S Rimsky

Ecole normale supérieure de Cachan, Cachon, Île-de-France, France

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Publications (24)136.78 Total impact

  • Sylvie Rimsky, Andrew Travers
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    ABSTRACT: Bacterial DNA is organised in a compact nucleoid body that is tightly associated with the coupled transcription and translation of mRNAs. This structure contains abundant DNA-binding proteins which perform both structural and regulatory roles, and, in Escherichia coli, serve to buffer and organise pervasive DNA superhelicity. We argue that NAPs coordinate regulation of gene expression and superhelicity at the global (or chromosomal) and at local (corresponding to promoter activity and genetic recombination) levels.
    Current opinion in microbiology 01/2011; 14(2):136-41. · 7.87 Impact Factor
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    ABSTRACT: For the structural determination of a ligand bound to an amorphous macromolecular system, solid-state NMR can be used to provide interatomic distances. It is shown here that selective labeling in discrete locations with tritium enables accurate measurement of long-range distances owing to the high gyromagnetic ratio of this nucleus, without structural modification of the molecule. This approach gives access to the largest NMR distance ever measured between two nuclei (14.4 A). (3)H MAS NMR appears to be a promising tool for structural applications in the biological and material sciences.
    Journal of the American Chemical Society 02/2010; 132(6):1734-5. · 10.68 Impact Factor
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    Ferric C Fang, Sylvie Rimsky
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    ABSTRACT: H-NS, a nucleoid-associated DNA-binding protein of enteric bacteria, was discovered 35 years ago and subsequently found to exert widespread and highly pleiotropic effects on gene regulation. H-NS binds to high-affinity sites and spreads along adjacent AT-rich DNA to silence transcription. Preferential binding to sequences with higher AT-content than the resident genome allows H-NS to repress the expression of foreign DNA in a process known as 'xenogeneic silencing.' Counter-silencing by a variety of mechanisms facilitates the evolutionary acquisition of horizontally transferred genes and their integration into pre-existing regulatory networks. This review will highlight recent insights into the mechanism and biological importance of H-NS-DNA interactions.
    Current Opinion in Microbiology 05/2008; 11(2):113-20. · 8.23 Impact Factor
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    ABSTRACT: H-NS is a protein of the bacterial nucleoid involved in DNA compaction and transcription regulation. In vivo, H-NS selectively silences specific genes of the bacterial chromosome. However, many studies have concluded that H-NS binds sequence-independently to DNA, leaving the molecular basis for its selectivity unexplained. We show that the negative regulatory element (NRE) of the supercoiling-sensitive Escherichia coliproU gene contains two identical high-affinity binding sites for H-NS. Cooperative binding of H-NS is abrogated by changes in DNA superhelical density and temperature. We further demonstrate that the high-affinity sites nucleate cooperative binding and establish a nucleoprotein structure required for silencing. Mutations in these sites result in loss of repression by H-NS. In this model, silencing at proU, and by inference at other genes directly regulated by H-NS, is tightly controlled by the cooperativity between bound H-NS molecules.
    Nature Structural & Molecular Biology 06/2007; 14(5):441-8. · 11.90 Impact Factor
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    ABSTRACT: We compared coupling approaches of SPR to LC-MS and ProteinChip-based mass spectrometry (SELDI) as a means of identifying proteins captured on DNA surfaces. The approach we outline has the potential to allow multiple, quantitative analysis of macromolecular interactions followed by rapid mass spectrometry identification of retained material.
    Nucleic Acids Research 02/2007; 35(6):e39. · 8.81 Impact Factor
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    ABSTRACT: The global transcriptional regulator H-NS selectively silences bacterial genes associated with pathogenicity and responses to environmental insults. Although there is ample evidence that H-NS binds preferentially to DNA containing curved regions, we show here that a major basis for this selectivity is the presence of a conserved sequence motif in H-NS target transcriptons. We further show that there is a strong tendency for the H-NS binding sites to be clustered, both within operons and in genes contained in the pathogenicity-associated islands. In accordance with previously published findings, we show that these motifs occur in AT-rich regions of DNA. On the basis of these observations, we propose that H-NS silences extensive regions of the bacterial chromosome by binding first to nucleating high-affinity sites and then spreading along AT-rich DNA. This spreading would be reinforced by the frequent occurrence of the motif in such regions. Our findings suggest that such an organization enables the silencing of extensive regions of the genetic material, thereby providing a coherent framework that unifies studies on the H-NS protein and a concrete molecular basis for the genetic control of H-NS transcriptional silencing.
    Nucleic Acids Research 02/2007; 35(18):6330-7. · 8.81 Impact Factor
  • Sylvie Rimsky, Malcolm Buckle
