A Dipolar Photoswitch Modulates Bacterial Membrane Potential and Reveals Context-dependent Bioelectrical Circuitry

Dynamic bioelectric signalling in bacteria regulates physiology and collective behaviours, yet tools to perturb microbial membrane voltage with high spatiotemporal control remain limited. Here we introduce MTP2, a non-genetic, membrane-targeting azobenzene photoswitch that enables optical modulation of bacterial membrane potential by tuning interfacial electrostatics in Bacillus subtilis. MTP2 associates strongly with the cell envelope and shifts the resting potential to more negative values in the dark, while 470-nm illumination evokes a robust, reversible depolarization at the single-cell level. Although MTP2 photoisomerization is ultrafast (picoseconds), the voltage waveforms unfold over seconds to minutes, indicating that the response is set by homeostatic ion transport rather than by MTP2 photochemistry. Genetic and pharmacological perturbations show that K+ conductance, Cl--sensitive pathways, and active transport reshape the amplitude, kinetics and even polarity of the optical response, revealing a context-dependent interplay between a passive molecular perturbation and endogenous bioelectric circuitry. As a functional proof of concept, kanamycin efficacy co-varies with the optically tuned voltage state. Together, these results establish MTP2 as a reversible chemical optostimulator for probing and controlling microbial electrophysiology.