The Experts below are selected from a list of 6840 Experts worldwide ranked by ideXlab platform
Susan Gottesman - One of the best experts on this subject based on the ideXlab platform.
-
Stress sigma factor RpoS degradation and translation are sensitive to the state of central metabolism
Proceedings of the National Academy of Sciences of the United States of America, 2015Co-Authors: Aurelia Battesti, Nadim Majdalani, Susan GottesmanAbstract:RpoS, the stationary phase/stress sigma factor of Escherichia coli, regulates a large cohort of genes important for the cell to deal with suboptimal conditions. Its level increases quickly in the cell in response to many stresses and returns to low levels when growth resumes. Increased RpoS results from increased translation and decreased RpoS degradation. Translation is positively regulated by small RNAs (sRNAs). Protein stability is positively regulated by anti-adaptors, which prevent the RssB adaptor-mediated degradation of RpoS by the ClpXP protease. Inactivation of aceE, a subunit of pyruvate dehydrogenase (PDH), was found to increase levels of RpoS by affecting both translation and protein degradation. The stabilization of RpoS in aceE mutants is dependent on increased transcription and translation of IraP and IraD, two known anti-adaptors. The aceE mutation also leads to a significant increase in RpoS translation. The sRNAs known to positively regulate RpoS are not responsible for the increased translation; sequences around the start codon are sufficient for the induction of translation. PDH synthesizes acetyl-CoA; acetate supplementation allows the cell to synthesize acetyl-CoA by an alternative, less favored pathway, in part dependent upon RpoS. Acetate addition suppressed the effects of the aceE mutant on induction of the anti-adaptors, RpoS stabilization, and RpoS translation. Thus, the bacterial cell responds to lowered levels of acetyl-CoA by inducing RpoS, allowing reprogramming of E. coli metabolism.
-
the RpoS mediated general stress response in escherichia coli
Annual Review of Microbiology, 2011Co-Authors: Aurelia Battesti, Nadim Majdalani, Susan GottesmanAbstract:Under conditions of nutrient deprivation or stress, or as cells enter stationary phase, Escherichia coli and related bacteria increase the accumulation of RpoS, a specialized sigma factor. RpoS-dependent gene expression leads to general stress resistance of cells. During rapid growth, RpoS translation is inhibited and any RpoS protein that is synthesized is rapidly degraded. The complex transition from exponential growth to stationary phase has been partially dissected by analyzing the induction of RpoS after specific stress treatments. Different stress conditions lead to induction of specific sRNAs that stimulate RpoS translation or to induction of small-protein antiadaptors that stabilize the protein. Recent progress has led to a better, but still far from complete, understanding of how stresses lead to RpoS induction and what RpoS-dependent genes help the cell deal with the stress.
-
mechanism of positive regulation by dsra and rpra small noncoding rnas pairing increases translation and protects RpoS mrna from degradation
Journal of Bacteriology, 2010Co-Authors: Colleen A Mccullen, Jihane N Benhammou, Nadim Majdalani, Susan GottesmanAbstract:Small noncoding RNAs (sRNAs) regulate gene expression in Escherichia coli by base pairing with mRNAs and modulating translation and mRNA stability. The sRNAs DsrA and RprA stimulate the translation of the stress response transcription factor RpoS by base pairing with the 5' untranslated region of the RpoS mRNA. In the present study, we found that the RpoS mRNA was unstable in the absence of DsrA and RprA and that expression of these sRNAs increased both the accumulation and the half-life of the RpoS mRNA. Mutations in dsrA, rprA, or RpoS that disrupt the predicted pairing sequences and reduce translation of RpoS also destabilize the RpoS mRNA. We found that the RpoS mRNA accumulates in an RNase E mutant strain in the absence of sRNA expression and, therefore, is degraded by an RNase E-mediated mechanism. DsrA expression is required, however, for maximal translation even when RpoS mRNA is abundant. This suggests that DsrA protects RpoS mRNA from degradation by RNase E and that DsrA has a further activity in stimulating RpoS protein synthesis. RpoS mRNA is subject to degradation by an additional pathway, mediated by RNase III, which, in contrast to the RNase E-mediated pathway, occurs in the presence and absence of DsrA or RprA. RpoS mRNA and RpoS protein levels are increased in an RNase III mutant strain with or without the sRNAs, suggesting that the role of RNase III in this context is to reduce the translation of RpoS even when the sRNAs are acting to stimulate translation.
