Veronique Arluison

Paris Diderot University | UP7

Under specific conditions, some proteins can self-assemble into fibrillar structures called amyloids. Initially, these proteins were associated with neurodegenerative diseases in eucaryotes. Nevertheless, they have now been identified in the three domains of life. In bacteria, they are involved in diverse biological processes and are usually useful...

Useful structural information about the conformation of nucleic acids can be quickly acquired by circular and linear dichroism (CD/LD) spectroscopy. These techniques, rely on the differential absorption of polarised light and are indeed extremely sensitive to subtle changes in the structure of chiral biomolecules. Many CD analyses of DNA or DNA:pro...

Due to their two-cell membranes, Gram-negative bacteria are particularly resistant to antibiotics. Recent investigations aimed at exploring new target proteins involved in Gram-negative bacteria adaptation helped to identify environmental changes encountered during infection. One of the most promising approaches in finding novel targets for antibac...

Microcin E492 (MccE492) is an antimicrobial peptide and proposed virulence factor produced by some Klebsiella pneumoniae strains, which, under certain conditions, form amyloid fibers, leading to the loss of its antibacterial activity. Although this protein has been characterized as a model functional amyloid, the secondary structure transitions beh...

Hfq is a pleitropic actor that serves as stress response and virulence factor in the bacterial cell. To execute its multiple functions, Hfq assembles into symmetric torus-shaped hexamers. Extending outward from the hexameric core, Hfq presents a C-terminal region, described as intrinsically disordered in solution. Many aspects of the role and the s...

Guanine-rich DNA can fold into highly stable four-stranded DNA structures called G-quadruplexes (G4). Originally identified in sequences from telomeres and oncogene promoters, they can alter DNA metabolism. Indeed, G4-forming sequences represent obstacles for the DNA polymerase, with important consequences for cell life as they may lead to genomic...

The possible carrier role of Outer Membrane Vesicles (OMVs) for small regulatory noncoding RNAs (sRNAs) has recently been demonstrated. Nevertheless, to perform their function, these sRNAs usually need a protein cofactor called Hfq. In this work we show, by using a combination of infrared and circular dichroism spectroscopies, that Hfq, after inter...

Hfq is a pleitropic actor, serving as stress response and virulence factor in the bacterial cell. It achieves this function mostly through its binding to small noncoding RNAs, conferring annealing with mRNAs. To execute these functions, Hfq assembles into a symmetric torus-shaped hexamer, called the Sm-core, with two concave structurally distinct s...

The Nucleic Acid Circular Dichroism Database (NACDDB) is a public repository that archives and freely distributes circular dichroism (CD) and synchrotron radiation CD (SRCD) spectral data about nucleic acids, and the associated experimental metadata, structural models, and links to literature. NACDDB covers CD data for various nucleic acid molecule...

The bacterial chromosomic DNA is packed within a membrane-less structure, the nucleoid, due to the association of DNA with proteins called Nucleoid Associated Proteins (NAPs). Among these NAPs, Hfq is one of the most intriguing as it plays both direct and indirect roles on DNA structure. Indeed, Hfq is best known to mediate post-transcriptional reg...

Interactions between proteins and single-stranded DNA (ssDNA) are crucial for many fundamental biological processes, including DNA replication and genetic recombination. Thus, understanding detailed mechanisms of these interactions is necessary to uncover regulatory rules occurring in all living cells. The RNA-binding Hfq is a pleiotropic bacterial...

Amyloids are protein assemblies based on fibrillar arrangements of β-sheets. They were initially identified in various proteins associated with neurodegenerative diseases. However, there is now increasing evidence of alternative roles for amyloids, with beneficial aspects for cells. In particular, prokaryotes use amyloids as functional assemblies,...

Hfq is a pleiotropic regulator that mediates several aspects of bacterial RNA metabolism. The protein notably regulates translation efficiency and RNA decay in Gram-negative bacteria, usually via its interaction with small regulatory RNAs. Previously, we showed that the Hfq C-terminal region forms an amyloid-like structure and that these fibrils in...

