Blue-light dependent inhibition of twitching motility in Acinetobacter baylyi ADP1: Additive involvement of three BLUF domain-containing proteins

Twitching motility in Acinetobacter baylyi ADP1 is inhibited by moderate intensities of blue light in a temperature-dependent manner (maximally at 20 degrees C. We analyzed the involvement of four predicted blue-light-sensing-using flavin (BLUF) domain-containing proteins encoded in the genome of this strain in the twitching motility phenotype. All four genes were expressed both in light and in darkness. A phylogenetic tree showed that one BLUF domain, ACIAD2110, grouped separately from the other three (ACIAD1499, ACIAD2125 and ACIAD2129). Individual knock-out mutants of the latter, but not of ACIAD2110, fully abolished the light dependency of the twitching motility response. Quantitative analysis of transcripts level of the three genes showed a decreased expression in the light, with dark/light ratios of 1.65 +/- 0.28, 1.79 +/- 0.21 and 2.69 +/- 0.39, for ACIAD2125, ACIAD2129 and ACIAD1499, respectively. Double and triple knock-outs of ACIAD1499, ACIAD2125 and ACIAD2129 confirmed the same phenotype as the corresponding single knock-outs. Complementation of all the single knock-outs and the triple knock-out mutants with any of the three BLUF-domain encoding genes fully restored the inhibition of twitching motility by blue light that is observed in the wild type strain. A. baylyi ADP1 therefore shows a high degree of redundancy in the genes that encode BLUF-containing photoreceptors. Moreover, all plasmid-complemented strains, expressing any of the BLUF- proteins irrespective of the specific set of deleted photoreceptors, displayed increased light-dependent inhibition of twitching motility, as compared to the wild type (p<<0.001). We conclude that the three genes ACIAD1499, ACIAD2125 and ACIAD2129 are jointly required to inhibit twitching motility under moderate blue light illumination.