-
the phop phoq two component system stabilizes the alternative sigma factor RpoS in salmonella enterica
Proceedings of the National Academy of Sciences of the United States of America, 2006Co-Authors: Tammy Latifi, Susan Gottesman, Alexandre Bougdour, Eduardo A GroismanAbstract:The sigma factor RpoS regulates the expression of many stress response genes and is required for virulence in several bacterial species. We now report that RpoS accumulates when Salmonella enterica serovar Typhimurium is growing logarithmically in media with low Mg2+ concentrations. This process requires the two-component regulatory system PhoP/PhoQ, which is specifically activated in low Mg2+. We show that PhoP controls RpoS protein turnover by serving as a transcriptional activator of the iraP (yaiB) gene, which encodes a product that enhances RpoS stability by interacting with RssB, the protein that normally delivers RpoS to the ClpXP protease for degradation. Mutation of the phoP gene rendered Salmonella as sensitive to hydrogen peroxide as an RpoS mutant after growth in low Mg2+. In Escherichia coli, low Mg2+ leads to only modest RpoS stabilization, and iraP is not regulated by PhoP/PhoQ. These findings add the sigma factor RpoS to the regulatory proteins and two-component systems that are elevated in a PhoP/PhoQ-dependent fashion when Salmonella face low Mg2+ environments. Our data also exemplify the critical differences in regulatory circuits that exist between the closely related enteric bacteria Salmonella and E. coli.
-
regulation and mode of action of the second small rna activator of RpoS translation rpra
Molecular Microbiology, 2002Co-Authors: Nadim Majdalani, David Hernandez, Susan GottesmanAbstract:Translation of the stationary phase sigma factor RpoS is stimulated by at least two small RNAs, DsrA and RprA. DsrA disrupts an inhibitory secondary structure in the RpoS leader mRNA by pairing with the upstream RNA. Mutations in rprA and compensating mutations in the RpoS leader demonstrate that RprA interacts with the same region of the RpoS leader as DsrA. This is the first example of two different small RNAs regulating a common target. Regulation of these RNAs differs. DsrA synthesis is increased at low temperature. We find that RprA synthesis is regulated by the RcsC/RcsB phosphorelay system, previously found to regulate capsule synthesis and promoters of ftsZ and osmC. An rcsB null mutation abolishes the basal level, whereas mutations in rcsC that activate capsule synthesis also activate expression of the rprA promoter. An essential site with similarity to other RcsB-regulated promoters was defined in the rprA promoter. Activation of the RcsC/RcsB system leads to increased RpoS synthesis, in an RprA-dependent fashion. This work suggests a new signal for RpoS translation and extends the global regulation effected by the RcsC/RcsB system to coregulation of RpoS with capsule and FtsZ.
Herb E. Schellhorn - One of the best experts on this subject based on the ideXlab platform.
-
phenotypic diversity caused by differential RpoS activity among environmental escherichia coli isolates
Applied and Environmental Microbiology, 2011Co-Authors: Sarah M Chiang, Tao G Dong, Thomas A Edge, Herb E. SchellhornAbstract:Enteric bacteria deposited into the environment by animal hosts are subject to diverse selective pressures. These pressures may act on phenotypic differences in bacterial populations and select adaptive mutations for survival in stress. As a model to study phenotypic diversity in environmental bacteria, we examined mutations of the stress response sigma factor, RpoS, in environmental Escherichia coli isolates. A total of 2,040 isolates from urban beaches and nearby fecal pollution sources on Lake Ontario (Canada) were screened for RpoS function by examining growth on succinate and catalase activity, two RpoS-dependent phenotypes. The RpoS sequence was determined for 45 isolates, including all candidate RpoS mutants, and of these, six isolates were confirmed as mutants with the complete loss of RpoS function. Similarly to laboratory strains, the RpoS expression of these environmental isolates was stationary phase dependent. However, the expression of RpoS regulon members KatE and AppA had differing levels of expression in several environmental isolates compared to those in laboratory strains. Furthermore, after plating RpoS+ isolates on succinate, RpoS mutants could be readily selected from environmental E. coli. Naturally isolated and succinate-selected RpoS mutants had lower generation times on poor carbon sources and lower stress resistance than their RpoS+ isogenic parental strains. These results show that RpoS mutants are present in the environment (with a frequency of 0.003 among isolates) and that, similarly to laboratory and pathogenic strains, growth on poor carbon sources selects for RpoS mutations in environmental E. coli. RpoS selection may be an important determinant of phenotypic diversification and, hence, the survival of E. coli in the environment.
