Pe Promoter Research Articles

A comparison of the DNA binding and bending capacities and the oligomeric states of the immunity repressors of heteroimmune coliphages P2 and W

Bacteriophages P2 and WΦ are heteroimmune members of the P2-like family of temperate Escherichia coli phages. Temperate phages can grow lytically or form lysogeny after infection. A transcriptional switch that contains two con-vergent promoters, Pe and Pc, and two repressors regulate what life mode to enter. The immunity repressor C is the first gene of the lysogenic operon, and it blocks the early Pe promoter. In this work, some characteristics of the C proteins of P2 and WΦ are compared. An in vivo genetic analysis shows that WΦ C, like P2 C, has a strong dimerization activity in the absence of its DNA target. Both C proteins recognize two directly repeated sequences, termed half-sites and a strong bending is induced in the respective DNA target upon binding. P2 C is unable to bind to one half-site as opposed to WΦ, but both half-sites are required for repression of WΦ Pe. A reduction from three to two helical turns between the centers of the half-sites in WΦ has no significant effect on the capacity to repress Pe. However, the protein–DNA complexes formed differ, as determined by electrophoretic mobility shift experiments. A difference in spontaneous phage production is observed in isogenic lysogens.

Evolution of immunity and host chromosome integration site of P2-like coliphages.

The amount and distribution of variation in the genomic region containing the genes in the lytic-lysogenic genetic switch and the sequence that determines the integration site into the host chromosome were analyzed for 38 P2-like phages from Escherichia coli. The genetic switch consists of two convergent mutually exclusive promoters, Pe and Pc, and two repressors, C and Cox. The immunity repressor C blocks the early Pe promoter, leading to the establishment of lysogeny. The Cox repressor blocks expression of Pc, allowing lytic growth. Phylogenetic analyses showed that the C and Cox proteins were distributed into seven distinct classes. The phylogenetic relationship differed between the two proteins, and we showed that homologous recombination plays a major role in creating alterations in the genetic switch, leading to new immunity classes. Analyses of the host integration site for these phages resulted in the discovery of a previously unknown site, and there were at least four regular integration sites. Interestingly, we found no case where phages of the same immunity class had different host attachment sites. The evolution of immunity and integration sites is complex, since it involves interactions both between the phages themselves and between phages and hosts, and often, both regulatory proteins and target DNA must change.

Identification of a response regulator gene for catabolite control from a PCB‐degrading β‐proteobacteria, Acidovorax sp. KKS102

Acidovorax sp. (formally Pseudomonas sp.) strain KKS102 carries a bph operon for the degradation of PCB/biphenyl. Transcription from the pE promoter for the bph operon was found to be under catabolite control, i.e. the promoter activity was at a lower level when succinate, fumarate or acetate was added to the culture. Some mutations in the immediate upstream region of the pE promoter resulted in catabolite-insensitive and constitutively low promoter activity, suggesting that a transcriptional activator was involved in catabolite control. A genetic screen for a pE promoter activator identified two tandemly arranged genes, bphP and bphQ, that encoded proteins homologous to the sensor kinases and response regulators, respectively, of two-component regulatory system. In the bphPQ double mutant, pE promoter activity was weak and catabolite-insensitive, and a supply of the bphQ gene alone led to the restoration of the catabolite response. The mechanism of catabolite repression in KKS102 is explained in terms of inhibition of activation by BphQ. The genes highly similar to bphQ were found from several beta-proteobacteria, such as Burkholderia cenocepacia J2315, B. multivorans ATCC17616, B. xenovorans LB400 and Ralstonia solanacearum RS1085.

Sp1 Is a Co-activator with Ets-1, and Net Is an Important Repressor of the Transcription of CTP:Phosphocholine Cytidylyltransferase α

