TraR, a Homolog of a RNAP Secondary Channel Interactor, Modulates Transcription

Recent structural and biochemical studies have identified a novel control mechanism of gene expression mediated through the secondary channel of RNA Polymerase (RNAP) during transcription initiation. Specifically, the small nucleotide ppGpp, along with DksA, a RNAP secondary channel interacting factor, modifies the kinetics of transcription initiation, resulting in, among other events, down-regulation of ribosomal RNA synthesis and up-regulation of several amino acid biosynthetic and transport genes during nutritional stress. Until now, this mode of regulation of RNAP was primarily associated with ppGpp. Here, we identify TraR, a DksA homolog that mimics ppGpp/DksA effects on RNAP. First, expression of TraR compensates for dksA transcriptional repression and activation activities in vivo. Second, mutagenesis of a conserved amino acid of TraR known to be critical for DksA function abolishes its activity, implying both structural and functional similarity to DksA. Third, unlike DksA, TraR does not require ppGpp for repression of the rrnB P1 promoter in vivo and in vitro or activation of amino acid biosynthesis/transport genes in vivo. Implications for DksA/ppGpp mechanism and roles of TraR in horizontal gene transfer and virulence are discussed

Stringent response of Escherichia coli: revisiting the bibliome using literature mining

Understanding the mechanisms responsible for cellular responses depends on the systematic collection and analysis of information on the main biological concepts involved. Indeed, the identification of biologically relevant concepts in free text, namely genes, tRNAs, mRNAs, gene products and small molecules, is crucial to capture the structure and functioning of different responses. Results In this work, we review literature reports on the study of the stringent response in Escherichia coli. Rather than undertaking the development of a highly specialised literature mining approach, we investigate the suitability of concept recognition and statistical analysis of concept occurrence as means to highlight the concepts that are most likely to be biologically engaged during this response. The co-occurrence analysis of core concepts in this stringent response, i.e. the (p)ppGpp nucleotides with gene products was also inspected and suggest that besides the enzymes RelA and SpoT that control the basal levels of (p)ppGpp nucleotides, many other proteins have a key role in this response. Functional enrichment analysis revealed that basic cellular processes such as metabolism, transcriptional and translational regulation are central, but other stress-associated responses might be elicited during the stringent response. In addition, the identification of less annotated concepts revealed that some (p)ppGpp-induced functional activities are still overlooked in most reviews. Conclusions In this paper we applied a literature mining approach that offers a more comprehensive analysis of the stringent response in E. coli. The compilation of relevant biological entities to this stress response and the assessment of their functional roles provided a more systematic understanding of this cellular response. Overlooked regulatory entities, such as transcriptional regulators, were found to play a role in this stress response. Moreover, the involvement of other stress-associated concepts demonstrates the complexity of this cellular response

Transcription regulation of the Escherichia coli pcnB gene coding for poly(A) polymerase I: roles of ppGpp, DksA and sigma factors

Poly(A) polymerase I (PAP I), encoded by the pcnB gene, is a major enzyme responsible for RNA polyadenylation in Escherichia coli, a process involved in the global control of gene expression in this bacterium through influencing the rate of transcript degradation. Recent studies have suggested a complicated regulation of pcnB expression, including a complex promoter region, a control at the level of translation initiation and dependence on bacterial growth rate. In this report, studies on transcription regulation of the pcnB gene are described. Results of in vivo and in vitro experiments indicated that (a) there are three σ70-dependent (p1, pB, and p2) and two σS-dependent (pS1 and pS2) promoters of the pcnB gene, (b) guanosine tetraphosphate (ppGpp) and DksA directly inhibit transcription from pB, pS1 and pS2, and (c) pB activity is drastically impaired at the stationary phase of growth. These results indicate that regulation of the pcnB gene transcription is a complex process, which involves several factors acting to ensure precise control of PAP I production. Moreover, inhibition of activities of pS1 and pS2 by ppGpp and DksA suggests that regulation of transcription from promoters requiring alternative σ factors by these effectors of the stringent response might occur according to both passive and active models

Insights into transcriptional regulation and σ competition from an equilibrium model of RNA polymerase binding to DNA

To explore scenarios that permit transcription regulation by activator recruitment of RNA polymerase and σ competition in vivo, we used an equilibrium model of RNA polymerase binding to DNA constrained by the values of total RNA polymerase (E) and σ(70) per cell measured in this work. Our numbers of E and σ(70) per cell, which are consistent with most of the primary data in the literature, suggest that in vivo (i) only a minor fraction of RNA polymerase (<20%) is involved in elongation and (ii) σ(70) is in excess of total E. Modeling the partitioning of RNA polymerase between promoters, nonspecific DNA binding sites, and the cytoplasm suggested that even weak promoters will be saturated with Eσ(70) in vivo unless nonspecific DNA binding by Eσ(70) is rather significant. In addition, the model predicted that σs compete for binding to E only when their total number exceeds the total amount of RNA polymerase (excluding that involved in elongation) and that weak promoters will be preferentially subjected to σ competition

Delayed-onset ataxia in mice lacking α-tocopherol transfer protein: Model for neuronal degeneration caused by chronic oxidative stress

α-Tocopherol transfer protein (α-TTP) maintains the concentration of serum α-tocopherol (vitamin E), one of the most potent fat-soluble antioxidants, by facilitating α-tocopherol export from the liver. Mutations of the α-TTP gene are linked to ataxia with isolated vitamin E deficiency (AVED). We produced a model mouse of AVED by deleting the α-TTP gene, which showed ataxia and retinal degeneration after 1 year of age. Because the brain α-TTP functions in maintaining α-tocopherol levels in the brain, α-tocopherol was completely depleted in the α-TTP(−/−) mouse brain, and the neurological phenotype of α-TTP(−/−) mice is much more severe than that of wild-type mice when maintained on an α-tocopherol-deficient diet. Lipid peroxidation in α-TTP(−/−) mice brains showed a significant increase, especially in degenerating neurons. α-Tocopherol supplementation suppressed lipid peroxidation and almost completely prevented the development of neurological symptoms. This therapy almost completely corrects the abnormalities in a mouse model of human neurodegenerative disease. Moreover, α-TTP(−/−) mice may prove to be excellent animal models of delayed onset, slowly progressive neuronal degeneration caused by chronic oxidative stress

Role of Lkb1, the causative gene of Peutz–Jegher's syndrome, in embryogenesis and polyposis

Peutz–Jeghers syndrome (PJS) is a dominantly inherited human disorder characterized by gastrointestinal hamartomatous polyposis and mucocutaneous melanin pigmentation. LKB1 (STK11) serine/threonine kinase is the product of the causative gene of PJS, which has been mapped to chromosome 19p13.3. However, several studies have produced results that are not consistent with a link between LKB1 gene mutation and PJS. We constructed a knockout gene mutation of Lkb1 to determine whether it is the causative gene of PJS and to examine the biological role of the Lkb1 gene. Lkb1(−/−) mice died in utero between 8.5 and 9.5 days postcoitum. At 9.0 days postcoitum, Lkb1(−/−) embryos were generally smaller than their age-matched littermates, showed developmental retardation, and did not undergo embryonic turning. Multiple gastric adenomatous polyps were observed in 10- to 14-month-old Lkb1(+/−) mice. Our results indicate that functional Lkb1 is required for normal embryogenesis and that it is related to tumor development. The Lkb1(+/−) mouse is suitable for studying molecular mechanism underlying the development of inherited gastric tumors in PJS