Progressive decline in old pole gene expression signal enhances phenotypic heterogeneity in bacteria

Cell growth and gene expression are heterogeneous processes at the single-cell level, leading to the emergence of multiple physiological states within bacterial populations. Cellular aging is a known deterministic driver of growth asymmetry, however its role on gene expression heterogeneity remains elusive. Here we show that aging mother cells undergo a progressive decline in old pole activity, generating asymmetry in product partitioning, gene expression, and cell morphology. We demonstrate that mother cells, when compared to their daughters, exhibit lower product inheritance and gene expression rates independently of promoter dynamics. The declining activity of maternal old poles generates gene expression gradients that manifest as mother-daughter asymmetry upon division, showing that asymmetry is built over time within the maternal intracellular environment. Moreover, old pole aging correlates with a gradual increase in cell length, leading to morphological asymmetry. These findings provide further evidence for aging as a mechanism to enhance phenotypic heterogeneity in bacterial populations, with possible consequences for stress response and survival.

Methods
The dataset contains processed data on single-cell microscopy observations of Escherichia coli MG1655 bacteria growing in the mother machine microfluidic device (Wang et al., 2010). These cells contain a fluorescent transcriptional reporter of RpoS expression (Zaslaver et al., 2006) or constitutive GFP expression, and were imaged in phase contrast (2 min intervals), GFP fluorescence (10 min intervals) and RFP fluorescence (10 min intervals, for cell lysis visualization with propidium iodide). Mother and daughter cells were tracked for 72h (100 generations) in M9 medium at 37°C. Raw images were processed using DeLTA, a pipeline for image segmentation using machine learning algorithms (Lugagne et al., 2020) and the data presented here was extracted from single-cell measurements of length at birth (right after division), length at division, time of birth, generations elapsed, elongation rates, etc. The dataset includes the code necessary to reproduce figures in the manuscript. Data files that include both mother and daughter cell information are organized as a list of two separate data frames, with each row referring to one generation.