    Encyclopedia of Molecular Cell Biology and Molecular Medicine, 09/2006; , ISBN: 9783527600908
  • Sylvie Rimsky
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    ABSTRACT: H-NS belongs to the group of histone-like proteins in Gram-negative bacteria and is also a pleiotropic regulator of genes implicated in many responses to environmental changes. It plays a dual role in structuring DNA and in regulating transcription. Recent advances have been made in elucidating the structure and oligomerisation properties of this protein, thus aiding in the understanding of the molecular relationship between its two major functions.
    Current Opinion in Microbiology 05/2004; 7(2):109-14. · 8.23 Impact Factor
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    ABSTRACT: The histone-like nucleoid structuring (H-NS) protein is a global modulator of gene expression in Gram-negative bacteria. VicH, the H-NS protein of Vibrio cholerae, regulates the expression of certain major virulence determinants implicated in the pathogenesis of cholera. We present here the 2.5A crystal structure of the N-terminal oligomerisation domain of VicH (VicH_Nt). VicH_Nt adopts the same fold and dimeric assembly as the NMR structure of Escherichia coli H-NS_Nt, thus validating this fold against conflicting data. The structural similarity of V.cholerae VicH_Nt and E.coli H-NS_Nt, despite differences in origin, system of expression, experimental conditions and techniques used, indicates that the fold determined in our studies is robust to experimental conditions. Structural analysis and homology modelling were carried out to further elucidate the molecular basis of the functional polyvalence of the N-terminal domain. Our analysis of members of the H-NS superfamily supports the suggestion that the oligomerisation function of H-NS_Nt is conserved even in more distantly related proteins.
    Journal of Molecular Biology 12/2003; 334(2):179-85. · 3.91 Impact Factor
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    ABSTRACT: H-NS, a protein found in Gram-negative bacteria, is involved in structuring the bacterial chromosome and acts as a global regulator for the expression of a wide variety of genes. These functions are correlated with both its DNA-binding and oligomerization properties. We have identified the minimal dimerization domain of H-NS, a 46 amino acid-long N-terminal fragment, and determined its structure using heteronuclear NMR spectroscopy. The highly intertwined structure of the dimer, reminiscent of a handshake, defines a new structural fold, which may offer a possibility for discriminating prokaryotic from eukaryotic proteins in drug design. Using mutational analysis, we also show that this N-terminal domain actively contributes to DNA binding, conversely to the current paradigm. Together, our data allows us to propose a model for the action of full length H-NS.
    Nature Structural Biology 04/2003; 10(3):212-8.
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    ABSTRACT: At several E. coli promoters, initiation of transcription is repressed by a tight nucleoprotein complex formed by the assembly of the H-NS protein. In order to characterize the relationship between the structure of H-NS oligomers in solution and on relevant DNA fragments, we have compared wild-type H-NS and several transdominant H-NS mutants using gel shift assays, DNase I footprinting, analytical ultracentrifugation, and reactivity toward a cross-linking reagent. In solution, oligomerization occurs through two protein interfaces, one necessary to construct a dimeric core (and involving residues 1-64) and the other required for subsequent assembly of these dimers. We show that, as well as region 64-95, residues present in the NH(2)-terminal coiled coil domain also participate in this second interface. Our results support the view that the same interacting interfaces are also involved on the DNA. We propose that the dimeric core recognizes specific motifs, with the second interface being critical for their correct head to tail assembly. The COOH-terminal domain of the protein contains the DNA binding motif essential for the discrimination of this specific functional assembly over competitive nonspecific H-NS polymers.
    Journal of Biological Chemistry 12/2002; 277(44):41657-66. · 4.65 Impact Factor
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    S Rimsky, F Zuber, M Buckle, H Buc
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    ABSTRACT: The H-NS protein is a major component of the bacterial nucleoid and plays a crucial role in the global gene regulation of enteric bacteria. Although H-NS does not exhibit a high DNA sequence specificity, a number of H-NS-responsive promoters have been shown to contain regions of intrinsic DNA curvature located either upstream or downstream of the transcription start point. We have studied H-NS binding to DNA and in vitro transcriptional regulation by H-NS at several synthetic promoters with or without curved sequences inserted upstream of the Pribnow box. We show how such inserts determine the final organization of H-NS-containing nucleoprotein complexes and how this affects transcription. We refine a two-step mechanism for the constitution of H-NS assemblies that are efficient in regulation.
    Molecular Microbiology 01/2002; 42(5):1311-23. · 5.03 Impact Factor