Phosphatidylcholine biosynthesis via the CDP-choline pathway is primarily regulated by CTP:phosphocholine cytidylyltransferase (CT) encoded by the Pcyt1a and Pcyt1b genes. Previously, we identified an Ets-1-binding site located at -49/-47 in the promoter of Pcyt1a as an important transcriptional element involved in basal CTalpha transcription (Sugimoto, H., Sugimoto, S., Tatei, K., Obinata, H., Bakovic, M., Izumi, T., and Vance, D. E. (2003) J. Biol. Chem. 278, 19716-19722). In this study, we determined whether or not there were other important elements and binding proteins for basal CTalpha transcription in the Pcyt1a promoter, and if other Ets family proteins bind to the Ets-1-binding site. The results indicate the formation of a ternary complex with Ets-1 binding at -49/-47 and Sp1 binding at -58/-54 of the Pcyt1a promoter that is important for activating CTalpha transcription. When nuclear extracts of COS-7 cells expressing various Ets family repressors were incubated with DNA probes, binding of Net to the probes was observed. Net dose-dependently depressed the promoter-luciferase activity by 98%, even when co-expressed with Ets-1. RNA interference targeting Net caused an increase of endogenous CTalpha mRNA. After synchronizing the cell cycle in NIH3T3 cells, CTalpha mRNA increased at the S-M phase corresponding to an increase of Ets-1 mRNA and a decrease of Net mRNA. These results indicated that Net is an important endogenous repressor for CTalpha transcription.

Role of the RNA polymerase alpha subunits in CII-dependent activation of the bacteriophage lambda pE promoter: identification of important residues and positioning of the alpha C-terminal domains.

The bacteriophage lambda CII protein stimulates the activity of three phage promoters, p(E), p(I) and p(aQ), upon binding to a site overlapping the -35 element at each promoter. Here we used preparations of RNA polymerase carrying a DNA cleavage reagent attached to specific residues in the C-terminal domain of the RNA polymerase alpha subunit (alphaCTD) to demonstrate that one alphaCTD binds near position -41 at p(E), whilst the other alphaCTD binds further upstream. The alphaCTD bound near position -41 is oriented such that its 261 determinant is in close proximity to sigma(70). The location of alphaCTD in CII-dependent complexes at the p(E) promoter is very similar to that found at many activator-independent promoters, and represents an alternative configuration for alphaCTD at promoters where activators bind sites overlapping the -35 region. We also used an in vivo alanine scan analysis to show that the DNA-binding determinant of alphaCTD is involved in stimulation of the p(E) promoter by CII, and this was confirmed by in vitro transcription assays. We also show that whereas the K271E substitution in alphaCTD results in a drastic decrease in CII-dependent activation of p(E), the p(I) and p(aQ) promoters are less sensitive to this substitution, suggesting that the role of alphaCTD at the three lysogenic promoters may be different.

Characterization of the developmental switch region of bacteriophage P2 Hy dis.

In this work, the DNA sequence of the transcriptional switch that affects the development of the P2 Hy dis bacteriophage was determined. The switch contains two face-to-face-located promoters and two repressors, Cox and C. The locations of the Pc and Pe promoters were determined by primer extension analysis. The P2 Hy dis homolog of the P2 multifunctional Cox protein was shown to be able to substitute for P2 Cox in repression of the P2 Pc promoter, excision of the P2 prophage, and activation of the satellite phage P4 PLL promoter. A directly repeated sequence, flanking the--35 region of the Pe promoter, was found to be important for C repressor binding as well as for repression. The P4 E protein was shown to derepress the developmental switch of P2 Hy dis in a plasmid-based derepression assay.

BphS, a Key Transcriptional Regulator of bph Genes Involved in Polychlorinated Biphenyl/Biphenyl Degradation inPseudomonas sp. KKS102

The bph genes in Pseudomonas sp. KKS102, which are involved in the degradation of polychlorinated biphenyl/biphenyl, are induced in the presence of biphenyl. In this study our goal was to understand the regulatory mechanisms involved in the inducible expression. The bph genes (bphEGF(orf4)A1A2A3BCD(orf1)A4R) constitute an operon, and its expression is strongly dependent on the pE promoter located upstream of the bphE gene. A bphS gene, whose deduced amino acid sequence showed homology with the GntR family transcriptional repressors, was identified at the upstream region of the bphE gene. Disruption of the bphS gene resulted in constitutive expression of bph genes, suggesting that the bphS gene product negatively regulated the pE promoter. The gel retardation and DNase footprinting analyses demonstrated specific binding of BphS to the pE promoter region and identified four BphS binding sites that were located within and immediately downstream of the -10 box of the pE promoter. The four binding sites were functional in repression because their respective elimination resulted in derepression of the pE promoter. The binding of BphS was abolished in the presence of 2-hydroxy-6-oxo-6-phenylhexa-2,4-dienoic acid, an intermediate compound in the biphenyl degradation pathway. We concluded that the negative regulator BphS plays a central role in the regulation of bph gene expression through its action at the pE promoter.