-
role of RpoS in virulence of pathogens
Infection and Immunity, 2010Co-Authors: Tao G Dong, Herb E. SchellhornAbstract:Understanding mechanisms of bacterial pathogenesis is critical for infectious disease control and treatment. Infection is a sophisticated process that requires the participation of global regulators to coordinate expression of not only genes coding for virulence factors but also those involved in other physiological processes, such as stress response and metabolic flux, to adapt to host environments. RpoS is a key response regulator to stress conditions in Escherichia coli and many other proteobacteria. In contrast to its conserved well-understood role in stress response, effects of RpoS on pathogenesis are highly variable and dependent on species. RpoS contributes to virulence through either enhancing survival against host defense systems or directly regulating expression of virulence factors in some pathogens, while RpoS is dispensable, or even inhibitory, to virulence in others. In this review, we focus on the distinct and niche-dependent role of RpoS in virulence by surveying recent findings in many pathogens.
-
global effect of RpoS on gene expression in pathogenic escherichia coli o157 h7 strain edl933
BMC Genomics, 2009Co-Authors: Tao G Dong, Herb E. SchellhornAbstract:RpoS is a conserved stress regulator that plays a critical role in survival under stress conditions in Escherichia coli and other γ-proteobacteria. RpoS is also involved in virulence of many pathogens including Salmonella and Vibrio species. Though well characterized in non-pathogenic E. coli K12 strains, the effect of RpoS on transcriptome expression has not been examined in pathogenic isolates. E. coli O157:H7 is a serious human enteropathogen, possessing a genome 20% larger than that of E. coli K12, and many of the additional genes are required for virulence. The genomic difference may result in substantial changes in RpoS-regulated gene expression. To test this, we compared the transcriptional profile of wild type and RpoS mutants of the E. coli O157:H7 EDL933 type strain. The RpoS mutation had a pronounced effect on gene expression in stationary phase, and more than 1,000 genes were differentially expressed (twofold, P < 0.05). By contrast, we found 11 genes expressed differently in exponential phase. Western blot analysis revealed that, as expected, RpoS level was low in exponential phase and substantially increased in stationary phase. The defect in RpoS resulted in impaired expression of genes responsible for stress response (e.g., gadA, katE and osmY), arginine degradation (astCADBE), putrescine degradation (puuABCD), fatty acid oxidation (fadBA and fadE), and virulence (ler, espI and cesF). For EDL933-specific genes on O-islands, we found 50 genes expressed higher in wild type EDL933 and 49 genes expressed higher in the RpoS mutants. The protein levels of Tir and EspA, two LEE-encoded virulence factors, were elevated in the RpoS mutants under LEE induction conditions. Our results show that RpoS has a profound effect on global gene expression in the pathogenic strain O157:H7 EDL933, and the identified RpoS regulon, including many EDL933-specific genes, differs substantially from that of laboratory K12 strains.
-
Control of RpoS in global gene expression of Escherichia coli in minimal media
Molecular Genetics and Genomics, 2009Co-Authors: Tao Dong, Herb E. SchellhornAbstract:RpoS, an alternative sigma factor, is critical for stress response in Escherichia coli . The RpoS regulon expression has been well characterized in rich media that support fast growth and high growth yields. In contrast, though RpoS levels are high in minimal media, how RpoS functions under such conditions has not been clearly resolved. In this study, we compared the global transcriptional profiles of wild type and an RpoS mutant of E. coli grown in glucose minimal media using microarray analyses. The expression of over 200 genes was altered by loss of RpoS in exponential and stationary phases, with only 48 genes common to both conditions. The nature of the RpoS-controlled regulon in minimal media was substantially different from that expressed in rich media. Specifically, the expression of many genes encoding regulatory factors (e.g., hfq , csrA , and rpoE ) and genes in metabolic pathways (e.g., lysA , lysC , and hisD ) were regulated by RpoS in minimal media. In early exponential phase, protein levels of RpoS in minimal media were much higher than that in Luria-Bertani media, which may at least partly account for the observed difference in the expression of RpoS-controlled genes. Expression of genes required for flagellar function and chemotaxis was elevated in the RpoS mutant. Western blot analyses show that the flagella sigma factor FliA was expressed much higher in RpoS mutants than in WT in all phase of growth. Consistent with this, the motility of RpoS mutants was enhanced relative to WT. In conclusion, RpoS and its controlled regulators form a complex regulatory network that mediates the expression of a large regulon in minimal media.
