Role of Integration Host Factor in the Transcriptional Activation of Flagellar Gene Expression in Caulobacter crescentus

Abstract

In the Caulobacter crescentus predivisional cell, class III and IV flagellar genes, encoding the extracytoplasmic components of the flagellum, are transcribed in the nascent swarmer compartment. This asymmetric expression pattern is attributable to the compartmentalized activity of the 54 -dependent transcriptional activator FlbD. Additionally, these temporally transcribed flagellar promoters possess a consensus sequence for the DNA-binding protein integration host factor (IHF), located between the upstream FlbD binding site and the promoter sequences. Here, we deleted the C. crescentus gene encoding the ␤-subunit of the IHF, ihfB (himD), and examined the effect on flagellar gene expression. The ⌬ihfB strain exhibited a mild defect in cell morphology and impaired motility. Using flagellar promoter reporter fusions, we observed that expression levels of a subset of class III flagellar promoters were decreased by the loss of IHF. However, one of these promoters, fliK-lacZ, exhibited a wild-type cell cycle-regulated pattern of expression in the absence of IHF. Thus, IHF is required for maximal transcription of several late flagellar genes. The ⌬ihfB strain was found to express significantly reduced amounts of the class IV flagellin, FljL, as a consequence of reduced transcriptional activity. Our results indicate that the motility defect exhibited by the ⌬ihfB strain is most likely attributable to its failure to accumulate the class IV-encoded 27-kDa flagellin subunit, FljL.Cell division in Caulobacter crescentus results in the generation of two dissimilar daughter cells, a sessile stalked cell and a motile swarmer cell. The synthesis of the single polarly localized flagellum occurs in a temporally and spatially regulated fashion over the course of each cell cycle (14,17,27). Over 50 C. crescentus genes are required for construction of the flagellum, with the majority organized within a trans-acting regulatory hierarchy of four classes (I to IV). The class I-encoded transcription factor CtrA activates the cell cycle-regulated expression of class II genes, encoding the cytoplasmic membrane-embedded MS ring, the flagellar switch, and the secretory apparatus of the flagellum. The expression and successful assembly of a class II-encoded structure is required for the transcriptional activation of class III genes, which encode the rod transversing the cytoplasmic membrane and the periplasm, the outer membrane rings, and the extracellular hook structure (37,48,65). The assembly of the hook structure is, in turn, required for translation of the flagellin subunits (class IV) that comprise the filament.The temporal regulation of class III and IV flagellar gene expression is influenced by both cell cycle events and the progression of flagellar assembly. These late flagellar genes share a conserved promoter sequence that includes a consensus binding site for RNA polymerase containing the 54 subunit (8,21,28,38,39,45,47) and one or more ftr enhancer elements recognized by the class II-encoded transcription factor FlbD (5,6,18,2...

Probing flagellar promoter occupancy in wild-type and mutant Caulobacter crescentus by chromatin immunoprecipitation

“…4a). FliX, the regulatory factor that links the status of flagellar assembly to FlbD activity (Muir & Gober, 2005), was present at the class II promoters, at higher levels than WT , in Δ fliG (170 ± 7%) and Δ tipF (144 ± 4%), consistent with the decreased levels of FlbD at the fliF promoter (Fig. 4a).…”

Linking structural assembly to gene expression: a novel mechanism for regulating the activity of a σ54 transcription factor

“…FlbD bound to enhancer DNA interacts with σ 54 ‐containing RNA polymerase (σ 54 ‐RNAP) bound to conserved promoter sequences located at −24 and −12 bp (indicated here as two black rectangles immediately to the left of the arrow) from the transcription start site. Interaction between FlbD and σ 54 ‐RNAP is facilitated by the DNA bending protein, IHF (Gober and Shapiro, 1990; Muir and Gober, 2005), that binds to promoter DNA located between (−75, −40) the enhancer sequences and the promoter. B.…”

Comparative analysis of Caulobacter chromosome replication origins

“…DnaA (Gorbatyuk & Marczynski, 2005), CtrA (Domian et al, 1997), IHF (Muir & Gober, 2005) and CcrM (Reisenauer et al, 1999) all have documented cellcycle variations in C. crescentus. Therefore, each protein could in principle contribute to replication control.…”