Changes in Spo0A~P pulsing frequency control biofilm matrix deactivation.

Under starvation conditions, B. subtilis survives by differentiating into one of two cell types: biofilm matrix-producing cells or sporulating cells. These two cell-differentiation pathways are activated by the same phosphorylated transcription factor - Spo0A~P. Despite sharing the activation mechanism, these cell fates are mutually exclusive at the single-cell level. This decision has been shown to be controlled by the effects of growth rate on gene dosage and protein dilution in the biofilm matrix production network. In this work, we explore an alternative mechanism of growth rate-mediated control of this cell fate decision. Namely, using deterministic and stochastic modeling, we investigate how the growth-rate-dependent pulsing dynamics of Spo0A~P affect biofilm matrix deactivation and activation. Specifically, we show that the Spo0A~P pulsing frequency tunes the biofilm matrix deactivation and activation probability. Interestingly, we found that DNA replication is the cell cycle stage that most substantially contributes to the deactivation of biofilm matrix production. Finally, we report that the deactivation of biofilm matrix production is not primarily regulated by the effects of growth rate on gene dosage and protein dilution. Instead, it is driven by changes in the pulsing period of Spo0A~P. In summary, our findings elucidate another mechanism governing biofilm deactivation during the late stages of starvation, thereby advancing our understanding of how bacterial networks interpret dynamic transcriptional regulatory signals to control stress-response pathways.