Small-angle scattering is a powerful technique to obtain structural information on biomacromolecules in aqueous solution at the sub-nanometer and nanometer length scales. It provides the sizes and overall shapes of the scattering particles. While small-angle X-ray scattering (SAXS) has often been used for structural analysis of a single-component s...

Hfq is a bacterial master regulator which promotes the pairing of nucleic acids. Due to the high molecular weight of the complexes formed between nucleic acids and the amyloid form of the protein, it is difficult to analyze solely by a gel shift assay the complexes formed, as they all migrate at the same position in the gel. In addition, precise ki...

Amyloid inhibitors, such as the green tea compound epigallocatechin gallate EGCG, apomorphine or curlicide, have antibacterial properties. Conversely, antibiotics such as tetracycline derivatives or rifampicin also affect eukaryotic amyloids formation and may be used to treat neurodegenerative diseases. This opens the possibility for existing drugs...

Bacterial chromosomal DNA is packed within a non-membranous structure, the nucleoid, thanks to nucleoid associated proteins (NAPs). The role of bacterial amyloid has recently emerged among these NAPs, particularly with the nucleoid-associated protein Hfq that plays a direct role in DNA compaction. In this chapter, we present a 3D imaging technique,...

The expression and conformation of bacterial proteins and peptides can be monitored in situ by Fourier transform infrared spectroscopy (FTIR), provided that the concentration of the protein of interest is sufficient. Here, we describe a simple protocol to analyze the conformation adopted by a specific amyloid protein in Escherichia coli cells, the...

Nanofluidics is an emerging methodology to investigate single biomacromolecules without functionalization and/or attachment of the molecules to a substrate. In conjunction with fluorescence microscopy, it can be used to investigate structural and dynamical aspects of amyloid-DNA interaction. Here, we summarize the methodology for fabricating lab-on...

Fourier transform infrared spectroscopy (FTIR), circular dichroism (CD), and orientated circular dichroism (OCD) are complementary spectroscopies widely used for the analysis of protein samples such as the amyloids commonly renowned as neurodegenerative agents. Determining the secondary structure content of proteins, such as aggregated β-sheets ins...

The mobility of protein is fundamental in the machinery of life. Here, we have investigated the effect of DNA binding in conjunction with DNA internal motion of the bacterial Hfq master regulator devoid of its amyloid C-terminus domain. Hfq is one of the most abundant nucleoid associated proteins that shape the bacterial chromosome and is involved...

Bacterial chromosomic DNA is packed within a membrane-less structure, the nucleoid, thanks to proteins called Nucleoid Associated Proteins (NAPs). The NAP composition of the nucleoid varies during the bacterial life cycle and is growth phase-dependent. Among these NAPs, Hfq is one of the most intriguing as it plays both direct and indirect roles on...

Nucleic acid amyloid proteins interactions have been observed in the past few years. These interactions often promote protein aggregation. Nevertheless, molecular basis and physiological consequences of these interactions are still poorly understood. Additionally, it is unknown whether the nucleic acid promotes the formation of self-assembly due to...

Hfq is a bacterial RNA chaperone which promotes the pairing of small noncoding RNAs to target mRNAs, allowing post-transcriptional regulation. This RNA annealing activity has been attributed for years to the N-terminal region of the protein that forms a toroidal structure with a typical Sm-fold. Nevertheless, many Hfqs, including that of Escherichi...

Hfq is a bacterial regulator with key roles in gene expression. The protein notably regulates translation efficiency and RNA decay in Gram-negative bacteria, thanks to its binding to small regulatory noncoding RNAs. This property is of primary importance for bacterial adaptation and survival in hosts. Small RNAs and Hfq are, for instance, involved...

Fourier transform infrared (FTIR) and circular dichroism (CD) spectroscopies are fast techniques providing important information about the conformation of nucleic acids and proteins. These vibrational and electronic absorption spectroscopies are extremely sensitive to any change in molecular structure. While numerous reviews describe how to analyze...

