micF RNA in ompB mutants of Escherichia coli: different pathways regulate micF RNA levels in response to osmolarity and temperature change (original) (raw)
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Journal of Bacteriology, 1987
To analyze the function of micF as an antisera RNA in the osmoregulatory expression of the ompF gene in Escherichia coli, we performed two experiments. In the first experiment, two strains were constructed in which the transcription initiation site of the ompF gene and the transcription termination site of the micF gene were separated by 186 and 4,100 base pairs, respectively, on the chromosome. These two strains showed almost the same profile of ompF expression as the wild-type strain in which the two genes are separated by 10(6) base pairs. When a high-copy-number plasmid carrying the micF gene was introduced into these strains, ompF expression was completely repressed, whereas no repression was observed with a low-copy-number plasmid carrying the micF gene. These results indicate that the distance between the two genes on the chromosome is not critical for the function of micF. In the second experiment, expression of the ompF gene was examined by pulse-labeling in both the micF+ ...
MicC, a Second Small-RNA Regulator of Omp Protein Expression in Escherichia coli
Journal of Bacteriology, 2004
In a previous bioinformatics-based search for novel small-RNA genes encoded by the Escherichia coli genome, we identified a region, IS063, located between the ompN and ydbK genes, that encodes an ϳ100nucleotide small-RNA transcript. Here we show that the expression of this small RNA is increased at a low temperature and in minimal medium. Twenty-two nucleotides at the 5 end of this transcript have the potential to form base pairs with the leader sequence of the mRNA encoding the outer membrane protein OmpC. The deletion of IS063 increased the expression of an ompC-luc translational fusion 1.5-to 2-fold, and a 10-fold overexpression of the small RNA led to a 2-to 3-fold repression of the fusion. Deletion and overexpression of the IS063 RNA also resulted in increases and decreases, respectively, in OmpC protein levels. Taken together, these results suggest that IS063 is a regulator of OmpC expression; thus, the small RNA has been renamed MicC. The antisense regulation was further demonstrated by the finding that micC mutations were suppressed by compensatory mutations in the ompC mRNA. MicC was also shown to inhibit ribosome binding to the ompC mRNA leader in vitro and to require the Hfq RNA chaperone for its function. We suggest that the MicF and MicC RNAs act in conjunction with the EnvZ-OmpR two-component system to control the OmpF/OmpC protein ratio in response to a variety of environmental stimuli.
Temperature-Dependent Regulation of the LPXT Gene in Escherichia Coli and Pseudomonas Aeruginosa
2017
Sommario PART I State of the Art 1-Temperature is a critical parameter affecting bacteria growth 1.1-In E. coli the major players of heat and cold shock responses are post-transcriptionally regulated through RNA-based mechanisms 12 1.2-The secondary structure of the TIR affects mRNA translation 1.3-RNA thermometers belong to heterogeneous structural classes 1.4-RNA thermometers and virulence 1.5-Searching new RNATs by in vitro and in vivo approaches 2-The Lipopolysaccharide modification systems 2.1-The bacterial Lipopolysaccharide 2.2-Lipid A modifications 2.3-The inner membrane protein LpxT Aim of the Project Main Results A:-Regulation by temperature of the lpxT gene in Pseudomonas aeruginosa B:-Regulation by temperature of the lpxT gene in Escherichia coli Discussion ! 5 Bibliography PART II Tet-Trap, a Genetic Approach to the Identification of Bacterial RNA Thermometers: Application to Pseudomonas aeruginosa PART III Temperature responsive regulation of Escherichia coli lpxT is controlled by a new RNA thermometer based on an unstable mRNA secondary structure and a sub-optimal Shine-Dalgarno sequence ! 6 ! 7
Antibiotics, 2017
Antibiotic resistant Gram-negative bacteria are a serious threat for public health. The permeation of antibiotics through their outer membrane is largely dependent on porin, changes in which cause reduced drug uptake and efficacy. Escherichia coli produces two major porins, OmpF and OmpC. MicF and MicC are small non-coding RNAs (sRNAs) that modulate the expression of OmpF and OmpC, respectively. In this work, we investigated factors that lead to increased production of MicC. micC promoter region was fused to lacZ, and the reporter plasmid was transformed into E. coli MC4100 and derivative mutants. The response of micC-lacZ to antimicrobials was measured during growth over a 6 h time period. The data showed that the expression of micC was increased in the presence of β-lactam antibiotics and in an rpoE depleted mutant. Interestingly, the same conditions enhanced the activity of an ompN-lacZ fusion, suggesting a dual transcriptional regulation of micC and the quiescent adjacent ompN. Increased levels of OmpN in the presence of sub-inhibitory concentrations of chemicals could not be confirmed by Western blot analysis, except when analyzed in the absence of the sigma factor σ E. We suggest that the MicC sRNA acts together with the σ E envelope stress response pathway to control the OmpC/N levels in response to β-lactam antibiotics.
