Transcriptional regulation of a periodically controlled flagellar gene operon in Caulobacter crescentus

Ohta, Noriko; Chen, Ling-Sing; Swanson, E C; Newton, Austin

doi:10.1016/0022-2836(85)90261-x

Cited by 79 publications

(108 citation statements)

References 31 publications

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“…The flaN and flbG operons (Fig. 1A) are transcribed at the same time in the cell cycle (5,29) and have similar genetic requirements for expression (5,23,30). Potential regulatory sequences in flaN and flbG have been identified either by their alignment with E. coli regulatory elements or by their similarity to sequence elements previously identified in C. crescentus (10,23,25).…”

Section: Resultsmentioning

confidence: 92%

“…These studies suggested that the C. crescentus fla genes, like those in Eschenchia coli (14,15) and Salmonella typhimurium (18), are organized into a regulatory hierarchy in which the expression of fla genes at each level depends on the expression of genes above them in the hierarchy (27,38). Moreover, recent results have shown that the time offla gene expression in the C. crescentus cell division cycle is determined by the relative position of a gene in the regulatory hierarchy (4,29 Thus, the adjacent and divergent flaN and flbG (hook) operons are expressed at about the same time in the cell cycle, and their transcription depends on flbF and genes in the flaO locus, which is now designated the fliF operon (5,23,30). The flbF and fliF operons are above flaN and flbG in the regulatory hierarchy (5,23) and are transcribed before them in the cell cycle (29,35).…”

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confidence: 99%

“…RNA was purified from C. crescentus cells as described before (30). DNA restriction fragment probes used in nuclease S1 protection assays were 5'-end-labeled with [-y-32P]ATP and T4 polynucleotide kinase (1).…”

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A sigma 54 promoter and downstream sequence elements ftr2 and ftr3 are required for regulated expression of divergent transcription units flaN and flbG in Caulobacter crescentus

Mullin

Newton

1993

J Bacteriol

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In this study, we investigated the cis-acting sequences required for transcription of the divergent, cell cycle-regulated flaN and flbG operons of Caulobacter crescentus. Previous work showed that transcription of flbG in vivo depends on a cr& promoter and a sequence element calledftrl that is located about 100 bp upstream from the transcription start site (D. A. Mullin and A. Newton, J. Bacteriol. 171:3218-3227, 1989). We now show that regulation offlaN transcription in vivo depends on a o54 promoter and two ftr elements located downstream of the transcription start site at +86 (ftr2) and +120 (ftr3). Mutations in or between the conserved elements at -24 and -12 in this or4 promoter reduced or abolishedflaN transcription, and one mutation that eliminated flaN expression led to an increased level offlbG transcript. Mutations in ftr2 resulted in greatly reduced levels offlaN transcript but had no noticeable effect on flbG transcript levels. All three mutations constructed inftr3 resulted in elevated flaN and flbG transcript levels. We conclude thatftr2 is required for positive regulation offlaN, whereasftr3 appears to play a negative regulatory role inflaN andflbG expression.To explain the coordinated positive activation and negative autoregulation of these two transcription units and the effect of mutations on gene expression, we propose a model in which theflaN andflbG promoters interact through alternative DNA looping to form structures that are transcriptionally active or inactive.Flagellum biosynthesis in Caulobacter crescentus requires more than 50 flagellar (fla) genes, and about half of these are organized in transcription units that are located in three major clusters on the chromosome. These fla genes are under strict temporal regulation, as indicated by the observation that they are transcribed periodically in the cell cycle at about the time that the gene products assemble to form the flagellum. The precisely ordered sequence of fla gene expression during flagellum biosynthesis and the availability of powerful biochemical and genetic tools have made C. crescentus an attractive model for studying the mechanism of temporal gene expression during cell differentiation. A central focus of these studies has been to explain, on the molecular level, how genes are turned on and off in a defined sequence during the cell division cycle (for a review, see reference 26).The role of trans-acting genes in the expression of C. crescentus fla genes was indicated first by epistasis experiments (3, 31) and later examined in detail by experiments that measured the levels of specific fla gene transcripts in different nonmotile mutants (27). These studies suggested that the C. crescentus fla genes, like those in Eschenchia coli (14, 15) and Salmonella typhimurium (18), are organized into a regulatory hierarchy in which the expression of fla genes at each level depends on the expression of genes above them in the hierarchy (27,38 identified, but there is evidence that a step in the DNA synthesis pathway is required for the...