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    ABSTRACT: During the last decade, the hns gene and its product, the H-NS protein, have been extensively studied in Escherichia coli. H-NS-like proteins seem to be widespread in gram-negative bacteria. However, unlike in E. coli and in Salmonella enterica serovar Typhimurium, little is known about their role in the physiology of those organisms. In this report, we describe the isolation of vicH, an hns-like gene in Vibrio cholerae, the etiological agent of cholera. This gene was isolated from a V. cholerae genomic library by complementation of different phenotypes associated with an hns mutation in E. coli. It encodes a 135-amino-acid protein showing approximately 50% identity with both H-NS and StpA in E. coli. Despite a low amino acid conservation in the N-terminal part, VicH is able to cross-react with anti-H-NS antibodies and to form oligomers in vitro. The vicH gene is expressed as a single gene from two promoters in tandem and is induced by cold shock. A V. cholerae wild-type strain expressing a vicHDelta92 gene lacking its 3' end shows pleiotropic alterations with regard to mucoidy and salicin metabolism. Moreover, this strain is unable to swarm on semisolid medium. Similarly, overexpression of the vicH wild-type gene results in an alteration of swarming behavior. This suggests that VicH could be involved in the virulence process in V. cholerae, in particular by affecting flagellum biosynthesis.
    Journal of Bacteriology 05/2000; 182(7):2026-32. · 3.19 Impact Factor
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    ABSTRACT: The chromosome of Escherichia coli K-12 contains a putative gene, yheB (chiA), at centisome 74.7, whose product shows sequence similarity with chitinases of bacterial and viral origin. We cloned the chiA (yheB) gene and demonstrated that it codes for a 94.5 kDa periplasmic protein with endochitinase/lysozyme activity. Under standard laboratory growth conditions, chiA expression is very low, as shown by the Lac- phenotype of a chiA transcriptional fusion to a promoterless lacZ reporter. To identify factors that control chitinase gene expression, we generated random Tn10 insertions in the chromosome of the fusion-containing strain, selecting for a Lac+ phenotype. The majority of the mutations that caused a Lac+ phenotype mapped to the hns gene, encoding the nucleoid-structuring protein H-NS. Transcription of chiA in vivo is driven by a single sigma70 promoter and is derepressed in an hns mutant. Using a competitive gel retardation assay, we demonstrated that H-NS binds directly and with high affinity to the chiA promoter region. In addition to hns, other E. coli mutations causing defects in global regulatory proteins, such as fis, crp or stpA in combination with hns, increased chiA expression to different extents, as did decreasing the growth temperature from 37 degrees C to 30 degrees C. A possible physiological function of ChiA (YheB) endochitinase in E. coli K-12 is discussed.
    Molecular Microbiology 04/2000; 35(6):1506-17. · 5.03 Impact Factor
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    ABSTRACT: Little is known about the molecular mechanism by which histone-like nucleoid-structuring (H-NS) protein and cyclic AMP-catabolite activator protein (CAP) complex control bacterial motility. In the present paper, we show that crp and hns mutants are nonmotile due to a complete lack of flagellin accumulation. This results from a reduced expression in vivo of fliA and fliC, which encode the specific flagellar sigma factor and flagellin, respectively. Overexpression of the flhDC master operon restored, at least in part, motility in crp and hns mutant strains, suggesting that this operon is the main target for both regulators. Binding of H-NS and CAP to the regulatory region of the master operon was demonstrated by gel retardation experiments, and their DNA binding sites were identified by DNase I footprinting assays. In vitro transcription experiments showed that CAP activates flhDC expression while H-NS represses it. In agreement with this observation, the activity of a transcriptional fusion carrying the flhDC promoter was decreased in the crp strain and increased in the hns mutant. In contrast, the activity of a transcriptional fusion encompassing the entire flhDC regulatory region extending to the ATG translational start codon was strongly reduced in both hns and crp mutants. These results suggest that the region downstream of the +1 transcriptional start site plays a crucial role in the positive control by H-NS of flagellum biosynthesis in vivo. Finally, the lack of complementation of the nonmotile phenotype in a crp mutant by activation-deficient CAP mutated proteins and characterization of cfs, a mutation resulting in a CAP-independent motility behavior, demonstrate that CAP activates flhDC transcription by binding to its promoter and interacting with RNA polymerase.
    Journal of Bacteriology 01/2000; 181(24):7500-8. · 3.19 Impact Factor
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    R M Williams, S Rimsky
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    ABSTRACT: The nucleoid-associated protein H-NS has a central role in the structuring and control of the enteric bacterial chromosome. This protein has been demonstrated to contribute to the regulation of expression for approximately thirty genes. In this article, the molecular aspects of H-NS structure and function are briefly reviewed. H-NS contains at least two independent structural domains: a C-terminal domain, involved in the DNA-protein interactions, and a N-terminal domain, likely involved in protein-protein interactions. Recent reports have revealed that H-NS is a key factor in a multi-component gene regulatory system. Factors have now been discovered which can backup or antagonise H-NS action at certain promoters. These recent findings are summarised and discussed in relationship to the role of H-NS in DNA packaging and nucleoid structure.
