Simultaneous measurements of translation rate and transcriptome uncovers linked regulation within an active bacterial cell population.

Cell-to-cell variation within clonal bacterial populations provides bacterial communities important advantages including opportunities for bet-hedging and metabolic division of labor. In recent years the extent of bacterial heterogeneity has been documented both at the transcriptome level and with physiological measurements of cell growth rate and translation rate. However, methods that link physiological parameters to a single cell's full transcriptomic state are lacking, making it difficult to identify the regulatory mechanisms that couple physiology and transcriptional output. Here we introduce a method that combines click-chemistry enabled labeling of nascent polypeptides to measure translation rates in single cells alongside microfluidic encapsulation and single cell transcriptomic measurements, providing a tandem measurement of translation rate and transcriptome in thousands of single Bacillus subtilis cells. In a culture experiencing nutrient limitation, we identified a subpopulation of cells with a higher rate of protein translation that uniquely overexpresses genes for several metabolic processes including acetoin production and arginine synthesis. Using a genetic approach informed by the gene expression in this subpopulation, we identified a regulatory mechanism that couples the increase in protein abundance of a transcriptional regulator AlsR with expression of alsR regulated genes in this subpopulation. Overall design: Before microfluidic encapsulation, bacteria were either treated or not treated with OPP as per instructions in the click-it OPP kit (thermofisher scientific) and fixed in 1% paraformaldehyde after OPP incubation in order to preserve transcripts and permeabilized by mild lysozyme treatment. OPP positive samples were processed with click-ligation of a probe using copper click reagents in the Click-it OPP kit. Permeabilized bacteria were then incubated with the corresponding DNA probe library. Non-hybridized probes were removed with repeated washes . Next, the bacteria were run through a 10X Chromium Controller, where the DNA probes were captured and barcoded in a manner analogous to the barcoding of the transcriptome of eukaryotic cells