Phototaxis refers to an organism’s movement toward a light source, while photophobotaxis involves movement into illuminated regions. Although phototaxis in cyanobacteria has been widely studied, photophobotaxis has been investigated in only a few species. In this study, we examined 18 cyanobacterial species and found that all single-celled and filamentous species exhibiting twitching motility performed photophobotaxis. In contrast, immobile Nostocales and a pilus-free Synechocystis did not display this behavior. The widespread occurrence of photophobotaxis aligns with the universality of photosystems, suggesting photosystems act as light sensors for this phenomenon. The photosystem II inhibitor DCMU disrupted photophobotaxis in single-celled cyanobacteria at a concentration of 10 µM, while filamentous species required ≥100 µM DCMU for inhibition. Despite the higher required concentration in filaments, the inhibition appears specific to photosystem II electron transport. This increased threshold may reflect heightened sensitivity due to collective sensing by multiple cells in the filament. Previous studies on spectral sensitivity and the cyanobacteriochrome PixJ in Phormidium lacuna identified PixJ as a negative regulator of photophobotaxis. In pixJ mutants, light sensitivity was increased compared to the wild type. Dual-wavelength experiments confirmed that yellow light induces PixJ to downregulate photophobotaxis. A paradox finding emerged in long-distance movements between two light sources: filaments moved faster than predicted by a random movement model, suggesting inter-filament communication. These results provide new insights into the mechanisms and regulation of photophobotaxis in cyanobacteria.

Cyanobacterium Phormidium lacuna filaments move from dark to illuminated areas by twitching motility. Time-lapse recordings demonstrated that this photophobotaxis response was based on random movements with movement reversion at the light-dark border. The filaments in the illuminated area form a biofilm attached to the surface. The wild-type and two mutants, pixJ and cphA, were investigated for photophobotaxis at diverse wavelengths and intensities. CphA is a cyanobacterial phytochrome; PixJ is a biliprotein with a methyl-accepting chemotaxis domain and is regarded as a phototaxis photoreceptor. The cphA mutant exhibits reduced biofilm surface binding. The pixJ mutant was characterized by greater photophobotaxis sensitivity. Thus, PixJ is a negative photophobotaxis regulator and does not serve as a light direction sensor. 3-(3,4-dichlorophenyl)1,1-dimethylurea (DCMU) blocks electron transfer in PS II. At concentrations of 100 and 1000 µM DCMU, photophobotaxis was inhibited, whereas motility was not significantly affected. This specific effect indicates PS II’s central role in light-direction sensing. It is argued that the intracellular concentrations of regular photoreceptors, including CphA or PixJ, are too small for a filament to sense rapid light intensity changes in very weak light. Three arguments, specific inhibition by DCMU, broad spectral sensitivity, and sensitivity against weak light, support photosynthesis pigments use as photophobotaxis sensors.