O operon pst de P. aeruginosa codifica um transportador de fosfato de alta afinida-de e também a proteína PhoU que, em conjunto, atuam como repressores da ex-pressão do regulon Pho dessa espécie. A atividade de PhoU está também associada ao metabolismo de polifosfato (poliP), dado que mutantes phoU nulos apresentam um vasto acúmulo do biopolímero. Ensaios de β-galactosidase mostraram uma alteração na expressão dos genes ppk e ppx, envolvidos no metabolismo de poliP, no mutante phoU. Observou-se que na cepa selvagem, a transcrição de ppk e de ppx não responde às limitações de Pi ou de nitrogênio, sendo esses genes altamente expressos em condições normais de crescimento. Além disso, determinou-se que ppk é co-transcrito com o gene hemB, os quais formam, portanto, um operon. O operon pst também foi analisado. Foi identificado por ensaios de northern blot o transcrito do primeiro gene do operon, pstS, que codifica uma proteína periplasmática. Também, foi identificado um promotor imediatamente a montante de phoU, o gene mais distal do operon, que permitiria sua expressão em condições normais do crescimento bacteriano. Por fim, determinou-se por ensaios de EMSA que as duas sequências consenso Pho box presentes no operon pst são completamente funcionais. / The pst operon in P. aeruginosa encodes a high-affinity phosphate transporter and the PhoU protein, which together act as repressors of Pho regulon of this species. The PhoU activity is also related with polyphosphate (polyP) metabolism, since phoU null mutants have a large accumulation of the biopolymer. β-galactosidase assays allowed to confirm a change in the expression of ppk and ppx genes, in-volved in PolyP metabolism, in the phoU mutant. It was also evidenced that in the wild type strain, the ppk and ppx transcription does not respond to Pi or nitrogen starvation, and that these genes are highly expressed under conditions of normal growth. In addition, it was determined that ppk is co-transcribed with hemB, a gene involved in the synthesis of porphyrins, and they constitute therefore an operon. The pst operon was also examined. Was identified by northern blot the transcript of the first gene in the operon, pstS, which encodes a periplasmic protein. Also, a promoter was identified immediately upstream of phoU, the most distal gene in the operon, allowing its expression in normal conditions of bacterial growth. Finally, it was determined by EMSA that the two consensus sequences Pho box present in the pst operon are fully functional.

<p>In recent years, the pervasiveness and importance of post-transcriptional regulation has reshaped the underlying principles of the organizational logic of gene expression. RNA-binding proteins (RBPs) and non-coding RNAs are the regulatory molecules primary responsible for interaction with target mRNAs and thereby regulating post-transcriptional processes eventually influencing characteristics of the encoded protein. Many of the mRNA targets of RBPs encode functionally related proteins, which for post-trascriptional operons, resulting in coordination of macromolecular complexes or specific cellular processes. Thus, identifying RNA targets, precise binding sites, and the dynamics of these interactions will reveal how these important regulatory factors contribute to gene regulatory networks.</p><p>ELAV family of human RBPs consist of 4 members, which all have 3 RRM (RNA-recognition motif) domains the last separated by a hinge region. It predominant role is to positively regulate the stability and translation of target mRNAs through binding to ARE (AU-rich elements) in the 3' UTR (untranslated region) of protein coding transcripts. In response to certain stimuli, HuR is subject to post-translational modifications and changes subcellular localization, which impacts its regulatory capacity. In this study on a transcriptome-wide level, we interrogate the RNA targets, precise binding sites, as well as the remodeling of these interactions in response to stimuli.</p><p>We utilized two complementary methods, RIP-chip and PAR-CLIP, to identify targets of HuR and high-resolution binding sites on a transcriptome-wide scale. We discovered that HuR-mRNA interactions are not restricted to the 3' UTR and there are thousands of intronic binding sites. A significant proportion of intronic binding sites are contained in the poly-pyrimidine tract near 3' splice sites. Binding sites in the 3' UTR and intron are often approximately 30 nucleotides apart. HuR can bind to both AU-rich and U-rich sequences, the former more prevalent in 3' UTRs and the latter more prevalent at the 3' splice site.</p><p>Next we integrated the binding data with transcriptomics of HuR siRNA mediated knockdown. We found that the degree of binding is proportional to the degree of HuR-dependent stabilization. Moreover the ability to stabilize mRNA is not restricted to 3' UTR binding sites, as intronic binding sites also exhibited the binding degree correlated stabilization. We observed that the spatial pattern of HuR binding sites relative to exons influences exon usage decisions. Specifically, binding sites upstream of the exon promote exclusion, while binding sites downstream of the exon promote inclusion.</p> / Dissertation

