RNA search engines empower the bacterial intranet

RNA acts not only as an information bearer in the biogenesis of proteins from genes, but also as a regulator that participates in the control of gene expression. In bacteria, small RNA molecules (sRNAs) play controlling roles in numerous processes and help to orchestrate complex regulatory networks. Such processes include cell growth and development, response to stress and metabolic change, transcription termination, cell-to-cell communication, and the launching of programmes for host invasion. All these processes require recognition of target messenger RNAs by the sRNAs. This review summarizes recent results that have provided insights into how bacterial sRNAs are recruited into effector ribonucleoprotein complexes that can seek out and act upon target transcripts. The results hint at how sRNAs and their protein partners act as pattern-matching search engines that efficaciously regulate gene expression, by performing with specificity and speed while avoiding off-target effects. The requirements for efficient searches of RNA patterns appear to be common to all domains of life

An allosteric transport mechanism for the AcrAB-TolC multidrug efflux pump

Bacterial efflux pumps confer multidrug resistance by transporting diverse antibiotics from the cell. In Gram-negative bacteria, some of these pumps form multi-protein assemblies that span the cell envelope. Here, we report the near-atomic resolution cryoEM structures of the Escherichia coli AcrAB-TolC multidrug efflux pump in resting and drug transport states, revealing a quaternary structural switch that allosterically couples and synchronizes initial ligand binding with channel opening. Within the transport-activated state, the channel remains open even though the pump cycles through three distinct conformations. Collectively, our data provide a dynamic mechanism for the assembly and operation of the AcrAB-TolC pump.This work was supported by the American Heart Association (16RNT29720001), and Grants from National Institutes of Health (P41GM103832, R01GM079429, R01GM072804, S10OD016279), the Wellcome trust, the Human Frontiers Science Program and MRC Mitochondrial Biology Unit (Grant number: U105663141). CFH is supported by a pre-doctoral training fellowship from the Keck Center of the Gulf Coast Consortia, on the NLM Training Program in Biomedical Informatics (Grant No. T15LM007093). We acknowledge the computing resources provided by the Center for Computational and Integrative Biomedical Research of Baylor College of Medicine and the Texas Advanced Computing Center at the University of Texas at Austin funded by the National Science Foundation (NSF) through grant ACI-1134872

Structural insights into RapZ-mediated regulation of bacterial amino-sugar metabolism

In phylogenetically diverse bacteria, the conserved protein RapZ plays a central role in RNA-mediated regulation of amino-sugar metabolism. RapZ contributes to the control of glucosamine phosphate biogenesis by selectively presenting the regulatory small RNA GlmZ to the essential ribonuclease RNase E for inactivation. Here, we report the crystal structures of full length Escherichia coli RapZ at 3.40 Å and 3.25 Å, and its isolated C-terminal domain at 1.17 Å resolution. The structural data confirm that the N-terminal domain of RapZ possesses a kinase fold, whereas the C-terminal domain bears closest homology to a subdomain of 6-phosphofructokinase, an important enzyme in the glycolytic pathway. RapZ self-associates into a domain swapped dimer of dimers, and in vivo data support the importance of quaternary structure in RNA-mediated regulation of target gene expression. Based on biochemical, structural and genetic data, we suggest a mechanism for binding and presentation by RapZ of GlmZ and the closely related decoy sRNA, GlmY. We discuss a scenario for the molecular evolution of RapZ through re-purpose of enzyme components from central metabolism.This work was supported by the Wellcome Trust [RG84381 to B.F.L]; and the Austrian Science Fund [P 26681-B22 and F4317 to B.G.]. GMG was supported by a Gates Cambridge Scholarship. Funding for open access charge: Wellcome Trust

Estrogen receptor transcription and transactivation: Structure-function relationship in DNA- and ligand-binding domains of estrogen receptors

Estrogen receptors are members of the nuclear receptor steroid family that exhibit specific structural features, ligand-binding domain sequence identity and dimeric interactions, that single them out. The crystal structures of their DNA-binding domains give some insight into how nuclear receptors discriminate between DNA response elements. The various ligand-binding domain crystal structures of the two known estrogen receptor isotypes (α and β) allow one to interpret ligand specificity and reveal the interactions responsible for stabilizing the activation helix H12 in the agonist and antagonist positions

Allosteric Analysis of Glucocorticoid Receptor-DNA Interface Induced by Cyclic Py-Im Polyamide: A Molecular Dynamics Simulation Study

