Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Factor For Inversion Stimulation Protein”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 73 records · Page 4Linked to original sources
PubMed2000

Coactivation of the RpoS-dependent proP P2 promoter by fis and cyclic AMP receptor protein.

The Escherichia coli proP P2 promoter, which directs the expression of an integral membrane transporter of proline, glycine betaine, and other osmoprotecting compounds, is induced upon entry into stationary phase to protect cells from osmotic shock. Transcription from the P2 promoter is completely dependent on RpoS (sigma(38)) and Fis. Fis activates transcription by binding to a site centered at -41, which overlaps the promoter, where it makes a specific contact with the C-terminal domain of the alpha subunit of RNA polymerase (alpha-CTD). We show here that Fis and cyclic AMP (cAMP) receptor protein (CRP)-cAMP collaborate to activate transcription synergistically in vitro. Coactivation both in vivo and in vitro is dependent on CRP binding to a site centered at -121.5, but CRP without Fis provides little activation. The contribution by CRP requires the correct helical phasing of the CRP site and a functional activation region 1 on CRP. We provide evidence that coactivation is achieved by Fis and CRP independently contacting each of the two alpha-CTDs. Efficient transcription in vitro requires that both activators must be preincubated with the DNA prior to addition of RNA polymerase.

Bacterial Proteins↗
PubMed2002

Factors affecting start site selection at the Escherichia coli fis promoter.

Transcription initiation with CTP is an uncommon feature among Escherichia coli sigma(70) promoters. The fis promoter (fis P), which is subject to growth phase-dependent regulation, is among the few that predominantly initiate transcription with CTP. Mutations in this promoter that cause a switch from utilization of CTP to either ATP or GTP as the initiation nucleotide drastically alter its growth phase regulation pattern, suggesting that the choice of the primary initiating nucleotide can significantly affect its regulation. To better understand what factors influence this choice in fis P, we made use of a series of promoter mutations that altered the nucleotide or position used for initiation. Examination of these promoters indicates that start site selection is determined by a combination of factors that include preference for a nucleotide distance from the -10 region (8 > 7 > 9 >> 6 >> 10 > 11), initiation nucleotide preference (A = G >> CTP > or = UTP), the DNA sequence surrounding the initiation region, the position of the -35 region, and changes in the intracellular nucleoside triphosphate pools. We describe the effects that each of these factors has on start site selection in the fis P and discuss the interplay between position and nucleotide preference in this important process.

Carrier Proteins↗
PubMed1990

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

The thrU(tufB) operon of Escherichia coli is endowed with a cis-acting region upstream of the promoter, designated UAS for Upstream Activator Sequence. A protein fraction has been isolated that binds specifically to DNA fragments of the UAS, thus forming three protein-DNA complexes corresponding to three binding sites on the UAS. It stimulates in vitro transcription of the operon by facilitating the binding of the RNA polymerase to the promoter. All three protein-DNA complexes contain one and the same protein. Dissociation constants for the three complexes have been determined, the lowest being in the sub-nanomolar range. The protein also binds to the UAS of the tyrT operon and to the UAS upstream of the P1 promoter of the rrnB operon, suggesting that transcription of the three operons, if not of more stable RNA operons, is activated by a common trans activator. We demonstrate that the E.coli protein FIS (Factor for Inversion Stimulation) also binds to the UAS of the thrU(tufB) operon forming three protein-DNA complexes. A burst of UAS- and FIS-dependent promoter activity is observed after reinitiation of growth of stationary cultures in fresh medium.

Carrier Proteins↗
PubMed1997

Activation of RpoS-dependent proP P2 transcription by the Fis protein in vitro.