Hyaluronic acid (HA) is a high value biopolymer that has numerous biomedical and cosmetic applications. It is currently derived from two sources, namely animal tissues and bacterial fermentation (Fong Chong et al. 2005). The molecular weight (Mw) of HA can vary from several hundred thousand dalton (Da) to approximately 8 MDa (Widner et al. 2005). High Mw HA has surgical applications, and therefore constitutes a major component of the lucrative HA market. The current need is largely met by extraction from animal tissues, e.g., rooster comb and bovine vitreous humor (Shiedlin et al. 2004). However, the potential of contamination with adventitious agents (e.g., viruses) have raised regulatory concerns regarding the use of animal extracts in pharmaceutical products. Moreover, with recent reports of zoonotic diseases (e.g., bovine spongiform encephalitis and avian influenza virus), pharmaceutical companies are moving towards microbial HA sources. Although HA obtained from bacterial fermentation does not have the problem of viral contamination, this approach has not yet resulted in a process where HA of sufficiently high Mw for surgical applications can be derived. While attempts have been made to produce higher Mw HA through cross-linking, cross-linked HA is undesirable for certain medical procedures (e.g., ophthalmic applications) which requires a natural polymer with a short half-life. Nevertheless, due to its availability and the relative ease of purification, bacterial fermentation has the potential of replacing extraction from animal tissues as a preferred commercial source of HA. This thesis presents a good example of a metabolic engineering study where modern techniques (e.g., molecular biology, fermentation and omics technologies) are used to explain complex cell metabolism. The hypothesis for this study was that the precursors to HA, i.e., UDP-glucuronic acid and UDP-N-acetylglucosamine and consequently the genes involved in precursor generation, are important for HA Mw. However, environmental manipulation, e.g., anaerobic versus aerobic or glucose versus maltose, often results in large global changes in metabolite concentration and enzyme activities. This makes it impossible to resolve issues related to Mw control. Classical statistical methods do not provide a meaningful inference as the number of explanatory variables always exceeds the number of independent observations. Hence, it is difficult to distinguish between causative and accidental correlation. This work first examined the influence of manipulation of metabolite concentrations in the hyaluronan pathway to find an explanation for the mechanism of Mw control. To achieve this, the five essential genes of the hyaluronan synthesis (has) operon in Streptococcus equi subsp. zooepidemicus (S. zooepidemicus) were first examined. These genes are involved in two pathways which lead to the production of either UDP-glucuronic acid or UDP-N-acetylglucosamine. Overexpression of genes involved in UDP-glucuronic acid biosynthesis decreased HA Mw, while overexpression of genes involved in UDP-N-acetylglucosamine biosynthesis increased HA Mw. The Mw variation generated provided a stepping stone for further understanding of Mw control of HA. The highest Mw observed was achieved with combined overexpression of pgi and glmU. This study proved that there is a positive correlation between UDP-N-acetylglucosamine and Mw. The first model for HA Mw control based on the concentration of activated sugar precursors is described in this study (Chapter 3). This correlation observed led to the hypothesis that high Mw HA can be achieved when an appropriate balance of the two HA precursor is maintained. Three genes in the two precursor pathways are not found in the has operon of S. zooepidemicus. To obtain a complete overview of all genes in the HA pathway, these genes were also examined using overexpression studies. Individual overexpression of these genes had negligible effects on HA Mw and production. Despite the positive correlation previously observed between UDP-N-acetylglucosamine and Mw, sequential overexpression of genes involved in the UDP-N-acetylglucosamine precursor pathway did not increase Mw of HA produced. This is surprising since the highest pool of UDP-N-acetylglucosamine was achieved in this case. This suggests that a threshold effect is present in the correlation between UDP-N-acetylglucosamine and Mw. This threshold effect may be defined by a balance between the two precursors. To investigate this phenomenon further, the precursor ratio was also manipulated by co-metabolising glucose and N-acetylglucosamine. Similar to the previous experiment, a significant increase in UDP-N-acetylglucosamine levels was observed despite only a marginal increase in Mw (Chapter 4). Surprisingly, an increase in Mw was observed with the introduction of a plasmid in S. zooepidemicus. This plasmid effect was studied on a global scale using transcriptome and proteome analysis to understand the changes occurring in the system. The increase in Mw due to the plasmid effect is independent of the functions, i.e., nisin promoter or antibiotic resistance, encoded in the plasmid. A gene involved in UDP-N-acetylglucosamine production, UDP-N-acetylglucosamide 1-carboxivinyltransferase (murA), was significantly down-regulated in both the plasmid bearing strain and the high Mw strain (pgi). In addition, overexpression of murA decreased both the concentration of activated sugar precursors and HA Mw. There was however no evidence of down-regulation of murA in the plasmid containing strain from transcriptomics data. This suggests that control is exerted either at the translation level or by protein degradation (Chapter 5). This thesis contributes and represents an ongoing effort to understand the elusive mechanism of Mw control of HA.