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Section: Resultsmentioning

confidence: 92%

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confidence: 99%

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A sigma 54 promoter and downstream sequence elements ftr2 and ftr3 are required for regulated expression of divergent transcription units flaN and flbG in Caulobacter crescentus

Mullin

Newton

1993

J Bacteriol

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“…flbD and flbF are of particular interest in relation to the regulatory hierarchy because they are required in trans for expression of several flagellar gene transcription units, including flaN, flbG, flgL, and flgK (30,32,38). Analysis of theflaO operon DNA sequence revealed thatflbD codes for a 52-kDa protein, and its inferred amino acid sequence and domain organization is very similar to that of the NtrC(NRI) protein (43,51).…”

Section: Discussionmentioning

confidence: 99%

“…Flagellum biosynthesis and function in C. crescentus is a complex process that requires at least 48 genes, and about half of these are contained in three major gene clusters (11). The C. crescentus flagellar genes (6,7,36,38,55), like those in Escherichia coli (23,24) and Salmonella typhimurium (26), can be arranged in a regulatory hierarchy in which trans-acting genes at one level are required for expression of genes at lower levels. The flbD and flbF genes occupy high levels in the C. crescentus regulatory hierarchy because they are required in trans for the expression of genes needed for the synthesis of the flagellar hook and filament (30,32,36,39).…”

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Characterization of the Caulobacter crescentus flbF promoter and identification of the inferred FlbF product as a homolog of the LcrD protein from a Yersinia enterocolitica virulence plasmid

1992

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We have investigated the organization and expression of the Caulobacter crescentus flbF gene because it occupies a high level in the flagellar gene regulatory hierarchy. The nucleotide sequence comprising the 3' end of theflaO operon and the adjacentflbF promoter and structural gene was determined, and the organization of transcription units within this sequence was investigated. We located the 3' ends of theflaO operon transcript by using a nuclease S1 protection assay, and the 5' end of theflbF transcript was precisely mapped by primer extension analysis. The nucleotide sequence upstream from the 5' end of the flbF transcript contains -10 and -35 elements similar to those found in promoters transcribed by C28 RNA polymerase in other organisms. Mutations that changed nucleotides in the -10 or -35 elements or altered their relative spacing resulted in undetectable levels of flbF transcript, demonstrating that these sequences contain nucleotides essential for promoter function. We identified a 700-codon open reading frame, downstream from theflbF promoter region, that was predicted to be theflbF structural gene. The amino-terminal half of the FlbF amino acid sequence contains eight hydrophobic regions predicted to be membrane-spanning segments, suggesting that the FlbF protein may be an integral membrane protein. The FlbF amino acid sequence is very similar to that of a transcriptional regulatory protein called LcrD that is encoded in the highly conserved low-calcium-response region of virulence plasmid pYV03 in Yersinia enterocolitica (A.-M. Viitanen, P. Toivanen, and M. Skurnik, J. Bacteriol. 172:3152-3162, 1990).The Caulobacter crescentus flagellum has been used as a model for investigating cell differentiation because its morphogenesis is under strict spatial and temporal control in the cell division cycle (for reviews, see references 35 and 48). Flagellum biosynthesis and function in C. crescentus is a complex process that requires at least 48 genes, and about half of these are contained in three major gene clusters (11). The C. crescentus flagellar genes (6, 7, 36, 38, 55), like those in Escherichia coli (23,24) and Salmonella typhimurium (26), can be arranged in a regulatory hierarchy in which trans-acting genes at one level are required for expression of genes at lower levels. The flbD and flbF genes occupy high levels in the C. crescentus regulatory hierarchy because they are required in trans for the expression of genes needed for the synthesis of the flagellar hook and filament (30,32,36,39). These genes that depend upon flbD and flbF for their transcription contain conserved cis-acting sequences that include ur54-type promoters, a sequence calledftr (30, 32, 33, 37), and another sequence that matches the consensus binding site for E. coli integration host factor protein (15). flbD is a member of the flaO operon (39), whose transcription in vivo depends upon a sequence similar to that of the -10 box of E. coli promoters that are transcribed by cr32 RNA polymerase (34,52). It is evident that the promot...

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Temporal and spatial regulation of developmentally expressed genes in Caulobacter

Gober

Shapiro

1991

BioEssays

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Fundamental problems in developmental biology, such as the generation of asymmetry, differential transcription, and the execution of positional information, are all exhibited by a prokaryotic systcm, the Caulobacter cell cycle. Progeny cells resulting from each cell division have different developmental programs with respect to patterns of transcription and the initiation of DNA replication. The binary fission that yields dissimilar progeny results from asymmetry generated in the parent cell. An aspect of this fundamental developmental event is the temporally-controlled biogenesis of a polar flagellum. Several flagellar promoters require the histone-like protein, integration host factor (IEIF) for transcription activation. IHF introduces bends in DNA resulting in the reorganization of the higher-order structure of the promoter region. These promoters appear to be transcribed from the chromosome in the swarmer pole of the predivisional cell. The newly replicated chromosomes in the predivisional cell differ in sedimentation coefficient and in the ability to initiate replication in the progeny cell. We hypothesize that the differential higher order structure of the two chromosomes contributes to both replication potential and transcription specificity.

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Transcriptional regulation of a periodically controlled flagellar gene operon in Caulobacter crescentus

Cited by 79 publications

References 31 publications

A sigma 54 promoter and downstream sequence elements ftr2 and ftr3 are required for regulated expression of divergent transcription units flaN and flbG in Caulobacter crescentus

A sigma 54 promoter and downstream sequence elements ftr2 and ftr3 are required for regulated expression of divergent transcription units flaN and flbG in Caulobacter crescentus

Characterization of the Caulobacter crescentus flbF promoter and identification of the inferred FlbF product as a homolog of the LcrD protein from a Yersinia enterocolitica virulence plasmid

Temporal and spatial regulation of developmentally expressed genes in Caulobacter

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