The proP gene, encoding a transporter of the osmoprotecting compounds proline and glycine betaine, is expressed from two promoters. Transcription of the P2 promoter occurs at a transient period in late exponential phase and is dependent upon Fis and the RpoS (sigma38) sigma factor. Here we characterize Fis-mediated activation of the P2 promoter in vitro. We find that this promoter displays unusually high specificity for sigma38. Fis strongly activates P2 when bound to site I centered at -41 within the promoter region. There is a complex relationship involving DNA supercoiling and potassium glutamate concentration on Fis activation, but most efficient transcription occurs under high salt conditions when the superhelical density is above -0.03. The major stimulatory effect of DNA supercoiling occurs between superhelical densities of 0 to -0.02 suggesting that, while supercoiling is mechanistically important, it may not be a physiologically relevant controlling factor. However, the stimulation of transcription by high potassium glutamate concentrations may contribute to the osmotic inducibility of the P2 promoter. We show that Fis and E sigma38 bind cooperatively on supercoiled DNA to form a stable complex at P2 that involves promoter melting. Fis also binds to a second site within the proP regulatory region. While binding to this site appears to play no role in Fis activation of the P2 promoter, it functions as a repressor of transcription initiating from the P1 promoter by either sigma70 or sigma38.

Bacterial Proteins↗
PubMed2005

Low-force DNA condensation and discontinuous high-force decondensation reveal a loop-stabilizing function of the protein Fis.

We report single-DNA-stretching experiments showing that the protein Fis, an abundant bacterial chromosome protein of E. coli, mediates a dramatic DNA condensation to zero length. This condensation occurs abruptly when DNA tension is reduced below a protein-concentration-dependent threshold f* < 1 pN. Following condensation, reopening under larger forces proceeds via a series of discrete jumps, indicating that Fis is able to stabilize DNA crossings. Our experiments suggest that Fis may play a role in vivo stabilizing the "loop-domain" structure of the bacterial chromosome.

DNA, Bacterial↗
PubMed1997

Fis is required for illegitimate recombination during formation of lambda bio transducing phage.

Specialized transducing particles of phage lambda are formed by illegitimate recombination during prophage induction. We examined the effects of an Esherichia coli int, xis, himA, himD, or fis mutation on illegitimate recombination during formation of lambda Spi- phage, a class of lambda bio transducing phage. This type of phage is distinguishable from the docL and docR particles, which contain one cohesive end and are formed by cutting of the cos site, by plaque formation of lambda bio on Escherichia coli P2 lysogens. The yields of lambda Spi- phage in the int, xis, int-xis deletion, and b2 deletion mutants were about 50- to 200-fold higher than that of the wild-type prophage when bacteria were irradiated with UV light. This result indicates that Int and Xis functions, and the att site, are not required for illegitimate recombination. The yield of lambda Spi- phage in the himA, himD, or fis mutant carrying lambda delta int-xis prophage was 2.6-, 3.3-, or 17-fold lower, respectively, than that in the wild-type bacteria under UV irradiation. Analysis of the nucleotide sequences of the junctions of the transducing phages indicates that recombination at the hotspots, as well as at non-hotspots, takes place between short homologous sequences. Because the growth of infecting phages was not suppressed by the himA, himD, or fis mutation, we conclude that Fis is required, but IHF is only partially required, for short-homology-dependent illegitimate recombination during the formation of lambda bio transducing phage.

Bacterial Proteins↗
PubMed2003

Proteome analysis of Salmonella enterica serovar Typhimurium fis mutant.

Salmonella enterica serovar Typhimurium is an enteric pathogen and a principal cause of gastroenteritis in humans. The factor-for-inversion stimulation protein (Fis) is known to play a pivotal role in the expression of Salmonella pathogenicity island (SPI)-1 genes in addition to various cellular processes such as recombination, replication, and transcription. In order to understand Fis function in pathogenicity of Salmonella, we performed two-dimensional gel electrophoresis and identified proteins whose expression pattern is affected by Fis using mass spectrometry. The results revealed various proteins that can be grouped according to their respective cellular functions. These groups include the genes involved in the metabolism of sugar, flagella synthesis, translation, and SPI expression. Changes in SPI expression suggest the possibility that regulation of genes in SPI-2 as well as SPI-1 is affected by Fis.

Bacterial Proteins↗
PubMed2001

Escherichia coli ribosome-associated protein SRA, whose copy number increases during stationary phase.