Thesis (Ph. D.)--University of Washington, 1999. / Vita. Includes bibliographical references (leaves 145-155).

Universiẗat, Diss., 2005--Düsseldorf.

O operon pst de P. aeruginosa codifica um transportador de fosfato de alta afinida-de e também a proteína PhoU que, em conjunto, atuam como repressores da ex-pressão do regulon Pho dessa espécie. A atividade de PhoU está também associada ao metabolismo de polifosfato (poliP), dado que mutantes phoU nulos apresentam um vasto acúmulo do biopolímero. Ensaios de &beta;-galactosidase mostraram uma alteração na expressão dos genes ppk e ppx, envolvidos no metabolismo de poliP, no mutante phoU. Observou-se que na cepa selvagem, a transcrição de ppk e de ppx não responde às limitações de Pi ou de nitrogênio, sendo esses genes altamente expressos em condições normais de crescimento. Além disso, determinou-se que ppk é co-transcrito com o gene hemB, os quais formam, portanto, um operon. O operon pst também foi analisado. Foi identificado por ensaios de northern blot o transcrito do primeiro gene do operon, pstS, que codifica uma proteína periplasmática. Também, foi identificado um promotor imediatamente a montante de phoU, o gene mais distal do operon, que permitiria sua expressão em condições normais do crescimento bacteriano. Por fim, determinou-se por ensaios de EMSA que as duas sequências consenso Pho box presentes no operon pst são completamente funcionais. / The pst operon in P. aeruginosa encodes a high-affinity phosphate transporter and the PhoU protein, which together act as repressors of Pho regulon of this species. The PhoU activity is also related with polyphosphate (polyP) metabolism, since phoU null mutants have a large accumulation of the biopolymer. &beta;-galactosidase assays allowed to confirm a change in the expression of ppk and ppx genes, in-volved in PolyP metabolism, in the phoU mutant. It was also evidenced that in the wild type strain, the ppk and ppx transcription does not respond to Pi or nitrogen starvation, and that these genes are highly expressed under conditions of normal growth. In addition, it was determined that ppk is co-transcribed with hemB, a gene involved in the synthesis of porphyrins, and they constitute therefore an operon. The pst operon was also examined. Was identified by northern blot the transcript of the first gene in the operon, pstS, which encodes a periplasmic protein. Also, a promoter was identified immediately upstream of phoU, the most distal gene in the operon, allowing its expression in normal conditions of bacterial growth. Finally, it was determined by EMSA that the two consensus sequences Pho box present in the pst operon are fully functional.