-
RpoS regulation of gene expression during exponential growth of Escherichia coli K12
Molecular Genetics and Genomics, 2007Co-Authors: Tao Dong, Mark G. Kirchhof, Herb E. SchellhornAbstract:RpoS is a major regulator of genes required for adaptation to stationary phase in E. coli. However, the exponential phase expression of some genes is affected by RpoS mutation, suggesting RpoS may also have an important physiological role in growing cells. To test this hypothesis, we examined the regulatory role of RpoS in exponential phase using both genomic and biochemical approaches. Microarray expression data revealed that, in the RpoS mutant, the expression of 268 genes was attenuated while the expression of 24 genes was enhanced. Genes responsible for carbon source transport (the mal operon for maltose), protein folding (dnaK and mopAB), and iron acquisition (fepBD, entCBA, fecI, and exbBD) were positively controlled by RpoS. The importance of RpoS-mediated control of iron acquisition was confirmed by cellular metal analysis which revealed that the intracellular iron content of wild type cells was two-fold higher than in RpoS mutant cells. Surprisingly, many previously identified RpoS stationary-phase dependent genes were not controlled by RpoS in exponential phase and several genes were RpoS-regulated only in exponential phase, suggesting the involvement of other regulators. The expression of RpoS-dependent genes osmY, tnaA and malK was controlled by Crl, a transcriptional regulator that modulates RpoS activity. In summary, the identification of a group of exponential phase genes controlled by RpoS reveals a novel aspect of RpoS function.
Regine Henggearonis - One of the best experts on this subject based on the ideXlab platform.
-
regulation of RpoS proteolysis in escherichia coli the response regulator rssb is a recognition factor that interacts with the turnover element in RpoS
Proceedings of the National Academy of Sciences of the United States of America, 1999Co-Authors: Gisela Becker, Eberhard Klauck, Regine HenggearonisAbstract:The degradation of the RpoS (σS) subunit of RNA polymerase in Escherichia coli is a prime example of regulated proteolysis in prokaryotes. RpoS turnover depends on ClpXP protease, the response regulator RssB, and a hitherto uncharacterized “turnover element” within RpoS itself. Here we localize the turnover element to a small element (around the crucial amino acid lysine-173) directly downstream of the promoter-recognizing region 2.4 in RpoS. Its sequence as well as its location identify the turnover element as a unique proteolysis-promoting motif. This element is shown to be a site of interaction with RssB. Thus, RssB is functionally unique among response regulators as a direct recognition factor in ClpXP-dependent RpoS proteolysis. Binding of RssB to RpoS is stimulated by phosphorylation of the RssB receiver domain, suggesting that environmental stress affects RpoS proteolysis by modulating RssB affinity for RpoS. Initial evidence indicates that lysine-173 in RpoS, besides being essential of RpoS proteolysis, may play a role in promoter recognition. Thus the same region in RpoS is crucial for proteolysis as well as for activity as a transcription factor.
-
the oxys regulatory rna represses RpoS translation and binds the hfq hf i protein
The EMBO Journal, 1998Co-Authors: Aixia Zhang, Shoshy Altuvia, Anita Tiwari, Liron Argaman, Regine Henggearonis, Gisela StorzAbstract:The OxyS regulatory RNA integrates the adaptive response to hydrogen peroxide with other cellular stress responses and protects against DNA damage. Among the OxyS targets is the RpoS-encoded sigma(s) subunit of RNA polymerase. Sigma(s) is a central regulator of genes induced by osmotic stress, starvation and entry into stationary phase. We examined the mechanism whereby OxyS represses RpoS expression and found that the OxyS RNA inhibits translation of the RpoS message. This repression is dependent on the hfq-encoded RNA-binding protein (also denoted host factor I, HF-I). Co-immunoprecipitation and gel mobility shift experiments revealed that the OxyS RNA binds Hfq, suggesting that OxyS represses RpoS translation by altering Hfq activity.
