Biology subjects

Gervais, T.

Publications and source records attributed to Gervais, T..

3 recordsLinked to original sources

bioRxiv2024

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.

microbiology↗

bioRxiv2022

Pixelated microfluidics for drug screening on tumour spheroids and ex vivo microdissected primary tissue

Anti-cancer drugs have the lowest success rate of approval in drug development programs. Thus, preclinical assays that closely predict the clinical responses to drugs are of utmost importance in both clinical oncology and pharmaceutical research. 3D tumour models preserve the tumoural architecture and are cost-, labour-, and time-efficient. However, the short-term longevity, limited throughput, and limitations to live imaging of these models have so far driven researchers towards simpler, less realistic tumour models such as monolayer cell cultures. Here, we present a static open-space microfluidic drug screening platform that enables the formation, culture, and multiplexed delivery of several reagents to various 3D tumour models, namely cancer cell line spheroids and ex vivo primary tumour fragments. Our platform utilizes an open-space microfluidic technology, a pixelated chemical display, which creates fluidic "pixels" of biochemical reagents that stream over tumour models in a contact-free fashion. Up to 9 different treatment conditions can be tested over 144 samples in a single experiment. We provide a proof-of-concept application by staining fixed and live tumour models with multiple cellular dyes. Furthermore, we demonstrate that the various responses of the tumour models to biological stimuli can be assessed using the proposed drug screening platform. The platform is amenable to various 3D tumour models, such as tumour organoids. Upscaling of the microfluidic platform to larger areas can lead to higher throughputs, and thus will have a significant impact on developing treatments for cancer.

bioengineering↗

bioRxiv2022

Growth rate controls the sensitivity of gene regulatory circuits

Unicellular organisms adapt to their changing environments by gene regulatory switches that sense chemical cues and induce specific target genes when the inducing signal is over a critical threshold. Using mathematical modeling we here show that, because growth rate sets the dilution rate of intracellular molecules, the sensitivity of gene regulatory switches automatically couples to growth rate, in a way that can be exploited by natural selection. We confirm the modeling predictions by experimentally demonstrating that, as nutrient quality is varied, the concentration of inducer required for activating the lac operon in E. coli increases quadratically with the populations growth rate. Our theory further predicts that, when growth rate is instead modulated by translation inhibition, critical inducer levels are invariant, and we experimentally validate this prediction as well. Moreover, we establish that this growth-coupled sensitivity allows bacteria to implement concentration-dependent sugar preferences, in which a new carbon source is used only if its concentration is high enough to improve upon the current growth rate of the cells. Using microfluidics in combination with time-lapse microscopy, we validate experimentally that this strategy governs how mixtures of glucose and lactose are used in E. coli at single-cell level. Overall, growth-coupled sensitivity provides a general mechanism through which cells can mute external signals in beneficial conditions when growth is fast, and become highly sensitive to alternative nutrients or stresses when growth is slow or arrested.

systems biology↗