Journal of bacteriology, 1993
The soxRS regulon is a cornerstone of the adaptive defense systems of Escherichia coli against oxidative stress. Unexpectedly, activation of this regulon also enhances bacterial resistance to multiple antibiotics that seem unrelated to oxygen radicals. We previously correlated this multiple antibiotic resistance with a reduced rate of synthesis of the OmpF outer membrane porin that does not affect the OmpC or OmpA porins. Studies presented here, with operon and gene fusions of ompF to lacZ, show that the soxRS-dependent repression of OmpF is achieved posttranscriptionally. We also show posttranscriptional repression of OmpF mediated by the soxQ1 mutation, which maps to the marA locus. These repressions are dependent on the micF gene, which encodes a small RNA partially complementary to the 5' end of the ompF message. Northern (RNA) blotting experiments show that micF transcription is strongly inducible by the superoxide-generating agent paraquat in a manner that depends complete...
Molecular Microbiology, 1993
The product of the m/cF gene is an endogenous antisense RNA which down-regulates the expression of a major outer membrane protein, OmpF, in E. coli. We report here that two DNA-binding factors compete for the same site in the promoter region of the micF gene: RSBF, a high-affinity redox-sensitive DNAbinding factor that responds to an active oxygen species other than hydrogen peroxide or superoxide anions; and HRBF a heat-resistant DNA-binding factor. Both RSBF and HRBF bind to the same DNA sequence, 5'-TTAAAATCAATAACTTATTCTTAA3-', located upstream of the transcription start site of the m/cF gene. We present evidence that RSBF could be the controlling factor of a novel regulon involved in the response to oxidative stress in E. coli.
Transcriptional Response of Escherichia coli to Temperature Shift
Biotechnology Progress, 2008
Temperature shift is often practiced in the cultivation of Escherichia coli to reduce undesired metabolite formation and to maximize synthesis of correctly folded heterologous protein. As the culture temperature is decreased below the optimal 37°C, growth rate decreases and many physiological changes occur. In this study, we investigated the gene expression dynamics of E. coli on switching its cultivation temperature from 37 to 33 and 28°C using whole genome DNA microarrays. Approximately 9% of the genome altered expression level on temperature shift. Overall, the alteration of transcription upon the downshift of temperature is rapid and globally distributed over a wide range of gene classes. The general trends of transcriptional changes at 28 and 33°C were similar. The largest functional class among the differentially expressed genes was energy metabolism. About 12% of genes in energy metabolism show a decrease in their level of expression, and ∼6% show an increase. Consistent with the decrease in the glucose uptake rate, many genes involved in glycolysis and the PTS sugar transport systems show decreased expression. Genes encoding enzymes related to amino acid biosynthesis and transport also have reduced expression levels. Such decrease in expression probably reflects the reduced growth rate and the accompanying reduction in energy and amino acid demand at lower temperatures. However, nearly all genes encoding enzymes in the TCA cycle have increased expression levels, which may well be compensating the reduction of the activity of TCA cycle enzymes at lower temperatures. Temperature shift also results in shift of the cytochromes from the high affinity cytochrome o system to the low affinity cytochrome d system. There is no evidence that protein processing genes are selectively altered to create favorable conditions for heterologous protein synthesis. Our results indicate that the beneficial effect of temperature shift in many biotechnological processes is likely to be attributed to the general effect of reduced growth and metabolism.
Molecular Microbiology, 1991
OmpC, a major outer-membrane protein, is highly expressed when Salmonella typhi is grown in nutrient broth (NB) of either low (NB + 0% sucrose) or high (NB + 20% sucrose) osmolarity. This contrasts with the expression of Escherichia coti OmpC, which is inhibited in low osmolarity and enhanced in high osmolarity, as has been described previously (van Alphen and Lugtenberg, 1977;. Nevertheless, expression of S. typhi OmpC is dependent on the £. coli OmpR transcriptional activator. These findings suggest differences between the mechanisms of osmoregulation of gene expression in both bacteria, although common effectors appear to be shared.
Changes in Escherichia coli transcriptome during acclimatization at low temperature
Research in microbiology, 2003
Upon cold shock Escherichia coli transiently stops growing and adapts to the new temperature (acclimatization phase). The major physiological effects of cold temperature are a decrease in membrane fluidity and the stabilization of secondary structures of RNA and DNA, which may affect the efficiencies of translation, transcription, and replication. Specific proteins are transiently induced in the acclimatization phase. mRNA stabilization and increased translatability play a major role in this phenomenon. Polynucleotide phosphorylase (PNPase) is one of the cold-induced proteins and is essential for E. coli growth at low temperatures. We investigated the global changes in mRNA abundance during cold adaptation both in wild type E. coli MG1655 and in a PNPase-deficient mutant. We observed a twofold or greater variation in the relative mRNA abundance of 20 genes upon cold shock, notably the cold-inducible subset of csp genes and genes not previously associated with cold shock response, among these, the extracytoplasmic stress response regulators rpoE and rseA, and eight genes with unknown function. Interestingly, we found that PNPase both negatively and positively modulated the transcript abundance of some of these genes, thus suggesting a complex role of PNPase in controlling cold adaptation.