The role of FIS in trans activation of stable RNA operons of E. coli.

Nilsson, Lars; Vanet, Anne; Vijgenboom, Erik; Bosch, L.

doi:10.1002/j.1460-2075.1990.tb08166.x

Cited by 182 publications

(181 citation statements)

References 33 publications

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“…A sequence similar to the E. coli factor for inversion stimulation (FIS) binding site (22) was found at positions 299 to 308 between the -35 regions of the trmA and btuB promoters (7 of 9 bp). FIS binding sites have also been found in the rrn promoters and are centered there at about -71, i.e., somewhat further upstream than the putative FIS binding site in the trmA promoter, which is centered at -56 (32,47). Cloning and sequence analysis of the trmA gene from S. typhimurium.…”

Section: Resultsmentioning

confidence: 99%

The trmA promoter has regulatory features and sequence elements in common with the rRNA P1 promoter family of Escherichia coli

et al. 1991

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The tRNA(m5U54)methyltransferase, whose structural gene is designated trmA, catalyzes the formation of 5-methyluridine in position 54 of all tRNA species in Escherichia coli. The synthesis of this enzyme has previously been shown to be both growth rate dependent and stringently regulated, suggesting regulatory features similar to those of rRNA. We have determined the complete nucleotide sequence of the trmA operon in E. coli and the sequence of the trmA promoter region in SalmoneUa typhimurium and also analyzed the transcriptional regulation of the gene. The trmA and the btuB (encoding the vitamin B12 outer membrane receptor protein) promoters are divergent promoters separated by 102 bp between the transcriptional start sites. The trmA promoters of both E. coli and S. typhimurium share promoter elements with the rRNA P1 promoter. The sequence downstream from the -10 region of the trmA promoter is homologous to the discriminatory region found in stringently regulated promoters. Next to and upstream from the -10 region is a sequence, TCCC, in the trmA promoter that is present in all of the seven rRNA P1 promoters and in some tRNA promoters but not in any other Co70 promoter. However, a similar motif is also found in promoters transcribed by the heat shock sigma factor CJ32. The trmA gene is transcribed as a monocistronic operon, and the 3' end of the transcript is shown to be located downstream from a dyad symmetry region not followed by a poly(U) stretch. Using a trmA-cat operon fusion, we show that the growth rate-dependent regulation of trmA resembles that of rRNA and operates at the level of transcription.

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Section: Resultsmentioning

confidence: 99%

The trmA promoter has regulatory features and sequence elements in common with the rRNA P1 promoter family of Escherichia coli

et al. 1991

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“…It was recently shown that FIS protein binds to an upstream activating sequence present in stable RNA operons, thereby activating transcription from downstream promoters (37). We therefore tested whether a deletion mutation infis affects proU expression.…”

Section: Methodsmentioning

confidence: 99%

“…Whether this is just a coincidence or whether it points to a functional role of the DNA bend for efficient proU expression is unclear at present. Sequence distributions associated with DNA curvature upstream of a number of procaryotic promoters with upstream activation of transcription have been described (44), and the binding of accessory proteins such as FIS and IHF to sequences upstream of promoters is implied in the activation of transcription (37,54). Our results with afis mutant show that FIS does not play any role in proU expression.…”

Section: Methodsmentioning

confidence: 99%

Characterization of mutations affecting the osmoregulated proU promoter of Escherichia coli and identification of 5' sequences required for high-level expression

Lucht

Bremer

1991

J Bacteriol

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Expression of the Escherichia coli proU operon, which encodes an efficient uptake system for the osmoprotectant glycine betaine, is strongly increased in cells grown at high osmolarity. We isolated 182 independent spontaneous mutants with elevated expression of the chromosomal 4'lproV-lacZ) (Hyb2) fusion at low osmolarity. Genetic analysis demonstrated that eight of these mutant strains carried mutations closely linked to the fusion, whereas all others carried mutations that appeared to be in osmZ. All of the mutations resulted in increased but still osmoregulated expression of the fI(proV-lacZ)(Hyb2) fusion. The proU-linked mutants carried an identical point mutation (proU603) which changes the -35 sequence of the proU promoter from TTGCCT to TTGACT and thereby increases the homology of the -35 region to the consensus sequence (TTGACA) of E. coli promoters. We also selected for mutants with decreased expression of the plasmid Escherichia coli and Salmonella typhimurium can adapt to high-osmolarity growth conditions by a variety of mechanisms (for recent overviews, see references 8 and 55). One of these mechanisms is the intracellular accumulation of the osmoprotectant glycine betaine, which is either synthesized from exogenously provided choline (28) or taken up from the environment (43). Two glycine betaine porters have been identified: the low-affinity ProP system (5, 32, 34) and the high-affinity ProU system (2,6,24,32).ProU is a binding-protein-dependent transport system and is encoded by the proU operon, which consists of three structural genes, proV, proW, and proX (20, 31, 41, 52). The level of proU expression is sensitively determined by the osmolarity of the growth medium. The basal transcription of proU is very low and is strongly stimulated upon a sudden osmotic upshock. The increased steady-state level of proU expression during growth at elevated osmolarity is directly correlated with the osmolarity of the growth medium (3,6,8,13,19,21,32

