E. coli leverages growth arrest to remodel its proteome upon entry into starvation

It is widely believed that due to nutrient limitations in natural environments, bacteria spend most of their life in non-growing states. However, very little is known about how bacteria change their phenotype during starvation and what controls the concentration of different gene products inside the cells. Here we used microfluidics with quantitative fluorescence microscopy to quantitatively monitor growth and gene expression in many independent single-cell E. coli lineages as cells were switched from exponential growth to carbon starvation. In contrast to the hypothesis that stationary phase at the population level may reflect a balance between continuing growth and death in different sub-populations, we found that all cells immediately enter growth arrest, that cells further in their cell cycle subsequently undergo reductive division, and no cell death occurs for more than two days. Second, we observed dramatic time-dependent changes in protein production that are highly homogeneous across single cells. Some promoters shut off protein production immediately, some show a slow exponential decay of production on a 10 h time scale, while others exhibit a transient burst of increased production before decaying exponentially at different rates. Notably, the reduction in protein production 30-60 h into starvation relative to production in exponential phase varies by more than two orders of magnitude across promoters and is highly correlated with production in the first 10 h of starvation. Control experiments show that protein degradation itself also decays expo-nentially and using mathematical modeling we show how the fold-change in a genes protein concentration between exponential phase and late starvation depends on the size of the expression burst at the onset of starvation, the rate of subsequent production decay, and the rate of degradation decay. For many genes, the expression in late starvation is driven by production during the first 10 h. Finally, we establish that this expression program at the onset of starvation is critical for cell viability. In particular, by inhibiting gene expression during different periods of starvation, we show that tolerance to stress later in starvation is determined by gene expression occurring during the first 10 h. Our study provides a foundation for quantitative studies of bacterial starvation by uncovering a gene expression program that fundamentally remodels the proteome during the first 10 h of starvation, is highly homogeneous across single cells, sets the proteome later in starvation, and is crucial for stress tolerance.