Protein D has previously been demonstrated to be associated with Escherichia coli ribosomes by the radical-free and highly reducing method of two-dimensional polyacrylamide gel electrophoresis. In this study, we show that protein D is exclusively present in the 30S ribosomal subunit and that its gene is located at 33.6 min on the E. coli genetic map, between ompC and sfcA. The gene consists of 45 codons, coding for a protein of 5,096 Da. The copy number of protein D per ribosomal particle varied during growth and increased from 0.1 in the exponential phase to 0.4 in the stationary phase. For these reasons, protein D was named SRA (stationary-phase-induced ribosome-associated) protein and its gene was named sra. The amount of SRA protein within the cell was found to be controlled mainly at the transcriptional level: its transcription increased rapidly upon entry into the stationary phase and was partly dependent on an alternative sigma factor (sigma S). In addition, global regulators, such as factor inversion stimulation (FIS), integration host factor (IHF), cyclic AMP, and ppGpp, were found to play a role either directly or indirectly in the transcription of sra in the stationary phase.

Amino Acid Sequence↗
PubMed1997

Induction and repair of cyclobutane pyrimidine dimers in the Escherichia coli tRNA gene tyrT: Fis protein affects dimer induction in the control region and suppresses preferential repair in the coding region of the transcribed strand, except in a short region near the transcription start site.

We analysed induction and repair of UV induced pyrimidine dimers in the Escherichia coli tRNA gene tyrT. In wild-type (WT) log or stationary phase different patterns of induction occurred in the three Fis binding sites and the core promoter -35 sequence of the control region: this was absent in fis- cells. In stationary WT cells, slow, similar rates of repair occurred throughout the non-transcribed strand (NTS). Faster repair occurred in the NTS control region in WT log phase. NTS repair in fis- cells was similar, except the control region differed less between phases. Heterogeneous repair occurred along the transcribed strand (TS). In the control region repair was faster than in the NTS. Repair in the TS coding region changed between growth phases or if repair took place in different media. When irradiated log phase WT cells were in rich medium, two TS domains were evident: a fast-repaired domain within 31 nucleotides from the transcription start site; and a more slowly repaired domain composed of the rest of the TS. A sharp gradient existed in the small domain with very fast repair at the beginning and diminished repair towards the end. Fast transcription coupled repair (TCR) in the small domain was absent in the TS large domain, where repair was similar to the NTS and to the entire TS in mfd- cells. In similarly treated stationary phase WT cells, TCR occurred in the large domain. Depletion of Fis reinstates TCR to a lesser extent, whilst a substitution of five nucleotides at the Fis binding sites in the upstream activating sequence reinstates TCR. Reinstatement of TCR was also achieved by incubating irradiated WT cells in minimal salt medium without the required amino acid. Our results suggest that Fis indirectly suppresses preferential repair in the TS large domain by stimulating transcription.

Base Sequence↗
PubMed1998

The FIS protein fails to block the binding of DnaA protein to oriC, the Escherichia coli chromosomal origin.

The Escherichia coli chromosomal origin contains several bindings sites for factor for inversion stimulation (FIS), a protein originally identified to be required for DNA inversion by the Hin and Gin recombinases. The primary FIS binding site is close to two central DnaA boxes that are bound by DnaA protein to initiate chromosomal replication. Because of the close proximity of this FIS site to the two DnaA boxes, we performed in situ footprinting with 1, 10-phenanthroline-copper of complexes formed with FIS and DnaA protein that were separated by native gel electrophoresis. These studies show that the binding of FIS to the primary FIS site did not block the binding of DnaA protein to DnaA boxes R2 and R3. Also, FIS appeared to be bound more stably to oriC than DnaA protein, as deduced by its reduced rate of dissociation from a restriction fragment containing oriC . Under conditions in which FIS was stably bound to the primary FIS site, it did not inhibit oriC plasmid replication in reconstituted replication systems. Inhibition, observed only at high levels of FIS, was due to absorption by FIS binding of the negative superhelicity of the oriC plasmid that is essential for the initiation process.

Bacterial Proteins↗
PubMed2004

The Escherichia coli Fis promoter is regulated by changes in the levels of its transcription initiation nucleotide CTP.