The transcriptional switch of bacteriophage WPhi, a P2-related but heteroimmune coliphage.

Phage WPhi is a member of the nonlambdoid P2 family of temperate phages. The DNA sequence of the whole early-control region and the int and attP region of phage WPhi has been determined. The phage integration site was located at 88.6 min of the Escherichia coli K-12 map, where a 47-nucleotide sequence was found to be identical in the host and phage genomes. The WPhi Int protein belongs to the Int family of site-specific recombinases, and it seems to have the same arm binding recognition sequence as P2 Int, but the core sequence differs. The transcriptional switch contains two face-to-face promoters, Pe and Pc, and two repressors, C and Cox, controlling Pe and Pc, respectively. The early Pe promoter was found to be much stronger than the Pc promoter. Furthermore, the Pe transcript was shown to interfere with Pc transcription. By site-directed mutagenesis, the binding site of the immunity repressor was located to two direct repeats spanning the Pe promoter. A point mutation in one or the other repeat does not affect repression by C, but when it is included in both, C has no effect on the Pe promoter. The Cox repressor efficiently blocks expression from the Pc promoter, but its DNA recognition sequence was not evident. Most members of the P2 family of phages are able to function as helpers for satellite phage P4, which lacks genes encoding structural proteins and packaging and lysis functions. In this work it is shown that P4 E, known to function as an antirepressor by binding to P2 C, also turns the transcriptional switch of WPhi from the lysogenic to the lytic mode. However, in contrast to P2 Cox, WPhi Cox is unable to activate the P4 Pll promoter.

Regulation of replication of lambda phage and lambda plasmid DNAs at low temperature.

It was previously demonstrated that while lysogenic development of bacteriophage lambda in Escherichia coli proceeds normally at low temperature (20-25 degrees C), lytic development is blocked under these conditions owing to the increased stability of the phage CII protein. This effect was proposed to be responsible for the increased stimulation of the pE promoter, which interferes with expression of the replication genes, leading to inhibition of phage DNA synthesis. Here we demonstrate that the burst size of phage lambda cIb2, which is incapable of lysogenic development, increases gradually over the temperature range from 20 to 37 degrees C, while no phage progeny are observed at 20 degrees C. Contrary to previous reports, it is possible to demonstrate that pE promoter activation by CII may be more efficient at lower temperature. Using density-shift experiments, we found that phage DNA replication is completely blocked at 20 degrees C. Phage growth was also inhibited in cells overexpressing cII, which confirms that CII is responsible for inhibition of phage DNA replication. Unexpectedly, we found that replication of plasmids derived from bacteriophage lambda is neither inhibited at 20 degrees C nor in cells overexpressing cII. We propose a model to explanation the differences in replication observed between lambda phage and lambda plasmid DNA at low temperature.

The Escherichia coli RNA polymerase alpha subunit and transcriptional activation by bacteriophage lambda CII protein.

Bacteriophage lambda is not able to lysogenise the Escherichia coli rpoA341 mutant. This mutation causes a single amino acid substitution Lys271Glu in the C-terminal domain of the RNA polymerase alpha subunit (alphaCTD). Our previous studies indicated that the impaired lysogenisation of the rpoA341 host is due to a defect in transcriptional activation by the phage CII protein and suggested a role for alphaCTD in this process. Here we used a series of truncation and point mutants in the rpoA gene placed on a plasmid to investigate the process of transcriptional activation by the cII gene product. Our results indicate that amino-acid residues 265, 268 and 271 in the a subunit may play an important role in the CII-mediated activation of the pE promoter (most probably residue 271) or may be involved in putative interactions between alphaCTD and an UP-like element near pE (most probably residues 265 and 268). Measurement of the activity of pE-lacZ, pI-lacZ and p(aQ)-lacZ fusions in the rpoA+ and rpoA341 hosts demonstrated that the mechanism of activation of these CII-dependent promoters may be in each case different.

The complete nucleotide sequence and functional organization of Bacillus subtilis bacteriophage SPP1.

