Cells of the Gram-negative bacterium Myxococcus xanthus respond to blue light by producing carotenoids, pigments that play a protective role against the oxidative effects of light. Blue light triggers a network of regulatory actions that lead to the transcriptional activation of the structural genes for carotenoid synthesis. The product of carF, similar to a family of proteins of unknown function called Kua, is an early regulator of this process. Previous genetic data indicate that CarF participates in the light-dependent inactivation of the antisigma factor CarR. In the dark, CarR sequesters the ECF-sigma factor CarQ to the membrane, thereby preventing the activation of the structural genes for carotenoid synthesis. Using a bacterial two-hybrid system, we show here that both CarF and CarQ physically interact with CarR. These results, together with the finding that CarF is located at the membrane, support the hypothesis that CarF acts as an anti-antisigma factor. Comparison of CarF with other Kua proteins shows a remarkable conservation of a number of histidine residues. The effects on CarF function of several histidine to alanine substitutions and of the truncation of specific CarF domains are also reported here.