Alteration of DNA supercoiling serves as a trigger of short-term cold shock repressed genes ofE. coli

Abstract

Cold shock adaptability is a key survival skill of gut bacteria of warm-blooded animals. Escherichia coli cold shock responses are controlled by a complex multi-gene, timely-ordered transcriptional program. We investigated its underlying mechanisms. Having identified short-term, cold shock repressed genes, we show that their responsiveness is unrelated to their transcription factors or global regulators, while their single-cell protein numbers’ variability increases after cold shock. We hypothesized that some cold shock repressed genes could be triggered by high propensity for transcription locking due to changes in DNA supercoiling (likely due to DNA relaxation caused by an overall reduction in negative supercoiling). Concomitantly, we found that nearly half of cold shock repressed genes are also highly responsive to gyrase inhibition (albeit most genes responsive to gyrase inhibition are not cold shock responsive). Further, their response strengths to cold shock and gyrase inhibition correlate. Meanwhile, under cold shock, nucleoid density increases, and gyrases and nucleoid become more colocalized. Moreover, the cellular energy decreases, which may hinder positive supercoils resolution. Overall, we conclude that sensitivity to diminished negative supercoiling is a core feature of E. coli’s short-term, cold shock transcriptional program, and could be used to regulate the temperature sensitivity of synthetic circuits.

Dynamics of bacterial operons during genome-wide stresses

“…E. coli has 833 operons (MG1655 strain 6,37,41 ), which account for 2708 of the 4724 genes. The nucleotide length of operons and the genes composing them are shown in Supplementary Figures S1A and S1B.…”

“…Operons are regulated by a total of 1174 recognized TSS's. However, they only have a total of 174 recognized transcription termination sites (TTSs) 6,37,42 . To investigate this, we studied the distance (in number of nucleotides) between operons and the next gene in the DNA, downstream from the end of the operon.…”

“…Here, we studied how the behavior of genes in operons in the context of genome wide perturbations that, by targeting RNAP and gyrase 36,37 , affect both primary as well as internal promoters of most operons, quasi-synchronously. We analyzed this dynamical data while accounting for information on the operon's composition of E. coli, which is likely the most dissected bacterial genome.…”

“…To study this, we investigated the genome-wide transcription dynamics of the operons of E. coli (Figure 1A) during stresses targeting RNAP and gyrase (Figure 1B). Disturbing their activity is known to influence (quasi-synchronously) several hundreds of transcription start sites (TSSs) 36,37 . We analyzed the response strength of each gene, in each operon, as a function of its position (Figure 1C) and as a function of the positioning of internal promoters.…”

The transcriptional response to low temperature is weakly conserved across theEnterobacteriaceae

“…A great deal is known about how E. coli responds to low temperature (1,3). A number of global regulatory factors have been implicated, including cold shock proteins (4,5), RNA degradation enzymes like RNase R and PNPase (5,6), nucleoid associated proteins like H-NS (7), global supercoiling levels (8), and the alternative sigma factor RpoS (2,9). Shifts between optimal temperature (37°C) and a lower temperature have substantial impacts on the transcriptome.…”