The complete nucleotide sequence of the B. subtilis bacteriophage SPP1 is described. The genome is 44,007 bp in size and has a base composition of 43.7% dG + dC. Only 32.2 kb are essential for phage amplification under laboratory conditions. Transcription using only the 'heavy strand' is asymmetric. Eighty-one orfs organized in five early and four late operons were identified. Experiments have shown that 25 orfs are essential. Of the remaining orfs, functions could be predicted for the products of five of the orfs on the basis of comparison of the deduced amino acid sequence to known proteins. Intergenic regions include most of the 5 PE and the 4 PL promoters. Transcripts are polycistronic. Transcription from the PE promoters is mediated by host RP, whereas recognition of the PL promoters requires an additional unidentified phage-encoded product. Translation of mRNA transcribed from most of the orfs seems to be initiated independently, each from its own ribosomal binding and initiation site, although a few cases of coupled translation have been reported. The organization of SPP1 genes involved in the replication, DNA packaging and phage assembly proteins resembles the organization of genes of equivalent regions of different E. coli double-stranded DNA phages. Absence of aa sequence similarity between analogous proteins of different phages suggested that the conserved gene organization is representative of a primordial bacteriophage.

Derepression of prophage P2 by satellite phage P4: cloning of the P4 epsilon gene and identification of its product.

Escherichia coli phage P4 lacks all of the genetic information necessary for capsid, tail, and lysis functions. P4 is therefore dependent on a helper phage, such as P2, for lytic propagation. During P4 superinfection of a P2 lysogen, the P2 prophage is derepressed by the action of the P4-encoded epsilon gene. We have cloned the epsilon gene and identified the 10-kDa E protein. The epsilon gene product is the only P4 protein required to derepress prophage P2, which leads to in situ P2 DNA replication. A two-plasmid derepression assay system has been developed to examine the derepression activity of E. The reporter plasmid contains the two face-to-face promoters, Pe and Pc, involved in the lysis-lysogeny transcriptional switch of phage P2 and the immunity repressor C. The Pe promoter is coupled to a cat reporter gene. In the construct, the C repressor is transcribed from the Pc promoter and represses the Pe promoter, which mimics the in situ-repressed P2 prophage. The E protein is supplied in trans from a compatible plasmid in which the epsilon gene is under the control of the T7 promoter. We show here that in the two-plasmid assay system, induction of the E protein derepresses the Pe promoter. The ash9 mutation, which is located upstream of the epsilon gene, enhances the E-mediated derepression of the Pe promoter. The purified E protein shows no specific DNA binding activity, and the implications of this are discussed.

Impaired lysogenisation of the Escherichia coli rpoA341 mutant by bacteriophage lambda is due to the inability of CII to act as a transcriptional activator.

The C-terminus of the alpha subunit of Escherichia coli RNA polymerase is known to function in transcriptional activation at certain promoters. This region was previously shown to be necessary for full activation of the pE promoter by the phage lambda CII protein in vitro. In this work we investigated the inability of phage lambda to follow the lysogenic pathway in cells carrying the point mutation rpoA341 (a change of lysine 271 to glutamic acid). We found that neither overexpression of the cII gene nor stabilisation of the CII protein by the can1 mutation or by cIII gene overexpression was able to suppress the block in lysogenisation. In contrast, the lambda cin1 phage, which carries a CII-independent promoter for the expression of the cI gene, was able to efficiently lysogenise the rpoA341 mutant strain. Furthermore, the rpoA341 mutation prevented the activation of pE-lacZ and pI-lacZ transcriptional fusions by CII. Therefore we conclude that transcriptional activation by the cII gene product is abolished by the rpoA341 mutation, most probably due to impaired interaction between the CII activator and mutant RNA polymerase. The inability of RNA polymerase to respond to CII results in the impairment of lysogenisation of the rpoA341 mutant by phage lambda.

Function of the C-terminal domain of the alpha subunit of Escherichia coli RNA polymerase in basal expression and integration host factor-mediated activation of the early promoter of bacteriophage Mu.

