Multiple structural proteins are required for both transcriptional activation and negative autoregulation of Caulobacter crescentus flagellar genes

Abstract

The periodic and sequential expression of flagellar (fla) genes in the Caulobacter crescentus cell cycle depends on their organization into levels I to IV of a regulatory hierarchy in which genes at the top of the hierarchy are expressed early in the cell cycle and are required for the later expression of genes below them. In these studies, we have examined the regulatory role of level IIfliF operon, which is located near the top of the hierarchy. The last gene in the fliF operon, flbD, encodes a transcriptional factor required for activation of j54-dependent promoters at levels III and IV and negative autoregulation of the level II fliF promoter. We have physically mapped the fliF operon, identified four new genes in the transcription unit, and determined that the organization of these genes is 5'-fliF-fliG-flbE-fliN-flbD-3'. Three of the genes encode homologs of the MS ring protein (FliF) and two switch proteins (FliG and FliN) of enteric bacteria, and the fourth encodes a predicted protein (FlbE) without obvious similarities to known bacterial proteins. We have introduced nonpolar mutations in each of the open reading frames and shown that all of the newly identified genes (fliFfliG,flbE, andfliN) are required in addition toflbD for activation of the u54-dependentflgK andflbG promoters at level III. In contrast,fliFfliG, andflbE, but notfliN, are required in addition toflbD for negative autoregulation of the level II fliF promoter. The simplest interpretation of these results is that the requirements of FlbD in transcriptional activation and repression are not identical, and we speculate that FlbD function is subject to dual or overlapping controls. We also discuss the requirement of multiple structural genes for regulation of levels II and III genes and suggest thatfla gene expression in C. crescentus may be coupled to two checkpoints in flagellum assembly.The procaryotic flagellum is a complex organelle composed of three structural elements: the basal body or motor, which is embedded in the membrane and peptidoglycan layers of the cell envelope; the external hook, which attaches the basal body to the flagellar filament; and the filament itself, which rotates to move the cell (35, 62). In the dimorphic bacterium Caulobacter crescentus, the flagellum is assembled at one pole of the dividing cell late in the cell cycle and then segregates with the motile swarmer cell at division. Its biosynthesis requires the activity of 40 to 50 flagellar (fla) genes (15), and, as in the enteric bacteria, expression of these genes is regulated by their organization in a regulatory hierarchy in which expression of genes at the top of the hierarchy is required for expression of genes lower in the hierarchy (reviewed in references 7 and 44). In the C. crescentus hierarchy, there is also evidence that fla gene interactions mediate the negative, as well as the positive, transcriptional control of gene expression (45,49,70).A unique feature of flagellum biosynthesis in C. crescentus is the cell cycle regulation of fla gene expre...

Regulation of the Caulobacter flagellar gene hierarchy; not just for motility

“…1B; Ramakrishnan et al, 1994). However, some of the flagellar genes appear to be unique to C. crescentus.…”

“…The requirement of multiple structural genes for the regulation of the Class III and Class IV genes has suggested that flagellar gene expression in C. crescentus is co-ordinated with flagellum biosynthesis by two assembly checkpoints: transcription of Class III genes requires completion of MS ring-switch assembly, and expression of the Class IV flagellin genes requires completion of basal body-hook assembly ( Fig. 2A; Ramakrishnan et al, 1994). Similar reasoning was invoked originally to propose that completion of the basal bodyhook structure is an assembly checkpoint controlling late flagellar gene transcription in E. coli ( Fig.…”

Aggregation in Azospirillum brasilense: effects of chemical and physical factors and involvement of extracellular components

“…Analysis of the DNA flanking the transposon insertion site revealed a high level of similarity to the fliG gene of CauZobacter crescentus (S. Moens, personal communication), which encodes a protein involved in the regulation of both the rotation and the biogenesis of the bacterial flagellum (Ramakrishnan et al, 1994). This, along with findings from the present work, indicates that even if the role of the polar flagellum in the aggregation process is not clear, both aggregation and flagellation processes could share one or more regulator genes.…”

Signal transduction mechanisms in Caulobacter crescentus development and cell cycle control

“…A potential kinase for FlbD is encoded by the £bE gene, which lies within the same operon as £bD and is required for activation of the c 54 -dependent class III and class IV promoters [25]. FlbE shares regions of homology with two-component histidine kinases and is able to phosphorylate FlbD in vitro [82].…”