    FEMS Microbiology Letters 11/1997; 156(2):175-85. · 2.05 Impact Factor
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    R M Williams, S Rimsky, H Buc
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    ABSTRACT: Twelve different dominant negative mutants of the Escherichia coli nucleoid-associated protein, H-NS, have been selected and characterized in vivo. The mutants are all severely defective in promoter repression activity in a strain lacking H-NS, and they all disrupt the repression normally exerted by H-NS at two of its target promoters. From the locations of the alterations in these mutants, which result in both large truncations and amino acid substitutions, we propose that H-NAS contains at least two distinct domains. The in vitro protein-protein cross-linking data presented in this report indicate that the proposed N-terminal domain of H-NS has a role in H-NS multimerization. StpA is a protein with known structural and functional homologies to H-NS. We have analyzed the extent of these homologies by constructing and studying StpA mutants predicted to be dominant negative. Our data indicate that the substitutions and deletions found in dominant negative H-NS have similar effects in the context of StpA. We conclude that the domain organizations and functions in StpA and H-NS are closely related. Furthermore, dominant negative H-NS can disrupt the activity of native StpA, and reciprocally, dominant negative StpA can disrupt the activity of native H-NS. We demonstrate that the N-terminal domain of H-NS can be chemically cross-linked to both full-length H-NS and StpA. We account for these observations by proposing that H-NS and StpA have the ability to form hybrid species.
    Journal of Bacteriology 09/1996; 178(15):4335-43. · 3.19 Impact Factor
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    ABSTRACT: Expression of the Escherichia coli StpA protein was investigated and a functional comparison undertaken with the structurally analogous nucleoid protein H-NS. Analysis of stpA and hns expression indicated that although stpA transcript levels are much lower than those of hns, the two gene products are capable of both negative autogenous control and cross-regulation. Examination of cellular proteins in stpA, hns, or stpA-hns backgrounds revealed that StpA can repress and activate a subset of H-NS-regulated genes. Mechanistic parallels in regulation of gene expression are indicated by the ability of both proteins to inhibit transcription from promoters containing curved DNA sequences, and to form nucleoprotein structures that constrain DNA supercoils. Despite their functional similarities, each molecule is capable of independent activities. Thus, H-NS regulates a class of genes that are unaffected by StpA in vivo, whereas StpA has much stronger RNA chaperone activity in vitro. We therefore propose that in addition to its role as a molecular back-up of H-NS, StpA's superior effect on RNA may be exploited under some specific cellular conditions to promote differential gene expression.
    The EMBO Journal 04/1996; 15(6):1340-9. · 9.82 Impact Factor
  • F Zuber, D Kotlarz, S Rimsky, H Buc
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    ABSTRACT: Replacement of the CRP-binding site of the gal control region by curved sequences can lead to the restoration of promoter strength in vivo. One curved sequence called 5A6A, however, failed to do so. The gene hns exerts a strong negative control on the resulting 5A6A gal promoter as well as on the distant bla promoter, specifically in a 5A6A gal context. The product of this gene, H-NS, displays a better affinity for this particular insert compared to other curved sequences. Mechanisms by which H-NS may repress promoters both at short and long distances from a favoured binding site are discussed.
    Molecular Microbiology 05/1994; 12(2):231-40. · 5.03 Impact Factor
  • S Rimsky, A Spassky
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    ABSTRACT: The H1 protein is a likely candidate for structuring DNA in the bacterial nucleoid. We have studied determinants leading to its binding to DNA (and in particular to Escherichia coli lac and gal promoters) in vitro through the pattern of attack of both DNaseI and the copper-o-phenanthroline complex [(OP)2Cu+]. The binding of H1 depends on the primary sequence of DNA. H1 also associates with recognition sites for specific proteins, in particular with the Pribnow box and the CRP binding site. Binding of H1 to the Pribnow box of the wild-type lac promoter does not change the pattern of nucleolytic digestion with (OP)2Cu+. In contrast, binding of H1 to the strong lac promoter mutants Ps and UV5 appears to change the conformational state of this DNA. Similar changes in accessibility of the minor groove surrounding the respective binding sites were observed for both H1-DNA and CRP-DNA complexes.
    Biochemistry 05/1990; 29(15):3765-71. · 3.38 Impact Factor

Publication Stats

1k Citations
136.78 Total Impact Points

Institutions

  • 2006–2011
    • Ecole normale supérieure de Cachan
      Cachon, Île-de-France, France
  • 2008
    • University of Washington Seattle
      • Department of Laboratory Medicine
      Seattle, WA, United States
  • 1996–2004
    • French National Centre for Scientific Research
      • Centre de Biochimie Structurale
      Paris, Ile-de-France, France
  • 1984
    • Institut Pasteur
      Lutetia Parisorum, Île-de-France, France