Integration host factor (IHF) can activate transcription from the early promoter (Pe) of bacteriophage Mu both directly and indirectly. Indirect activation occurs through alleviation of H-NS-mediated repression of the Pe promoter (P. Van Ulsen, M. Hillebrand, L. Zulianello, P. Van de Putte, and N. Goosen, Mol. Microbiol. 21:567-578, 1996). The direct activation involves the C-terminal domain of the alpha subunit (alphaCTD) of RNA polymerase. We investigated which residues in the alphaCTD are important for IHF-mediated activation of the Pe promoter. Initial in vivo screening, using a set of substitution mutants derived from an alanine scan (T. Gaal, W. Ross, E. E. Blatter, T. Tang, X. Jia, V. V. Krishnan, N. Assa-Munt, R. Ebright, and R. L. Gourse, Genes Dev. 10:16-26, 1996; H. Tang, K. Severinov, A. Goldfarb, D. Fenyo, B. Chait, and R. H. Ebright, Genes Dev. 8:3058-3067, 1994), indicated that the residues, which are required for transcription activation by the UP element of the rrnB P1 promoter (T. Gaal, W. Ross, E. E. Blatter, T. Tang, X. Jia, V. V. Krishnan, N. Assa-Munt, R. Ebright, and R. L. Gourse, Genes Dev. 10:16-26, 1996), are also important for Pe expression in the presence of IHF. Two of the RNA polymerase mutants, alphaR265A and alphaG296A, that affected Pe expression most in vivo were subsequently tested in in vitro transcription experiments. Mutant RNA polymerase with alphaR265A showed no IHF-mediated activation and a severely reduced basal level of transcription from the Pe promoter. Mutant RNA polymerase with alphaG296A resulted in a slightly reduced transcription from the Pe promoter in the absence of IHF but could still be activated by IHF. These results indicate that interaction of the alphaCTD with DNA is involved not only in the IHF-mediated activation of Pe transcription but also in maintaining the basal level of transcription from this promoter. Mutational analysis of the upstream region of the Pe promoter identified a sequence, positioned from -39 to -51 with respect to the transcription start site, that is important for basal Pe expression, presumably through binding of the alphaCTD. The role of the alphaCTD in IHF-mediated stimulation of transcription from the Pe promoter is discussed.

Drastically decreased transcription from CII-activated promoters is responsible for impaired lysogenization of the Escherichia coli rpoA341 mutant by bacteriophage λ

It was demonstrated previously that a mutation, rpoA341, in the gene encoding the a subunit of Escherichia coli RNA polymerase prevents lysogenization by bacteriophage X. The rpoA341 allele is known to be responsible for impaired transcription of some positively regulated E. coli chromosomal operons. Here we demonstrate that the inhibition of lysogenization of the rpoA341 mutant is a result of drastically decreased transcription from positively regulated phage promoters. We were unable to detect any transcripts originating from the CII-activated PE. pI and paQ promoters (important for lysogenic development) in the phage-infected rpoA341 mutant, in contrast to an otherwise isogenic rpoA + strain. The results are discussed in the light of other reports showing that activation of the pE promoter by CII protein in vitro is decreased only about fivefold when the native a subunit is replaced by truncated a polypeptides.

Drastically decreased transcription from CII-activated promoters is responsible for impaired lysogenization of the Escherichia coli rpoA341 mutant by bacteriophage lambda.

It was demonstrated previously that a mutation, rpoA341, in the gene encoding the alpha subunit of Escherichia coli RNA polymerase prevents lysogenization by bacteriophage lambda. The rpoA341 allele is known to be responsible for impaired transcription of some positively regulated E. coli chromosomal operons. Here we demonstrate that the inhibition of lysogenization of the rpoA341 mutant is a result of drastically decreased transcription from positively regulated phage promoters. We were unable to detect any transcripts originating from the CII-activated pE, pI and paQ promoters (important for lysogenic development) in the phage-infected rpoA341 mutant, in contrast to an otherwise isogenic rpoA+ strain. The results are discussed in the light of other reports showing that activation of the pE promoter by CII protein in vitro is decreased only about fivefold when the native alpha subunit is replaced by truncated alpha polypeptides.

Integration host factor alleviates the H-NS-mediated repression of the early promoter of bacteriophage Mu.

Integration host factor (IHF), which is a histone-like protein, has been shown to positively regulate transcription in two different ways. It can either help the formation of a complex between a transcription factor and RNA polymerase or it can itself activate RNA polymerase without the involvement of other transcription factors. In this study, we present a third mechanism for IHF-stimulated gene expression, by counteracting the repression by another histone-like protein, H-NS. The early (Pe) promoter of bacteriophage Mu is specifically inhibited by H-NS, both in vivo and in vitro. For this inhibition, H-NS binds to a large DNA region overlapping the Pe promoter. Binding of IHF to a binding site just upstream of Pe alleviates the H-NS-mediated repression of transcription. This same ihf site is also involved in the direct activation of Pe by IHF. In contrast to the direct activation by IHF, however, the alleviating effect of IHF appears not to be dependent on the relevant position of the ihf site on the DNA helix, and it also does not require the presence of the C-terminal domain of the alpha subunit of RNA polymerase. Footprint analysis shows that binding of IHF to the ihf site destabilizes the interaction of H-NS with the DNA, not only in the IHF-binding region but also in the DNA regions flanking the ihf site. These results suggest that IHF disrupts a higher-order nucleoprotein complex that is formed by H-NS and the DNA.

