Single-Cell Biophysics: Measurement, Modulation, and Modeling

Single-Cell Biophysics: Measurement, Modulation, and Modeling

Sunday Speaker Abstracts

Bacterial Transcription Meets Chromosome Organization: A Single-Molecule Perspective Achillefs Kapanidis , Mathew Stracy. University of Oxford, Oxford, United Kingdom. Despite the fundamental importance of transcription, a comprehensive analysis of RNA polymerase behavior and its role in the nucleoid organization in vivo is lacking, in part due to lack of sensitivity of conventional imaging and to its inability to resolve fine cellular structures. To address this challenge, my group is using superresolution fluorescence microscopy to study the localization and dynamics of the transcription machinery and the bacterial chromosome in live bacterial cells, both at the single-molecule and the population level. Specifically, we use photo-activated single-molecule tracking to discriminate between diffusing RNA polymerases and RNA polymerases specifically bound to DNA, either on promoters or transcribed genes. We find that transcription can cause spatial reorganization of the nucleoid, with movement of gene loci out of the bulk of DNA as levels of transcription increase. We also studied the degree and mode of interaction of RNAP with the DNA during on the promoter search process, showing that RNAP interacts substantially with non-specific DNA. Current work focuses on identifying the identity and organisation of genes that are highly transcribed, as well as on developing assays to study the non-specific interactions of DNA-binding proteins with chromosomal DNA. Our work provides a global view of the organization of transcription and its interplay with chromosome organisation in living bacteria.

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