Expression of the Escherichia coli nucleoid-associated protein Fis (factor for inversion stimulation) is controlled at the transcriptional level in accordance with the nutritional availability. It is highly expressed during early logarithmic growth phase in cells growing in rich medium but poorly expressed in late logarithmic and stationary phase. However, fis mRNA expression is prolonged at high levels throughout the logarithmic and early stationary phase when the preferred transcription initiation site (+1C) is replaced with A or G, indicating that initiation with CTP is a required component of the regulation pattern. We show that RNA polymerase-fis promoter complexes are short lived and that transcription is stimulated over 20-fold from linear or supercoiled DNA if CTP is present during formation of initiation complexes, which serves to stabilize these complexes. Use of fis promoter fusions to lacZ indicated that fis promoter transcription is sensitive to the intracellular pool of the predominant initiating NTP. Growth conditions resulting in increases in CTP pools also result in corresponding increases in fis mRNA levels. Measurements of NTP pools performed throughout the growth of the bacterial culture in rich medium revealed a dramatic increase in all four NTP levels during the transition from stationary to logarithmic growth phase, followed by reproducible oscillations in their levels during logarithmic growth, which later decrease during the transition from logarithmic to stationary phase. In particular, CTP pools fluctuate in a manner consistent with a role in regulating fis expression. These observations support a model whereby fis expression is subject to regulation by the availability of its initiating NTP.

Base Sequence↗
PubMed1995

The transcriptional activator protein FIS: DNA interactions and cooperative interactions with RNA polymerase at the Escherichia coli rrnB P1 promoter.

The E. coli rrnB P1 promoter owes its strength, in part, to the transcriptional activator protein FIS. FIS binds to three sites upstream of the RNA polymerase (RNAP) binding site and increases transcription in vivo four to ten-fold. In this report, hydroxyl radical and DMS footprinting analyses show that FIS binds to its three sites along one side of the DNA helix, and that FIS bound at the promoter-proximal site (site I) and RNAP bound at the promoter are in close proximity. The binding of FIS at site I and RNAP at the promoter are mutually cooperative. These observations support a model for direct interaction between the FIS protein bound at site I and RNAP in transcription activation at rrnB P1. We also find that FIS does not bind cooperatively to its three sites upstream of rrnB P1, and that the relatively small activation associated with FIS bound at sites II and III does not result indirectly by facilitation of binding of FIS to site I.

Base Sequence↗
PubMed1995

Effects of base substitutions on the binding of a DNA-bending protein.

In order to investigate whether the 2-amino group of guanine, which lies in the minor groove of the B-form helix, can directly influence DNA flexibility and major groove recognition by proteins we have examined the properties of DNA molecules containing inosine and/or 2,6-diaminopurine (DAP) residues. Appropriately substituted tyrT(A93) DNA fragments were prepared by the polymerase chain reaction. Their mobility in non-denaturing gels was affected, consistent with changed anisotropic flexibility leading to increased curvature due to G-->I substitution and decreased curvature due to replacement of adenine with DAP. Band-shift assays of FIS protein binding revealed facilitated interaction with inosine-containing DNA and markedly reduced binding to DAP-containing DNA, attributable to altered bendability. DNase footprinting experiments confirmed that fewer sites would bind FIS in DAP-containing DNA at a given protein concentration, whereas higher levels of binding occurred with inosine-containing molecules. Thus base substitutions which affect the placement and presence of the purine 2-amino group in the minor groove can affect both the intrinsic curvature and the bendability of DNA.

2-Aminopurine↗
PubMed2002

Architecture of Fis-activated transcription complexes at the Escherichia coli rrnB P1 and rrnE P1 promoters.

The transcription factor Fis activates the Escherichia coli rRNA promoters rrnB P1 and rrnE P1 by binding to sites centered at -71 and -72, respectively, and interacting with the C-terminal domain of the alpha subunit of RNA polymerase (RNAP alphaCTD). To understand the mechanism of activation by Fis at these promoters, we used oriented alpha-heterodimeric RNAPs and heterodimers of Fis to determine whether one or both subunits of alpha and Fis participate in the alphaCTD-Fis interaction. Our results imply that only one alphaCTD in the alpha dimer and only one activation-proficient subunit in the Fis dimer are required for activation by Fis. A library of alanine substitutions in alpha was used to identify the alphaCTD determinants required for Fis-dependent transcription at rrnB P1 and rrnE P1. We propose that the transcriptional activation region of the promoter-proximal subunit of the Fis dimer interacts with a determinant that includes E273 of one alphaCTD to activate transcription. We further suggest that the Fis contact to alphaCTD results in alphaCTD interactions with DNA that differ somewhat from those that occur at UP elements in the absence of Fis. The accompanying paper shows that the 273 determinant on alphaCTD is also targeted by Fis at the proP P2 promoter where the activator binds overlapping the -35 hexamer. Thus, similar Fis-alphaCTD interactions are used for activation of transcription when the activator is bound at very different positions on the DNA.