Transcription activation by histone-like protein integration host factor

Publisher Summary This chapter describes methods used to study the mechanism of transcription activation by integration host factor (IHF), using the Pe promoter of bacteriophage Mu as an example. IHF is a member of the class of so-called histone-like proteins from Escherichia coli . Proteins belonging to this class are small, relatively abundant, and able to bend or wrap the DNA, and have therefore been implicated in the condensation of the bacterial chromosome. In addition to IHF, other well-characterized histone-like proteins are HU, H-NS, and FIS. IHF and FIS are site-specific DNA-binding proteins, whereas HU and H-NS interact with DNA nonspecifically. For the FIS and IHF proteins, however, it has been shown that they can also have a more direct role—that is, in the activation of transcription by interacting with RNA polymerase. The chapter discusses the detection of IHF-RNA polymerase complexes in solution.

Genetic analysis of the DNA recognition sequence of the P2 Cox protein

The Cox protein of temperate Escherichia coli phage P2 is involved in three important biological processes: (i) excision of the integrated prophage genome (G. Lindahl and M. Sunshine, Virology 49:180-187, 1972), (ii) transcriptional repression of the P2 Pc promoter, which controls the expression of the immunity repressor C and the integrase (S. Saha, E. Haggård-Ljungquist, and K. Nordström, EMBO J. 6:3191-3199, 1987), and (iii) transcriptional activation of the late PII promoter of the unrelated satellite phage P4 (S. Saha, E. Haggård-Ljungquist, and K. Nordström, Proc. Natl. Acad. Sci. USA 86:3973-3977, 1989). A comparison of the DNA regions protected by Cox from DNaseI degradation has revealed a presumptive Cox recognition sequence (Saha et al., Proc. Natl. Acad. Sci. USA). The binding region of Cox in the P2 Pc promoter contains three presumptive recognition sequences, "Cox boxes," located in tandem. P2 vir3 and P2 vir24 are virulent deletion mutants unable to plate on Cox-producing strains, most likely because the deletions locate the new early promoters too close to the Cox-binding region (Saha et al., EMBO J.). In this report, spontaneous P2 vir3 and vir24 mutants, no longer sensitive to repression by the Cox protein, have been isolated. These mutants plate with equal efficiency on strains with or without a Cox-producing plasmid, and they have been named cor for cox resistance. Three types are recognized; the four P2 vir3 cor mutants have a 1-base deletion in the first Cox box, while the P2 vir24 cor mutants were of two types; four have a base substitution in the first Cox box, and one has a base substitution in the second Cox box. The effect of the Cox protein on the mutated P2 vir3 and vir24 promoters was analyzed in vivo by using fusions to a promoterless cat (chloramphenicol acetyltransferase) gene. The activities of the P2 vir3 and vir24 early promoters, as opposed to the wild-type early Pe promoter, are drastically reduced by the Cox protein, and the cor mutation renders them as resistant to Cox as the wild-type Pe promoter. Thus, at least the first two Cox boxes are essential for binding of the Cox protein.

Integration host factor is necessary for lysogenization of Escherichia coli by bacteriophage P2

Whether infection by bacteriophage P2 results in lysogenization of the host or vegetative growth of the phage depends upon a race between transcription from the repressor promoter Pc and the early promoter Pe; transcription from these promoters is mutually exclusive, since the Pc repressor Cox is formed from the Pe transcript and the Pe repressor C from the Pc transcript. The involvement of integration host factor (IHF) in the lysogenization of Escherichia coli K12 by P2 was tested by comparing wild-type and IHF-deficient (himA and himD) mutants. No lysogenic clones were formed following infection of the mutant bacteria. A switch plasmid that contains Pc-C-cat and Pe-cox-kan was used to test the choice for expression of Pc versus Pe. In the wild-type K12 bacteria, 20% of the clones expressed Pe transcription and 80% Pc transcription, whereas all transformed IHF-defective clones expressed transcription from Pe only. The effects of IHF on the in vivo expression of the Pe and Pc promoters were only marginal. The IHF protein was found to bind upstream of the Pe promoter, where a potential ihf sequence is located.