Background: It has been extensively developed in recent years that cell-permeable small molecules, such as polyamide, can be programmed to disrupt transcription factor-DNA interfaces and can silence aberrant gene expression. For example, cyclic pyrrole-imidazole polyamide that competes with glucocorticoid receptor (GR) for binding to glucocorticoid response elements could be expected to affect the DNA dependent binding by interfering with the protein-DNA interface. However, how such small molecules affect the transcription factor-DNA interfaces and gene regulatory pathways through DNA structure distortion is not fully understood so far. Methodology/Principal Findings: In the present work, we have constructed some models, especially the ternary model of polyamides+DNA+GR DNA-binding domain (GRDBD) dimer, and carried out molecular dynamics simulations and free energy calculations for them to address how polyamide molecules disrupt the GRDBD and DNA interface when polyamide and protein bind at the same sites on opposite grooves of DNA. Conclusions/Significance: We found that the cyclic polyamide binding in minor groove of DNA can induce a large structural perturbation of DNA, i.e. a.4 A ˚ widening of the DNA minor groove and a compression of the major groove by more than 4A ˚ as compared with the DNA molecule in the GRDBD dimer+DNA complex. Further investigations for the ternary system of polyamides+DNA+GRDBD dimer and the binary system of allosteric DNA+GRDBD dimer revealed that the compression o

Transcriptional Regulation of Human Dual Specificity Protein Phosphatase 1 (DUSP1) Gene by Glucocorticoids

Background: Glucocorticoids are potent anti-inflammatory agents commonly used to treat inflammatory diseases. They convey signals through the intracellular glucocorticoid receptor (GR), which upon binding to ligands, associates with genomic glucocorticoid response elements (GREs) to regulate transcription of associated genes. One mechanism by which glucocorticoids inhibit inflammation is through induction of the dual specificity phosphatase-1 (DUSP1, a.k.a. mitogen-activated protein kinase phosphatase-1, MKP-1) gene. Methodology/Principal Findings: We found that glucocorticoids rapidly increased transcription of DUSP1 within 10 minutes in A549 human lung adenocarcinoma cells. Using chromatin immunoprecipitation (ChIP) scanning, we located a GR binding region between 21421 and 21118 upstream of the DUSP1 transcription start site. This region is active in a reporter system, and mutagenesis analyses identified a functional GRE located between 21337 and 21323. We found that glucocorticoids increased DNase I hypersensitivity, reduced nucleosome density, and increased histone H3 and H4 acetylation within genomic regions surrounding the GRE. ChIP experiments showed that p300 was recruited to the DUSP1 GRE, and RNA interference experiments demonstrated that reduction of p300 decreased glucocorticoid-stimulated DUSP1 gene expression and histone H3 hyperacetylation. Furthermore, overexpression of p300 potentiated glucocorticoid-stimulated activity of a reporter gene containing the DUSP1 GRE, and this coactivation effect was compromised when the histone acetyltransferase domain was mutated. ChIP-reChIP experiments using GR followed by p300 antibodies showed significant enrichment of the DUSP1 GRE upon glucocorticoid treatment, suggesting that GR and p300 are in the same protein complex recruited to the DUSP1 GRE. Conclusions/Significance: Our studies identified a functional GRE for the DUSP1 gene. Moreover, the transcriptional activation of DUSP1 by glucocorticoids requires p300 and a rapid modification of the chromatin structure surrounding the GRE. Overall, understanding the mechanism of glucocorticoid-induced DUSP1 gene transcription could provide insights into therapeutic approaches against inflammatory diseases. © 2010 Shipp et al

Conceiving “personality”: Psychologist’s challenges and basic fundamentals of the Transdisciplinary Philosophy-of-Science Paradigm for Research on Individuals

Scientists exploring individuals, as such scientists are individuals themselves and thus not independent from their objects of research, encounter profound challenges; in particular, high risks for anthropo-, ethno- and ego-centric biases and various fallacies in reasoning. The Transdisciplinary Philosophy-of-Science Paradigm for Research on Individuals (TPS-Paradigm) aims to tackle these challenges by exploring and making explicit the philosophical presuppositions that are being made and the metatheories and methodologies that are used in the field. This article introduces basic fundamentals of the TPS-Paradigm including the epistemological principle of complementarity and metatheoretical concepts for exploring individuals as living organisms. Centrally, the TPS-Paradigm considers three metatheoretical properties (spatial location in relation to individuals’ bodies, temporal extension, and physicality versus “non-physicality”) that can be conceived in different forms for various kinds of phenomena explored in individuals (morphology, physiology, behaviour, the psyche, semiotic representations, artificially modified outer appearances and contexts). These properties, as they determine the phenomena’s accessibility in everyday life and research, are used to elaborate philosophy-of-science foundations and to derive general methodological implications for the elementary problem of phenomenon-methodology matching and for scientific quantification of the various kinds of phenomena studied. On the basis of these foundations, the article explores the metatheories and methodologies that are used or needed to empirically study each given kind of phenomenon in individuals in general. Building on these general implications, the article derives special implications for exploring individuals’ “personality”, which the TPS-Paradigm conceives of as individual-specificity in all of the various kinds of phenomena studied in individuals