# A delivered DNase toxin creates population heterogeneity through transient intoxication of siblings

## TL;DR

Bacteria use toxins to create population diversity by temporarily poisoning sibling cells, leading to varied responses and potential survival advantages.

## Contribution

The study reveals a molecular mechanism by which bacterial toxin delivery generates population heterogeneity through transient intoxication of kin cells.

## Key findings

- Toxin delivery causes heterogeneous gene expression in isogenic E. coli populations.
- Intoxicated cells can recover through de novo immunity protein synthesis.
- Toxin-induced heterogeneity affects redox status and DNA damage responses.

## Abstract

Population heterogeneity is important for multicellular behavior, as well as bet-hedging strategies. Recent findings suggest a role for bacterial toxin delivery in generating population heterogeneity, but the molecular mechanisms by which this occurs are not well understood. Here, we address if and how delivery of bacterial CdiA toxins generates heterogeneity in an isogenic population of Escherichia coli (E. coli) cells. Using a DNase toxin as a proxy, we find that E. coli populations able to deliver the toxin show a heterogeneous expression of the SOS-response gene sulA, whereas those incapable of kin-delivery remain homogeneous. Heterogeneity results from excessive delivery of toxin into some cells, which become intoxicated due to insufficient immunity. A low level of intoxication by this toxin is transiently reversible, and intoxicated cells can be rescued by the de novo synthesis of cognate immunity protein. The fraction of cells experiencing toxicity is increased by liberating the receptor responsible for toxin import from its tasks in outer-membrane biogenesis, suggesting that kin-intoxication is limited by receptor availability. Expression of sulA is regulated by both DNA damage and redox status. Interestingly, kin-delivery changes redox status, whereas intoxicated non-kin cells induce the SOS DNA damage response. The former results in changed expression of metabolic genes, whereas the latter induces prophage excision, which may promote horizontal gene transfer. In conclusion, we identify a molecular mechanism by which heterogeneity is generated through toxin delivery among kin, and some of the consequences of said heterogeneity.

Population heterogeneity is important for multicellularity, as well as for bet-hedging strategies. A heterogeneous population allows cells with the same genotype to respond differently to environmental cues and stresses. For multicellularity, heterogeneity originates from coordinated signaling, whereas bet-hedging strategies can arise stochastically due to cell-to-cell variation in the concentration of signaling molecules. However, recent advances suggest a role for bacterial toxin delivery in the generation of population heterogeneity. How toxins mediate heterogeneity mechanistically is, however, unclear. Here, we show that kin cells transiently intoxicate each other with CdiA toxins, resulting in physiological changes. These changes are specific to the toxic activity, i.e., other toxins with different activities are likely to give rise to other responses. Thus, we find that the arsenal of toxins that bacteria harbor could affect their ability to participate in bet-hedging strategies, as well as in multicellular behavior.