Negative regulatory loci coupling flagellin synthesis to flagellar assembly in Salmonella typhimurium

Abstract

The complex regulation of flagellin gene expression in Salmonella typhimurium was characterized in vivo by using lac transcriptional fusions to the two flagellin structural genes (fliC [Hl] andfljB [H2]). Phase variation was measured as the rate of switching of flagellin gene expression. Switching frequencies varied from 1/500 per cell per generation to 1/10,000 per cell per generation depending on the particular insertion and the direction of switching. There is a 4-to 20-fold bias in favor of switching from thefljB(On) to thefljB(Of) orientation. Random TnlOdTc insertions were isolated which failed to express flagellin. While most of these insertions mapped to loci known to be required for flagellin expression, several new loci were identified. The presence of functional copies of all of the genes responsible for complete flagellar assembly, except the hook-associated proteins (flgK,flgL, andfliD gene products), were required for expression of thefliC orfljB flagellin genes. Two novel loci involved in negative regulation offliC andfljB infla mutant backgrounds were identified. One of these loci, designated the flgR locus, mapped to the flg operon at 23 min on the Salmonella linkage map. An flgR insertion mutation resulted in relief of repression of thefliC andfljB genes in allfla mutant backgrounds except for mutants in the positive regulatory loci (flhC, flhD, and fli4 genes).Flagellar phase variation in Salmonella typhimurium results from the reversible inversion of a 996-bp chromosomal segment of DNA ( rium and with the homologous genes in Escherichia coli has uncovered a complex hierarchy of regulation for these genes (26,31,32,53). In S. typhimurium one and presumably also the other of the flagellin genes are at the bottom of this regulatory scheme and are grouped in the class of late genes (Fig. 2) (32). The members of this class are dependent on the early and middle gene classes for their expression. If any of the genes in the early and middle classes do not express functional proteins, the genes in the late class are not expressed.Similarly, the genes in the middle class are dependent upon the early class for their expression but are regulated independently of any defects in the late genes (32). The early genes include the flhD and flhC genes, which are believed to encode positive activators of flagellar expression (29). These early genes are required for expression of the middle genes.The fliA gene, shown recently to encode a flagellar-specific sigma factor (38), is included among the middle genes. The fliA gene product is thought to be required for the expression of the late genes, including flagellin.At the top of this regulatory scheme is the cAMP-Catabolite gene activator protein complex, which acts at the early genes (flhDC operon) (29, 42). In cya or crp mutant strains or in wild-type cells grown in the presence of glucose, no flagella are synthesized (1, 54). These proposed classes or levels of regulation of flagellar assembly in S. typhimurium are similar to those proposed for E. coli by Kome...

Stress‐induced host membrane remodeling protects from infection by non‐motile bacterial pathogens

“…To test the relevance of bacterial motility in conditions of host cell stress, we used the Salmonella motility‐deficient mutant strain Δ fliC , which lacks the fliC gene but retains expression of FljB. However, the frequency of flagellin switching is low [approximately 1/5,000 per cell per generation, biased toward the fljB OFF orientation (Gillen & Hughes, 1991)], and thus, the Salmonella Δ fliC mutant strain is essentially non‐motile. Additionally, we used a completely non‐motile Salmonella mutant (Δ flhC ), due to an insertion of the MudJ transposon into the flagella master regulator flhC gene.…”

Characterization of the sigD transcription unit of Bacillus subtilis

“…In the enteric bacteria an intact hook-basal body complex must be assembled before flagellin is expressed (21,36), and genetic studies have demonstrated a hierarchy of flagellar assembly, motility, and chemotaxis gene expression responsible for this structural regulation (13,14,22,23). A negative regulator of flagellin expression, designated FlgM, that is responsible for the coupling of hook-basal body assembly to flagellin expression has been identified in S. typhimurium (13,14).…”

“…The cF factor of E. coli displays a promoter specificity similar to that of aD and has been shown to transcribe certain flagellar and chemotaxis genes in that organism (3). The gene for the uF factor was identified in S. typhimurium as being thefliA locus (32), part of a class of fla, che, and mot genes termed class II genes (13,14,22,23).…”

“…Class II gene expression is dependent on the expression of class I gene products, which have been proposed to act as transcription activators (5). Class II gene products are all required for transcription of class III genes (13,14,22,23), and this regulation is based on the need for the functional assembly of class II structural proteins into the hook and basal body and on the expression offliA (13,14,22,23). In the absence of such assembly, the FlgM repressor shuts off synthesis of class III genes by interaction with the fliA u factor (13,14).…”

“…Class II gene products are all required for transcription of class III genes (13,14,22,23), and this regulation is based on the need for the functional assembly of class II structural proteins into the hook and basal body and on the expression offliA (13,14,22,23). In the absence of such assembly, the FlgM repressor shuts off synthesis of class III genes by interaction with the fliA u factor (13,14). In the enteric bacteria, therefore, there appears to be a complex set of transcriptional and structural regulatory mechanisms that ensures that the flagellin gene is expressed and the flagellar filament is assembled only when a functional hook-basal body complex has been formed.…”