The bacteroides group is a subdivision in the <I>Cytophaga-Flavobacterium-Bacteroides</I> phylum. This group is as phylogenetically distinct from other Gram-negative enterics, including <I>Escherichia coli</I>, as they are from Gram-positive organisms. Furthermore, there is no cross expression between genes of <I>E. coli</I> and <I>Bacteroides</I> species. It is thought that this difference in gene expression lies in part at the level of transcription initiation and is due to the sequences within the promoter region itself. A putative consensus sequence for <I>Bacteroides</I> promoters has been published by C. Jeff Smith&#146;s research group based on alignments of the sequences upstream of certain regulated genes. However, this consensus has not been found within all putative <I>Bacteroides</I> promoters. In this study, the promoter structure and function of a strong housekeeping <I>B. thetaiotaomicron</I> 16S rRNA promoter was examined and compared to an <I>E. coli</I> 16S rRNA promoter. Our hypothesis is that there are significant differences between the promoters of these two organisms. Analysis of <I>B. thetaiotaomicron</I> sequence upstream of the 16S rRNA gene has revealed the same overall structure known for <I>E. coli</I> 16S rRNA promoters in that there are two putative promoters separated by approximately 150 bp. However, the <I>B. thetaiotaomicron</I> 16S rRNA promoter contains the proposed <I>Bacteroides</I> &#151;7 and &#151;33 consensus sequences instead of the well known <I>E. coli</I> &#151;10 and &#151;35 consensus sequences. The biological activity of the<I> B. thetaiotaomicron</I> 16S rRNA full-length promoter was confirmed using a <I>Bacteroides lux</I> reporter system. A newly designed <I>Bacteroides lux</I> reporter was used to analyze specific regions of the <I>B. thetaiotaomicron</I> 16S rRNA promoter. In addition, by pairing the <I>B. thetaiotaomicron</I> 16S rRNA promoter with an <I>E. coli</I> ribosomal binding site, and vice-versa, the improved <I>lux</I> reporter was used to further confirm that the difference in gene expression between the two species lies at the level of transcription in <I>E. coli</I>. In <I>Bacteroides</I>, however, transcription and translation may work together to create a barrier to efficient gene expression of foreign genes. </P> / Master of Science

Arsenic contamination of groundwater remains a serious health concern in many areas of the world. Developing countries such as Bangladesh and Nepal are particularly affected because access to high quality water infrastructure is low. Since the 1970s, most water in these countries is sourced from shallow tube wells installed to reduce the spread of diseases associated with poor water hygiene. In this goal they were successful, however by the mid 1990s it became apparent that many of these wells were contaminated by arsenic and that these countries’ rural poor were being slowly poisoned. No simple, cheap, and reliable test for arsenic exists, and efforts to mitigate arsenic contamination have been severely limited by this over the past two decades. Government backed well-testing efforts using commercially available field kits have many issues with reliability, safety, rigour, and transparency, and have lost their urgency over the past decade, while the expensive field test kits remain out of the reach of most ordinary people in these areas. Synthetic Biology offers the technology to develop a new class of biosensor by exploiting bacteria’s natural ability to sense and respond to levels of arsenic considerably lower than commercially available kits which are based on analytical chemistry. In order to reach this goal, we must first develop our understanding of the natural response to arsenic in our chosen host, B. subtilis. Although we have a reasonably good qualitative understanding of the operon responsible for arsenic sensing, very little quantitative analysis has been carried out, and a robust system for ratiometric characterisation has not been established in the bacteria. In this work, a robust platform for rapid ratiometric characterisation is established in B. subtilis. A rigorous mathematical model of the ars operon is developed and analysed before being verified experimentally. This new knowledge is then used to explore synthetic permutations to the natural system aimed at improving the sensor properties of the system. Finally, a biological architecture for an easily tunable biosensor with good characteristics is recommended.

Submitted by Gabriela Lopes (gmachadolopesufpel@gmail.com) on 2017-04-24T11:45:18Z No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Dissertação Tassiana Ramires.pdf: 1728606 bytes, checksum: a63d0903f11cb3f858c5a7fd4e32ad97 (MD5) / Approved for entry into archive by Aline Batista (alinehb.ufpel@gmail.com) on 2017-05-04T18:58:35Z (GMT) No. of bitstreams: 2 Dissertação Tassiana Ramires.pdf: 1728606 bytes, checksum: a63d0903f11cb3f858c5a7fd4e32ad97 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Made available in DSpace on 2017-05-04T18:58:35Z (GMT). No. of bitstreams: 2 Dissertação Tassiana Ramires.pdf: 1728606 bytes, checksum: a63d0903f11cb3f858c5a7fd4e32ad97 (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2017-02-20 / Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPq / Campylobacter termofílicos