-
posttranscriptional osmotic regulation of the sigma s subunit of rna polymerase in escherichia coli
Journal of Bacteriology, 1996Co-Authors: A Muffler, Roland Lange, D D Traulsen, Regine HenggearonisAbstract:The sigma(s) subunit of RNA polymerase (encoded by the RpoS gene) is a master regulator in a complex regulatory network that governs the expression of many stationary-phase-induced and osmotically regulated genes in Escherichia coli. RpoS expression is itself osmotically regulated by a mechanism that operates at the posttranscriptional level. Cells growing at high osmolarity already exhibit increased levels of sigma(s) during the exponential phase of growth. Osmotic induction of RpoS can be triggered by addition of NaCl or sucrose and is alleviated by glycine betaine. Stimulation of RpoS translation and a change in the half-life of sigma(s) from 3 to 50 min both contribute to osmotic induction. Experiments with lacZ fusions inserted at different positions within the RpoS gene indicate that an element required for sigma(s) degradation is encoded between nucleotides 379 and 742 of the RpoS coding sequence.
-
identification of transcriptional start sites and the role of ppgpp in the expression of RpoS the structural gene for the sigma s subunit of rna polymerase in escherichia coli
Journal of Bacteriology, 1995Co-Authors: Roland Lange, Daniela Fischer, Regine HenggearonisAbstract:RpoS is the structural gene for the sigma S subunit of RNA polymerase which controls the expression of a large number of genes in Escherichia coli that are induced during entry into stationary phase or in response to increased medium osmolarity. Using a combination of primer extension experiments and a 59 deletion analysis of the region upstream of RpoS, we show that RpoS transcription is mainly driven by a single promoter (RpoSp1) located within the nlpD gene upstream of RpoS (the two relatively weak nlpD promoters contribute to the low level of RpoS expression during early exponential phase). In addition, we demonstrate that the expression of both transcriptional and translational RpoS::lacZ fusions as well as the level of RpoS mRNA originating at RpoSp1 is strongly reduced in ppGpp-deficient relA spoT mutants. However, experiments with the 59 deletion constructs indicate that a lack of ppGpp does affect transcriptional elongation rather than initiation.
-
osmotic regulation of RpoS dependent genes in escherichia coli
Journal of Bacteriology, 1993Co-Authors: Regine Henggearonis, Roland Lange, N Henneberg, Daniela FischerAbstract:The RpoS gene, which encodes a putative alternative sigma factor (sigma S), is essential for the expression of a variety of stationary-phase-induced genes as well as for stationary-phase-specific multiple-stress resistance. As previously shown for the otsA and otsB genes (R. Hengge-Aronis, W. Klein, R. Lange, M. Rimmele, and W. Boos, J. Bacteriol. 173:7918-7924, 1991), we demonstrate here that additional RpoS-controlled genes (bolA, csi-5) as well as at least 18 proteins on two-dimensional O9Farrell gels could be induced in growing cells by osmotic upshift via an RpoS-dependent mechanism. Also, RpoS-dependent thermotolerance and resistance against hydrogen peroxide could be osmotically stimulated. In contrast, the expression of glgS, while exhibiting strong stationary-phase induction, was only weakly increased by elevated osmolarity, and several RpoS-dependent proteins previously identified on two-dimensional gels were not osmotically induced. During osmotic induction of RpoS-dependent genes, RpoS transcription and the level of sigma S remained unchanged. We conclude that osmotically regulated genes represent a subfamily within the RpoS regulon that requires differential regulation in addition to that provided by sigma S. Images
Nadim Majdalani - One of the best experts on this subject based on the ideXlab platform.