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“…However, it cannot be predicted if Fis binding might occlude or enhance binding of other proteins at the ␥ origin. Prebinding of Fis at its own promoter excludes RNA polymerase (1a), but Fis binding enhances RNA polymerase binding elsewhere (62,68).…”

Section: Discussionmentioning

confidence: 99%

Preponderance of Fis-binding sites in the R6K gamma origin and the curious effect of the penicillin resistance marker on replication of this origin in the absence of Fis

Ehley

et al. 1996

J Bacteriol

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Fis protein is shown here to bind to 10 sites in the ␥ origin of plasmid R6K. The Fis-binding sites overlap all the previously identified binding sites in the ␥ origin for the plasmid-encoded initiator protein and three host-encoded proteins, DnaA, integration host factor, and RNA polymerase. However, the requirement of Fis for R6K replication depends on the use of copy-up -protein variants and, oddly, the antibiotic resistance marker on the plasmid. In Fis-deficient cells, copy-up variants cannot drive replication of R6K ␥-origin plasmids carrying the bla gene encoding resistance to penicillin (Pen r ) but can drive replication of plasmids with the same origin but carrying the chloramphenicol acetyltransferase gene encoding chloramphenicol resistance (Cm r ). In contrast, R6K replication driven by wild-type is unaffected by the antibiotic resistance marker in the absence of Fis protein. Individually, none of these elements (copy-up , Fis deficiency, or drug markers) prevents R6K replication. The replication defect is not caused by penicillin in the medium or runaway replication and is unaffected by the orientation of the bla gene relative to the origin. Replication remains inhibited when part of the bla coding segment is deleted but the bla promoter is left intact. However, replication is restored by insertion of transcriptional terminators on either side of the ␥ origin, suggesting that excess transcription from the bla gene may inactivate replication driven by copy-up mutants in the absence of Fis. This study suggests that vector sequences such as drug markers may not be inconsequential in replication studies, as is generally assumed.The small, abundant DNA-binding proteins Fis integration host factor (IHF), and HU (for reviews, see references 14, 24, 25, and 69) have roles in a variety of DNA transactions, including replication of many replicons. Fis binds and bends oriC, the origin of replication of the chromosome (22, 30), and is involved in chromosome replication (22). IHF also binds oriC (21, 63), and both HU and IHF assist DnaA in unwinding the origin (37) and participate in replication of oriC minichromosomes (21,42). IHF also has roles in the replication and stable maintenance of phage f1 (33, 34) and plasmids pSC101 (3-5, 27, 74, 75) and R6K (11-13, 15, 17, 45).Plasmid R6K encodes resistance to penicillin (Pen r ) and streptomycin (49) and contains three origins of replication: ␣, ␥, and ␤ (Fig. 1). Each origin requires in cis the 277-bp core segment and an additional segment unique to that origin (47,48,(70)(71)(72). The ␥ origin, for example, includes the core and the adjacent enhancer (for a recent review, see reference 16). The enhancer contains DnaA box 1 (79, 80) and a small segment, stb, involved in stable maintenance of ␥-origin-containing plasmids (81). We subdivided the core into three distinct regions (16): (i) the AT-rich region, bound by and IHF (11,13,15,17,19,45,52); (ii) seven 22-bp direct repeats (DRs) bound by (19,23,28,60); and (iii) a multiprotein-binding region interacting with D...

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The role of FIS in trans activation of stable RNA operons of E. coli.

Cited by 182 publications

References 33 publications

The trmA promoter has regulatory features and sequence elements in common with the rRNA P1 promoter family of Escherichia coli

The trmA promoter has regulatory features and sequence elements in common with the rRNA P1 promoter family of Escherichia coli

Characterization of mutations affecting the osmoregulated proU promoter of Escherichia coli and identification of 5' sequences required for high-level expression

Preponderance of Fis-binding sites in the R6K gamma origin and the curious effect of the penicillin resistance marker on replication of this origin in the absence of Fis

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