FimW Is a Negative Regulator Affecting Type 1 Fimbrial Expression inSalmonella entericaSerovar Typhimurium

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Abstract
Type 1 fimbriae are proteinaceous surface appendages that carry adhesins specific for mannosylated glycoproteins. These fimbriae are found on most members of the familyEnterobacteriaceaeand are known to facilitate binding to a variety of eukaryotic cells, including those found on the mucosal surfaces of the alimentary tract. We have shown that the regulation of type 1 fimbrial expression inSalmonella entericaserovar Typhimurium is controlled, in part, by the products of four genes found within thefimgene cluster:fimZ,fimY,fimW, andfimU. To better understand the specific role of FimW in fimbrial expression, a mutation was constructed in this gene by the insertion of a kanamycin resistance DNA cassette into the chromosome. The resultingfimWmutation was characterized by mannose-sensitive hemagglutination and agglutination with fimbria-specific antiserum. Assays suggested that this mutant was more strongly fimbriate than the parental strain, exhibiting a four- to eightfold increase in fimbrial production. ThefimWmutation was introduced into a second strain ofSalmonella entericaserovar Typhimurium, and this mutant was also found to be strongly fimbriate compared to the parental strain. Consistent with the role of this protein as a negative regulator,fimA-lacZexpression in serovar Typhimurium, as well as inEscherichia coli, was increased twofold in the absence of functional FimW. Primer extension analysis determined thatfimWtranscription is initiated from its own promoter 31 bp upstream of the translation start site. Analysis using afimW-lacZreporter indicated thatfimWexpression in serovar Typhimurium was increased under conditions that select for poorly fimbriate bacteria and lowfimAexpression. FimW also appears to act as an autoregulator, since expression from thefimW-lacZreporter was increased in afimWmutant. FimW was partially purified by fusion with theE. colimaltose-binding protein. Use of this FimW protein extract, as well as others, in DNA-binding assays was unable to identify a specific binding site for FimW in thefimA,fimZ,fimY, orfimWpromoter regions. To analyze protein-protein interactions, FimW was expressed in a LexA-based two-hybrid system inE. coli. A significant interaction between FimW and the DNA-binding activator protein, FimZ, was detected using this system. These results indicate that FimW is a negative regulator of serovar Typhimurium type 1 fimbrial expression and may function by interfering with FimZ-mediated activation offimAexpression.