Base Sequence↗
PubMed1989

Bent DNA is needed for recombinational enhancer activity in the site-specific recombination system Cin of bacteriophage P1. The role of FIS protein.

A series of recombinational enhancer mutants was constructed by manipulating the ClaI site between the two FIS binding sites of the Hin enhancer. These mutants include insertions from two to 12 base-pairs and two deletions of one or two base-pairs. Recombinational enhancer activity was found only with four mutants carrying either a four base-pair substitution, ten base-pair insertions or a one base-pair deletion, respectively; two other ten base-pair insertion mutants, however, were inactive, although FIS protein binding was unaffected. So, besides binding of FIS protein to its specific sites within the enhancer sequence and the correct helical positioning of these sites on the DNA, another criterion for enhancer activity must be fulfilled. DNA bending assays identify this requirement as a change of the enhancer DNA conformation, which FIS protein is able to induce and to stabilize. This conformational change of the DNA can be blocked by mutations in the central segment between the two FIS binding sites of the Hin enhancer. This sequence has special functions for the recombinational enhancer activity.

Bacteriophages↗
PubMed1997

A model for Fis N-terminus and Fis-invertase recognition.

In this modeling, we show that the elusive Fis N-terminus involved in invertase-mediated site-specific DNA inversion is, by all indications, a hinged flapping loop. The prediction is based on a combined sequence and secondary structure alignment against known structures of protein segments, as well as a tetrapeptide fragment observed crystallographically. Its validity is strongly supported by the ability to interpret consistently the available mutagenesis data pertaining to this region including, especially, a series of deletion mutants which until this work had been a puzzle in the search for structural explanations. A model for Fis-invertase recognition is also proposed.

Amino Acid Sequence↗
PubMed2001

Genome organisation and chromatin structure in Escherichia coli.

We have analysed the complete sequence of the Escherichia coli K12 isolate MG1655 genome for chromatin-associated protein binding sites, and compared the predicted location of predicted sites with experimental expression data from 'DNA chip' experiments. Of the dozen proteins associated with chromatin in E. coli, only three have been shown to have significant binding preferences: integration host factor (IHF) has the strongest binding site preference, and FIS sites show a weak consensus, and there is no clear consensus site for binding of the H-NS protein. Using hidden Markov models (HMMs), we predict the location of 608 IHF sites, scattered throughout the genome. A subset of the IHF sites associated with repeats tends to be clustered around the origin of replication. We estimate there could be roughly 6000 FIS sites in E. coli, and the sites tend to be localised in two regions flanking the replication termini. We also show that the regions upstream of genes regulated by H-NS are more curved and have a higher AT content than regions upstream of other genes. These regions in general would also be localised near the replication terminus.

Bacterial Proteins↗
PubMed1996

Identification of new Fis binding sites by DNA scission with Fis-1,10-phenanthroline-copper(I) chimeras.

The chimeric nuclease Fis-OP has been used to identify novel Fis binding sites. Tethering the chemical nuclease OP-Cu+ to position 73 of the protein with a newly developed longer acetyl-beta-alanylamino spacer has facilitated the localization of two high-affinity Fis binding sequences in a 3 kb pUC19 plasmid. The shorter acetamido linker has allowed the chimeric nuclease to locate two strong Fis binding sites in the 50 kb phage lambda genome. All four sites reside in biologically interesting loci and have been confirmed by gel-retardation and DNase I footprint analyses. A newly discovered site resides in the lac operon of Escherichia coli. The binding of Fis to this site may antagonize repression by the LacI repressor. These studies demonstrate the feasibility of applying chimeric chemical nucleases to the task of identifying functional protein binding sites of biological interest within genomes without any assumption about their sequence preference.

Bacteriophage lambda↗