CATM ENCODES A LYSR-TYPE TRANSCRIPTIONAL ACTIVATOR REGULATING CATECHOL DEGRADATION IN ACINETOBACTER-CALCOACETICUS

On the basis of the constitutive phenotypes of two catM mutants of Aci netobacter calcoaceticus, the CatM protein was proposed to repress exp ression of two different loci involved in catechol degradation, catA a nd catBCIJFD (E. Neidle, C. Hartnett; and L. N. Ornston, J. Bacteriol. 171:5410-5421, 1989). In spite of its proposed negative role as a rep ressor, CatM is similar in amino acid sequence to positive transcripti onal activators of the LysR family, Investigating this anomaly, we fou nd that insertional inactivation of catM did not cause the phenotype e xpected for the disruption of a repressor-encoding gene: in an interpo son-generated catM mutant, no cat genes were expressed constitutively, but rather catA and catB were still inducible by muconate. Moreover, this catM mutant grew poorly on benzoate, a process requiring the expr ession of all cat genes. The inducibility of the cat genes in this cat M mutant was completely eliminated by a 3.5-kbp deletion 10 kbp upstre am of catM. In this double mutant, catM in a ans restored muconate ind ucibility to both catA and catB. These results suggested the presence of an additional regulatory locus controlling cat gene expression. The ability of CatM to function as an activator was also suggested by the se results. In support of this hypothesis, in vivo methylation protect ion assays showed that CatM protects two guanines in a dyad 65 nucleot ides upstream of the catB transcriptional start site, in a location an d pattern typical of LysR-type transcriptional activators. Gel mobilit y shift assays indicated that CatM also binds to a region upstream of catA. DNA sequence analysis revealed a nucleotide near the 3' end of c atM not present in the published sequence. Translation of the correcte d sequence resulted in the deduced CatM protein being 52 residues long er than previously reported. The size, amino acid sequence, and mode o f action of CatM now appear similar to, and typical of, what has been found for transcriptional activators in the LysR family. Analysis of o ne of the constitutive alleles of catM previously thought to encode a dysfunctional repressor indicated instead that it encodes an inducer-i ndependent transcriptional activator.