-
Stress sigma factor RpoS degradation and translation are sensitive to the state of central metabolism
Proceedings of the National Academy of Sciences of the United States of America, 2015Co-Authors: Aurelia Battesti, Nadim Majdalani, Susan GottesmanAbstract:RpoS, the stationary phase/stress sigma factor of Escherichia coli, regulates a large cohort of genes important for the cell to deal with suboptimal conditions. Its level increases quickly in the cell in response to many stresses and returns to low levels when growth resumes. Increased RpoS results from increased translation and decreased RpoS degradation. Translation is positively regulated by small RNAs (sRNAs). Protein stability is positively regulated by anti-adaptors, which prevent the RssB adaptor-mediated degradation of RpoS by the ClpXP protease. Inactivation of aceE, a subunit of pyruvate dehydrogenase (PDH), was found to increase levels of RpoS by affecting both translation and protein degradation. The stabilization of RpoS in aceE mutants is dependent on increased transcription and translation of IraP and IraD, two known anti-adaptors. The aceE mutation also leads to a significant increase in RpoS translation. The sRNAs known to positively regulate RpoS are not responsible for the increased translation; sequences around the start codon are sufficient for the induction of translation. PDH synthesizes acetyl-CoA; acetate supplementation allows the cell to synthesize acetyl-CoA by an alternative, less favored pathway, in part dependent upon RpoS. Acetate addition suppressed the effects of the aceE mutant on induction of the anti-adaptors, RpoS stabilization, and RpoS translation. Thus, the bacterial cell responds to lowered levels of acetyl-CoA by inducing RpoS, allowing reprogramming of E. coli metabolism.
-
the RpoS mediated general stress response in escherichia coli
Annual Review of Microbiology, 2011Co-Authors: Aurelia Battesti, Nadim Majdalani, Susan GottesmanAbstract:Under conditions of nutrient deprivation or stress, or as cells enter stationary phase, Escherichia coli and related bacteria increase the accumulation of RpoS, a specialized sigma factor. RpoS-dependent gene expression leads to general stress resistance of cells. During rapid growth, RpoS translation is inhibited and any RpoS protein that is synthesized is rapidly degraded. The complex transition from exponential growth to stationary phase has been partially dissected by analyzing the induction of RpoS after specific stress treatments. Different stress conditions lead to induction of specific sRNAs that stimulate RpoS translation or to induction of small-protein antiadaptors that stabilize the protein. Recent progress has led to a better, but still far from complete, understanding of how stresses lead to RpoS induction and what RpoS-dependent genes help the cell deal with the stress.
-
mechanism of positive regulation by dsra and rpra small noncoding rnas pairing increases translation and protects RpoS mrna from degradation
Journal of Bacteriology, 2010Co-Authors: Colleen A Mccullen, Jihane N Benhammou, Nadim Majdalani, Susan GottesmanAbstract:Small noncoding RNAs (sRNAs) regulate gene expression in Escherichia coli by base pairing with mRNAs and modulating translation and mRNA stability. The sRNAs DsrA and RprA stimulate the translation of the stress response transcription factor RpoS by base pairing with the 5' untranslated region of the RpoS mRNA. In the present study, we found that the RpoS mRNA was unstable in the absence of DsrA and RprA and that expression of these sRNAs increased both the accumulation and the half-life of the RpoS mRNA. Mutations in dsrA, rprA, or RpoS that disrupt the predicted pairing sequences and reduce translation of RpoS also destabilize the RpoS mRNA. We found that the RpoS mRNA accumulates in an RNase E mutant strain in the absence of sRNA expression and, therefore, is degraded by an RNase E-mediated mechanism. DsrA expression is required, however, for maximal translation even when RpoS mRNA is abundant. This suggests that DsrA protects RpoS mRNA from degradation by RNase E and that DsrA has a further activity in stimulating RpoS protein synthesis. RpoS mRNA is subject to degradation by an additional pathway, mediated by RNase III, which, in contrast to the RNase E-mediated pathway, occurs in the presence and absence of DsrA or RprA. RpoS mRNA and RpoS protein levels are increased in an RNase III mutant strain with or without the sRNAs, suggesting that the role of RNase III in this context is to reduce the translation of RpoS even when the sRNAs are acting to stimulate translation.