são, atualmente, as principais bactérias causadoras de doenças gastrointestinais em todo o mundo. Esse grupo é assim denominado devido a sua temperatura ótima de multiplicação oscilar entre 42 °C e 43 °C, sendo Campylobacter jejuni, C. coli, C. lari e C. upsaliensis, as principais espécies envolvidas nos casos de campilobacteriose em humanos. Dentre essas espécies, a mais relacionada à essa doença é C. jejuni, seguida por C. coli. O principal reservatório desses micro-organismos são as aves, principalmente os frangos, possivelmente pela temperatura corporal desses animais ser similar à temperatura ótima para Campylobacter termofílicos. Com isso, o objetivo desse estudo foi avaliar a ocorrência, a diversidade genética, o perfil de resistência a antimicrobianos e a presença de genes associados à virulência em isolados de Campylobacter termofílicos provenientes de frangos de corte e na cama de aviário em granjas aviárias da região sul do Rio Grande do Sul, Brasil. Um total de 48 amostras foram coletadas em três diferentes granjas (A, B e C), incluindo uma amostra de swab de arrasto da cama do aviário e 15 pools de amostras de swab de cloaca em cada granja. Das três granjas amostradas, apenas a granja C apresentou contaminação por Campylobacter termofílicos, sendo todos os isolados identificados por técnicas fenotípicas e moleculares como C. jejuni. Dessas 16 amostras positivas, obtiveram-se 28 isolados, sendo 16 pelo isolamento em ágar Preston e 12 do ágar mCCD. A diversidade genética entre os isolados foi avaliada por PFGE, verificando-se que todos os isolados apresentaram um único padrão de macrorestrição, sugerindo clonalidade entre os isolados e a presença de apenas uma fonte de infecção por Campylobacter nessa granja. O perfil de resistência a antimicrobianos foi avaliado pelo teste de disco difusão em ágar, utilizando-se oito antimicrobianos distintos, de três classes diferentes: tetraciclina, quinolonas e macrolídeos. Os isolados apresentaram perfil similar de resistência a antimicrobianos, sendo resistentes às quinolonas e tetraciclinas e sensíveis aos macrolídeos. Devido a relação clonal e ao perfil de resistência similar, um isolado representativo foi selecionado para detecção dos genes de virulência. A técnica de PCR foi utilizada para detectar a presença dos genes ciaB, cadF, cdtA, cdtB e cdtC, sendo o isolado selecionado positivo todos os genes pesquisados. Dessa forma, a presença de C. jejuni resistente a antimicrobianos e com potencial de virulência em frangos de corte prontos para o abate e na cama de aviário durante o período de produção é um risco à saúde pública, pois esses micro-organismos podem ser introduzidos no ambiente do abatedouro e contaminar as carcaças durante o abate. / Thermophilic Campylobacter are currently the leading bacteria causing of gastrointestinal diseases worldwide. This group is so named because its optimal multiplication temperature oscillates between 42 °C and 43 °C, being Campylobacter jejuni, C. coli, C. lari and C. upsaliensis, the main species involved in cases of human campylobacteriosis. Among these species, the most related to this disease is C. jejuni, followed by C. coli. The main reservoir of these microorganisms are birds, especially chickens, possibly because the body temperature of these animals coincides with the optimal temperature for thermophilic Campylobacter. Therefore, the aim of this study was to verify the occurrence, genetic relationship, antimicrobial susceptibility, and the presence of virulence genes in thermophilic Campylobacter from broilers and broiler bedding from the southern region of Rio Grande do Sul, Brazil. A total of 48 samples were collected in three different farms (A, B and C), which comprising one sample of drag swab and 15 pools of cloacal swabs in each farm. From the three farms sampled, only the farm C showed thermophilic Campylobacter contamination. All isolates were identified by phenotypic and molecular techniques such as C. jejuni. Of these 16 positive samples, 28 isolates were obtained, 16 being isolated by Preston agar and 12 by mCCD agar. The genetic diversity among the isolates was evaluated by PFGE, and it was observed that all the isolates belonged to the same macrorestriction pattern, suggesting clonality among the isolates and the presence of only one source of Campylobacter infection in this farm. The antimicrobial resistance profile was evaluated by the agar disc diffusion test using eight distinct antimicrobial agents from three different classes: tetracyclines, quinolones and macrolides. The isolates presented a similar antimicrobial resistance profile, being resistant to quinolones and tetracyclines and susceptible to macrolides. As the isolates shared the same PFGE pattern and similar resistance profile, a representative isolate was chosed for investigation of virulence genes. A PCR assay was carried out aiming to identify the presence of ciaB, cadF, cdtA, cdtB and cdtC virulence genes and all the genes evaluated were found. Thus, the presence of C. jejuni resistant to antimicrobial agents and harboring virulence genes in broilers and broiler farm during the broiler production period may represents a potential risk to public health, because these microorganisms can be introduced into the abattoir environment and may contaminate the carcasses during slaughter.

