In situ single-molecule dynamics of the SOS-repressor LexA during antibiotic stress

Datum

15.10.2020

Zeit

11:00 - 12:00

Sprecher

Michael Schlierf

Zugehörigkeit

Technische Universität Dresden, Center for Molecular and Cellular Bioengineering (CMCB) B CUBE, Germany

Serie

MPI-CBG Thursday Seminar

Sprache

en

Hauptthema

Biologie

Host

Stefan Diez

Beschreibung

In bacteria, the key mechanism facilitating survival, mutation and adaptation upon DNA damage is the SOS response. Through autoproteolytic digestion triggered by single-stranded DNA, the repressor LexA controls over 50 SOS genes. Efforts to inhibit this response and thereby combat antibiotic resistance rely on a broad understanding of its behavior in vivo, which is still limited. Here, we use single-molecule localization microscopy to directly visualize LexA in Escherichia coli. With single-particle tracking PALM, we observe the repressor under antibiotic stress of different intensity and identify four diffusive populations. Using a set of LexA mutants, we assign operator bound, target-searching, freely diffusing and cleaved repressors. Measuring their time-evolution and applying a kinetic model, we obtain in vivo cleavage and binding rates in dependence of antibiotic dosage. A strong SOS response is even observed at low antibiotic stress. We further develop a strategy to count molecules in fixed cells and found that LexA is much more abundant during the late SOS response than previously thought.

Letztmalig verändert: 16.10.2020, 00:09:58