DOI: 10.5061/dryad.vq83bk42p
title: Data from: Dispensability of extrinsic DnaA regulators in Escherichia coli cell-cycle control

description: Investigating a long-standing conceptual question in bacterial physiology
 we examine why DnaA, the bacterial master replication initiator protein,
 exists in both ATP and ADP forms, despite only the ATP form being
 essential for initiation. We engineered the Δ4 Escherichia coli strain,
 devoid of all known external elements facilitating the DnaA-ATP/ADP
 conversion and found that these cells display nearly wild-type behaviors
 under nonoverlapping replication cycles. However, during rapid growth with
 overlapping cycles, Δ4 cells exhibit initiation instability. This aligns
 with our model predictions, suggesting that the intrinsic ATPase activity
 of DnaA alone is sufficient for robust initiation control in E. coli and
 the DnaA-ATP/ADP conversion regulatory elements extend the robustness to
 multifork replication, indicating an evolutionary adaptation. Moreover,
 our experiments revealed constant DnaA concentrations during steady-state
 cell elongation in both wild-type and Δ4 cells. These insights not only
 advance our understanding of bacterial cell-cycle regulation and DnaA but
 also highlight a fundamental divergence from eukaryotic cell-cycle
 controls, emphasizing protein copy-number sensing in bacteria versus
 programmed protein concentration oscillations in eukaryotes.
description: Mother machine experiments under fluorescence
 microscopy. Flow cytometry experiments.
description: # Data from: Dispensability of extrinsic DnaA regulators in *Escherichia
 coli* cell-cycle control
 [https://doi.org/10.5061/dryad.vq83bk42p](https://doi.org/10.5061/dryad.vq83bk42p) This is a collection of raw data from the mother-machine experiments (.zip files) and the flow-cytometry experiments (.xlsx files) in the paper. ## Description of the data and file structure **Mother-machine data:** * Each .zip file contains full images of a mother-machine experiment for the corresponding strain. For example, file "20220331_ALO7918_ABTglc_channels.zip" contains all the images from the mother-machine experiment for ALO7918 in the ABT-glucose condition that was conducted on 3/31/2022. For the strains' genotypes, please check our paper's SI appendix. * All images are in TIFF form. The original images have been preprocessed: first sliced for single channels and then stacked over time. No channel has been filtered out during preprocessing. In the suffix, "c1" means phase contrast images, and "c2" and "c3" are fluorescence images. **Flow-cytometry data:** * File "20220831_WT_Delta2ATP_Delta2ADP_Delta4_Triplicates.xlsx": All the experiments were conducted in the same growth condition -- ABT-glucose. The first sheet contains the raw flow cytometry data, and the second sheet shows the mean values of some parameters based on the flow cytometry experiments. "WT" means wild-type (MG1655), "Δ2" or "DARS" means Δ2-ATP in the paper, and "Δ2' " or "HDADAT" means Δ2-ADP in the paper. Color highlights are only for grouping experiments of the same strain for eye guidance. * File "20221109_WT_Delta4_4media.xlsx": The first sheet contains the raw flow cytometry data. The second sheet plots each column in the first sheet. The third one shows the mean values of some parameters based on the flow cytometry experiments, while the color highlights the values (red for small and green for large) for direct comparison. The fourth sheet shows the doubling times with the color highlighting the growth rates (red for slow growth and green for fast growth) for direct comparison. * File "20231101_WT_Delta4_pLR40mutants_fc.xlsx": The first sheet contains notes for some growth situations and strain information (see more details in SI appendix of the paper). The second sheet contains the raw data from flow cytometry. The third sheet shows plots of each column in the second sheet. The fourth one shows the mean values of some parameters based on the flow cytometry experiments or protein purification experiments, while the color highlights the values (red for small and green for large) for direct comparison. The last sheet is the results from western blots. * File "20240416_bgaldnaap.xlsx": "W_1", "W_2", "W_3" are three replicates for the wildtype. "DeltaADP" means Δ2-ADP in the paper, and "DeltaATP" means Δ2-ATP in the paper.