each other.
 

 

 

Our central goal is to use the new sub-diffraction-limit techniques of fluorescence microscopy to study the spatial organization and the motion of key components of the transcription and translation machinery in live E. coli cells in two and three dimensions with high spatial resolution. By localizing and tracking single molecules of photo-activatable proteins that label components of the transcription/translation machinery, we correlate motion of individual components with their location inside E. coli. This will provide much more detailed and quantitative picture of the organization of transcription/translation machinery, communication between them, and their association to the bacterial nucleoid and cytoplasmic membrane.