The genetic systems involved in the utilisation of aromatic β-glucosides in E. coli consist of the bgl, asc, and chb operons and the locus bglA encoding phospho-β-glucosidase A. The bgl and asc operons are known as cryptic or silent systems since their expression is not sufficient for utilisation of these sugars in wild type strains of E. coli. Their transcriptional activation by different classes of mutations confers a Bgl+ phenotype to the mutant. The maintenance of cryptic genes without accumulating deleterious mutation in spite of being silent is an evolutionary puzzle. Several observations have suggested the possibility that these genes may be expressed under specific physiological conditions conferring a fitness advantage to the organism. The main aim of this study was to investigate the possible role of aromatic β-glucoside catabolic systems of E. coli in combating nutrient stress and microaerobic growth conditions. The results presented in Chapter 2 address the evolution of aromatic β-glucoside catabolic systems when exposed to a novel β-glucoside as the sole substrate. The results indicate that the bgl opeon, the primary system involved in the utilisation of the aromatic β-glucosides arbutin and salicin, is also involved in esculin utilisation. In the absence of bglB encoding the enzyme phospho-β-glucosidase B, activation of the silent asc operon enables esculin utilisation. The bglA gene encoding phospho-β-glucosidase A specific for arbutin, can undergo successive mutations to evolve the ability to hydrolyse esculin and salicin sequentially when bglB and ascB are absent. The Esc+ and Sal+ mutants retain their arbutin+ phenotype, indicating that the mutations enhance the promiscuity of the enzyme. Sequencing data indicate that the first step Esc+ mutant carries a four base insertion within the promoter of the bglA gene that results in enhanced transcription of bglA. RT-PCR studies confirm that both the steady-state levels as well as the half-life of the bglA mRNA are enhanced in the mutant. This is further corroborated by the observation that overexpression of wild type bglA in the parent strain using a multicopy plasmid confers an Esc+ phenotype. The second step Sal+ mutant carries a point mutation within bglA ORF, a thymine to guanine transversion at position 583 (T583G) of the bglA gene, resulting in an amino acid change from cysteine to glycine at position 195 (C195G) of the BglA ORF close to the active site. Presence of a plasmid carrying the T583G mutation, introduced by site-directed mutagenesis, results in a Sal+ phenotype, confirming the role of the transversion in conferring the Sal+ phenotype. Based on docking studies, the positioning of salicin into the substrate binding site of the mutant BglA enzyme is different compared to wild type BglA due to the loss of stearic hindrance for the binding of salicin when C195 is replaced by the smaller amino acid glycine in the mutant protein. These observations indicate that under conditions of nutrient deprivation, exposure to novel substrates can result in the evolution of new metabolic capabilities by the sequential modification of a pre-existing genetic system. In the case of one novel substrate, the mutation results in the overexpression of the hydrolytic enzyme, while in the case of the second substrate, a mutation close to its active site increases its substrate specificity. Results presented in Chapter 3 specifically deal with the involvement of the bgl operon under low levels of oxygen. Earlier observations have shown that there is a 22 fold enhancement in the expression of the bgl operon under anaerobic condition. The present results provide evidence that bgl expression has a physiological role under low levels of oxygen and in addition suggest a possible mechanism for the overexpression of the bgl operon that involves the ArcAB two component system known to mediate regulation under microaerobic and static conditions. Transcription studies using a lacZ reporter fused to the wild type bgl promoter show that there is enhanced transcription from the bgl promoter under microaerobic and static conditions in the presence of arcA encoding the response regulator compared to that in its absence. The positive effect of arcA on the expression of the bgl operon is dispensable in the absence of H-NS since presence or absence of arcA does not change the expression of the bgl operon in an hns-null background, implying that the involvement of ArcA is via antagonizing H-NS. Competition experiments indicate that there is growth advantage associated with the activated allele of the bgl operon under low levels of oxygen since Bgl+ strains carrying the activated allele of the bgl operon as well as strains expressing BglG constitutively can out-compete wild-type strains. Presence of the wild type arcA allele results in a strong growth advantage compared to its absence under static conditions but not aerobic condition. The bgl operon seems to be one of the possible downstream targets of ArcA under static condition since absence of the bgl operon results in a modest reduction of the growth advantage (GASP) phenotype conferred by arcA. The up-regulation of the bgl operon is likely to enable the cells to scavenge available nutrients from their niche more efficiently. These experiments also show that the GASP phenotype associated with BglG constitutive strains under static conditions involves downstream genes that are different from oppA known to be one of the downstream targets during aerobic growth. It is possible that under low level of oxygen, the bgl operon is regulating a different set of downstream genes involving a different mechanism. In summary, the results of this investigation show that the aromatic β-glucoside catabolic systems in E. coli play a role in the generation of new metabolic capabilities via mutations in pre-existing genetic systems as well as through changes in gene expression patterns. The mechanisms outlined in this study are likely to be of broader significance applicable to microbial evolution under stress in general.