-
regulation and mode of action of the second small rna activator of RpoS translation rpra
Molecular Microbiology, 2002Co-Authors: Nadim Majdalani, David Hernandez, Susan GottesmanAbstract:Translation of the stationary phase sigma factor RpoS is stimulated by at least two small RNAs, DsrA and RprA. DsrA disrupts an inhibitory secondary structure in the RpoS leader mRNA by pairing with the upstream RNA. Mutations in rprA and compensating mutations in the RpoS leader demonstrate that RprA interacts with the same region of the RpoS leader as DsrA. This is the first example of two different small RNAs regulating a common target. Regulation of these RNAs differs. DsrA synthesis is increased at low temperature. We find that RprA synthesis is regulated by the RcsC/RcsB phosphorelay system, previously found to regulate capsule synthesis and promoters of ftsZ and osmC. An rcsB null mutation abolishes the basal level, whereas mutations in rcsC that activate capsule synthesis also activate expression of the rprA promoter. An essential site with similarity to other RcsB-regulated promoters was defined in the rprA promoter. Activation of the RcsC/RcsB system leads to increased RpoS synthesis, in an RprA-dependent fashion. This work suggests a new signal for RpoS translation and extends the global regulation effected by the RcsC/RcsB system to coregulation of RpoS with capsule and FtsZ.
-
regulation of RpoS by a novel small rna the characterization of rpra
Molecular Microbiology, 2001Co-Authors: Nadim Majdalani, Susan Gottesman, Suann Chen, Jonathan R Murrow, Kristin St JohnAbstract:Translational regulation of the stationary phase sigma factor RpoS is mediated by the formation of a double-stranded RNA stem-loop structure in the upstream region of the RpoS messenger RNA, occluding the translation initiation site. The interaction of the RpoS mRNA with a small RNA, DsrA, disrupts the double-strand pairing and allows high levels of translation initiation. We screened a multicopy library of Escherichia coli DNA fragments for novel activators of RpoS translation when DsrA is absent. Clones carrying rprA (RpoS regulator RNA) increased the translation of RpoS. The rprA gene encodes a 106 nucleotide regulatory RNA. As with DsrA, RprA is predicted to form three stem-loops and is highly conserved in Salmonella and Klebsiella species. Thus, at least two small RNAs, DsrA and RprA, participate in the positive regulation of RpoS translation. Unlike DsrA, RprA does not have an extensive region of complementarity to the RpoS leader, leaving its mechanism of action unclear. RprA is non-essential. Mutations in the gene interfere with the induction of RpoS after osmotic shock when DsrA is absent, demonstrating a physiological role for RprA. The existence of two very different small RNA regulators of RpoS translation suggests that such additional regulatory RNAs are likely to exist, both for regulation of RpoS and for regulation of other important cellular components.
Thomas Elliott - One of the best experts on this subject based on the ideXlab platform.
-
dksa affects ppgpp induction of RpoS at a translational level
Journal of Bacteriology, 2002Co-Authors: Larissa Brown, Thomas Elliott, Daniel R Gentry, Michael CashelAbstract:The RpoS sigma factor (also called σS or σ38) is known to regulate at least 50 genes in response to environmental sources of stress or during entry into stationary phase. Regulation of RpoS abundance and activity is complex, with many factors participating at multiple levels. One factor is the nutritional stress signal ppGpp. The absence of ppGpp blocks or delays the induction of RpoS during entry into stationary phase. Artificially inducing ppGpp, without starvation, is known to induce RpoS during the log phase 25- to 50-fold. Induction of ppGpp is found to have only minor effects on RpoS transcript abundance or on RpoS protein stability; instead, the efficiency of RpoS mRNA translation is increased by ppGpp as judged by both RpoS pulse-labeling and promoter-independent effects on lacZ fusions. DksA is found to affect RpoS abundance in a manner related to ppGpp. Deleting dksA blocks RpoS induction by ppGpp. Overproduction of DksA induces RpoS but not ppGpp. Deleting dksA neither alters regulation of ppGpp in response to amino acid starvation nor nullifies the inhibitory effects of ppGpp on stable RNA synthesis. Although this suggests that dksA is epistatic to ppGpp, inducing ppGpp does not induce DksA. A dksA deletion does display a subset of the same multiple-amino-acid requirements found for ppGpp0 mutants, but overproducing DksA does not satisfy ppGpp0 requirements. Sequenced spontaneous extragenic suppressors of dksA polyauxotrophy are frequently the same T563P rpoB allele that suppresses a ppGpp0 phenotype. We propose that DksA functions downstream of ppGpp but indirectly regulates RpoS induction.
