Genome‐scale quantitative characterization of bacterial protein localization dynamics throughout the cell cycle

Abstract

Bacterial cells display both spatial and temporal organization, and this complex structure is known to play a central role in cellular function. Although nearly one-fifth of all proteins in Escherichia coli localize to specific subcellular locations, fundamental questions remain about how cellular-scale structure is encoded at the level of molecular-scale interactions. One significant limitation to our understanding is that the localization behavior of only a small subset of proteins has been characterized in detail. As an essential step toward a global model of protein localization in bacteria, we capture and quantitatively analyze spatial and temporal protein localization patterns throughout the cell cycle for nearly every protein in E. coli that exhibits nondiffuse localization. This genome-scale analysis reveals significant complexity in patterning, notably in the behavior of DNA-binding proteins. Complete cell-cycle imaging also facilitates analysis of protein partitioning to daughter cells at division, revealing a broad and robust assortment of asymmetric partitioning behaviors.

SuperSegger: robust image segmentation, analysis and lineage tracking of bacterial cells

“…SuperSegger is a powerful automated image processing and analysis package, well suited for high‐throughput time‐lapse fluorescence microscopy of cells (LeRoux et al ., ; O'Connor et al ., ; Kuwada et al ., ; Russell et al ., ; Garmendia‐Torres et al ., ; Javer et al ., ; Kuwada et al ., ; Lampo et al ., ; LeRoux et al ., ; Stylianidou et al ., ; Cass et al ., ). It provides reliable and flexible segmentation which can be trained to optimize performance in a wide variety of experimental contexts.…”

Probing bacterial cell biology using image cytometry

“…To demonstrate the analysis potential of the Clist and gateTool in the context of a high‐throughput time‐lapse analysis, we analyzed data from a near‐complete library of fluorescent fusions. We had previously imaged the ASKA library (Kitagawa et al ., ), a genome‐scale, plasmid‐based collection of inducible fluorescence fusions to genes of E. coli (Kuwada et al ., ) to characterize protein localization dynamics in a model bacterium. An interesting potential feature of the ASKA library is the ability to characterize the effect of protein expression on cell morphology, where protein expression is quantified via fluorescence.…”

“…For previous studies, our lab imaged all ASKA fusions displaying non‐diffuse localization (Kuwada et al ., 2015a,b). We reanalyzed the data from these high‐throughput imaging experiments, which included both the FtsZ and FtsA fusions.…”

Escherichia coli Chromosomal Loci Segregate from Midcell with Universal Dynamics

“…These images show a clear asymmetry in DNA positioning, with a strong bias toward the single new pole at the beginning of the cell cycle and toward the cell center (which will become the two new poles) near the end of the cell cycle. To get an average nucleoid shape distribution throughout the cell cycle, we build a consensus localization pattern, as described in (49).…”