The transition from a planktonic (free-swimming) existence to growth attached to a surface in a biofilm occurs in response to environmental factors, including the availability of nutrients. We show that the catabolite repression control (Crc) protein, which plays a role in the regulation of carbon metabolism, is necessary for biofilm formation in Pseudomonas aeruginosa. Using phase-contrast microscopy, we found that a crc mutant only makes a dispersed monolayer of cells on a plastic surface but does not develop the dense monolayer punctuated by microcolonies typical of the wild-type strain. This is a phenotype identical to that observed in mutants defective in type IV pilus biogenesis. Consistent with this observation, crc mutants are defective in type IV pilus-mediated twitching motility. We show that this defect in type IV pilus function is due (at least in part) to a decrease in pilA (pilin) transcription. We propose that nutritional cues are integrated by Crc as part of a signal transduction pathway that regulates biofilm development.

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Crc (catabolite repression control) protein of Pseudomonas aeruginosa has shown to be involved in carbon regulation of several pathways. In this study, the role of Crc in catabolite repression control has been studied in Pseudomonas putida. The bkd operons of P. putida and P. aeruginosa encode the inducible multienzyme complex branched-chain keto acid dehydrogenase, which is regulated in both species by catabolite repression. We report here that this effect is mediated in both species by Crc. A 13-kb cloned DNA fragment containing the P. putida crc gene region was sequenced. Crc regulates the expression of branched-chain keto acid dehydrogenase, glucose-6-phosphate dehydrogenase, and amidase in both species but not urocanase, although the carbon sources responsible for catabolite repression in the two species differ. Transposon mutants affected in their expression of BkdR, the transcriptional activator of the bkd operon, were isolated and identified as crc and vacB (rnr) mutants. These mutants suggested that catabolite repression in pseudomonads might, in part, involve control of BkdR levels.Pseudomonads play an important role in nature because of their ability to metabolize natural and manufactured organic chemicals. Many of these compounds are environmental pollutants, such as benzene, toluene, xylene, ethylbenzene, styrene, and chlorobenzoates (18), and their removal has been named bioremediation. Although the enzymic pathways responsible for degradation of these pollutants may be effective when the target compound is the sole growth-supporting substrate, in nature these compounds are present as mixtures, and some substrates may be degraded preferentially. Catabolite repression control refers to the ability of an organism to preferentially metabolize one carbon source over another when both are present in the growth medium. Because of the importance of pseudomonads to bioremediation efforts, understanding the control of catabolite repression is important so that more efficient, genetically modified organisms can be utilized in the removal of these environmental pollutants.The molecular mechanisms of catabolite repression control have been extensively characterized in enteric bacteria, where glucose is the preferred carbon source. In these organisms, enzymes of the phosphoenolpyruvate-dependent phosphotransferase system mediate catabolite repression control by regulation of cyclic AMP (cAMP) concentration via adenylate cyclase activity (22). The strongest repressing substrates in Pseudomonas spp. are acetate, tricarboxylic acid cycle intermediates, and glucose (4, 10, 26). Unlike Escherichia coli, in Pseudomonas species adenylate cyclase activity, cAMP phosphodiesterase activity, and cAMP pools do not fluctuate with carbon source, nor does the addition of cAMP relieve repression of catabolite responsive pathways (21, 25). In addition, only one phosphotransferase system (fructose) has been identified in Pseudomonas (5), suggesting that PTS components are not involved in catabolite repression control in pseudomo...