-
dsra rna regulates translation of RpoS message by an anti antisense mechanism independent of its action as an antisilencer of transcription
Proceedings of the National Academy of Sciences of the United States of America, 1998Co-Authors: Nadim Majdalani, Thomas Elliott, Darren D. Sledjeski, Christofer Cunning, Susan GottesmanAbstract:DsrA RNA regulates both transcription, by overcoming transcriptional silencing by the nucleoid-associated H-NS protein, and translation, by promoting efficient translation of the stress σ factor, RpoS. These two activities of DsrA can be separated by mutation: the first of three stem-loops of the 85 nucleotide RNA is necessary for RpoS translation but not for anti-H-NS action, while the second stem-loop is essential for antisilencing and less critical for RpoS translation. The third stem-loop, which behaves as a transcription terminator, can be substituted by the trp transcription terminator without loss of either DsrA function. The sequence of the first stem-loop of DsrA is complementary with the upstream leader portion of RpoS messenger RNA, suggesting that pairing of DsrA with the RpoS message might be important for translational regulation. Mutations in the RpoS leader and compensating mutations in DsrA confirm that this predicted pairing is necessary for DsrA stimulation of RpoS translation. We propose that DsrA pairing stimulates RpoS translation by acting as an anti-antisense RNA, freeing the translation initiation region from the cis-acting antisense RNA and allowing increased translation.
-
mutations that increase expression of the RpoS gene and decrease its dependence on hfq function in salmonella typhimurium
Journal of Bacteriology, 1997Co-Authors: L. Brown, Thomas ElliottAbstract:The RpoS transcription factor (also called sigmaS or sigma38) is required for the expression of a number of stationary-phase and osmotically inducible genes in enteric bacteria. RpoS is also a virulence factor for several pathogenic species, including Salmonella typhimurium. The activity of RpoS is regulated in response to many different signals, at the levels of both synthesis and proteolysis. Previous work with RpoS-lac protein fusions has suggested that translation of RpoS requires hfq function. The product of the hfq gene, host factor I (HF-I), is a ribosome-associated, site-specific RNA-binding protein originally characterized for its role in replication of the RNA bacteriophage Qbeta of Escherichia coli. In this study, the role of HF-I was explored by isolating suppressor mutations that map to the region directly upstream of RpoS. These mutations increase RpoS-lac expression in the absence of HF-I and also confer substantial independence from HF-I. DNA sequence analysis of the mutants suggests a model in which the RNA secondary structure near the ribosome binding site of the RpoS mRNA plays an important role in limiting expression in the wild type. Genetic tests of the model confirm its predictions, at least in part. It seems likely that the mutations analyzed here activate a suppression pathway that bypasses the normal HF-I-dependent route of RpoS expression; however, it is also possible that some of them identify a sequence element with an inhibitory function that is directly counteracted by HF-I.
-
Efficient translation of the RpoS sigma factor in Salmonella typhimurium requires host factor I, an RNA-binding protein encoded by the hfq gene.
Journal of Bacteriology, 1996Co-Authors: L. Brown, Thomas ElliottAbstract:The RpoS transcription factor (also called sigma Sor sigma 38) is required for the expression of a number of stationary-phase and osmotically inducible genes in Escherichia coli. RpoS is also a virulence factor for several pathogenic bacteria, including Salmonella typhimurium. The activity of RpoS is regulated in response to several different signals, at the transcriptional and translational levels as well as by proteolysis. Here we report that host factor I (HF-I), the product of the hfq gene, is required for efficient expression of RpoS in S. typhimurium. HF-I is a small, heat-stable, site-specific RNA-binding protein originally characterized for its role in replication of the RNA bacteriophage Q beta of E. coli. Its role in the uninfected bacterial cell has previously been unknown. Assays of Beta-galactosidase in strains with RpoS-lac fusions, Western blot (immunoblot) analysis, and pulse-labeling and immunoprecipitation of both fusion proteins and native RpoS show that an S. typhimurium hfq mutant has a four- to sevenfold reduction in expression of RpoS that is attributable primarily to a defect in translation. These results add a new level of complexity to the regulation of RpoS activity.