Amyloid fibrils are composed of aggregated peptides or proteins in a fibrillary structure with a higher β-sheet content than their native structure. Attenuated total reflection Fourier transform infrared spectroscopy only provides bulk analysis of a sample therefore it is impossible to discriminate between different aggregated structures. To overco...

G-rich DNA repeats that can form G-quadruplex structures are prevalent in bacterial genomes and are frequently associated with regulatory regions of genes involved in virulence, antigenic variation, and antibiotic resistance. These sequences are also inherently mutagenic and can lead to changes affecting cell survival and adaptation. Transcription...

Front Cover: Polyionenes (PI) have been covalently grafted onto glass surfaces via a process combining polydopamine coating, diazonium‐induced polymerization and polyaddition. The presence of positive charges and aliphatic spacers in the PI grafted chains causes disruption of the bacterial membrane. Microbial adhesion studies demonstrated the proad...

Polyionenes (PI) with stable positive charges and tunable hydrophobic spacers in the polymer backbone, are shown to be particularly efficient regarding antimicrobial properties. This effect can be modulated since it increases with the length of hydrophobic spacers, i.e., the number of methylene groups between quaternary ammoniums. Now, to further e...

Molecular transport of biomolecules plays a pivotal role in the machinery of life. Yet, this role is poorly understood due the lack of quantitative information. Here, the role and properties of the C-terminal region of Escherichia coli Hfq is reported, involved in controlling the flow of a DNA solution. A combination of experimental methodologies h...

Molecular transport of biomolecules plays a pivotal role in the machinery of life. Yet, this role is poorly understood due the lack of quantitative information. Here, the role and properties of the C-terminal region of Escherichia coli Hfq is reported, involved in controlling the flow of a DNA solution. A combination of experimental methodologies h...

Circular dichroism (CD) spectroscopy is a fast and simple technique providing important information about the conformation of nucleic acids, proteins, sugars, lipids, and their interactions between each other. This electronic absorption spectroscopy method is extremely sensitive to any change in molecular structure containing asymmetric molecules....

Atomic force and transmission electron microscopies (AFM/TEM) are powerful tools to analyze RNA-based nanostructures. While cryo-TEM analysis allows the determination of near-atomic resolution structures of large RNA complexes, this chapter intends to present how RNA nanostructures can be analyzed at room temperature on surfaces. Indeed, TEM and AF...

Fourier transform infrared (FTIR) spectroscopy has been widely used for the analysis of both protein and nucleic acid secondary structure. This is one of the vibration spectroscopy methods that are extremely sensitive to any change in molecular structure. While numerous reports describe how to proceed to analyze protein and deoxyribonucleic acid (D...

This volume looks at the different spectroscopic and biophysical methods used by researchers to study the structure and folding of RNA, and to follow their interactions with proteins. The chapters in this book cover topics such as single-molecule spectroscopy of multiple RNA species; surface plasmon resonance, MS or microcalorimetry for investigati...

Certain G-rich DNA repeats can form quadruplex in bacterial chromatin that can present blocks to DNA replication and, if not properly resolved, may lead to mutations. To understand the participation of quadruplex DNA in genomic instability in Escherichia coli (E. coli), mutation rates were measured for quadruplex-forming DNA repeats, including (G3T...

The Hfq protein is reported to be involved in environmental adaptation and virulence of several bacteria. In Gram-negative bacteria, Hfq mediates the interaction between regulatory noncoding RNAs and their target mRNAs. Besides these RNA-related functions, Hfq is also associated with DNA and is a part of the bacterial chromatin. Its precise role in...

Amyloids are supramolecular protein assemblies based on fibrillar arrangements of β‐sheets that were first found as linked to neurodegenerative and systemic human diseases. However, there is now overwhelming evidence on alternative roles of amyloids as functional assemblies and as epigenetic determinants of beneficial traits, both in Fungi and Meta...

Hfq is a bacterial protein that regulates gene expression at the post-transcriptional level in Gram-negative bacteria. We have previously shown that Escherichia coli Hfq protein, and more precisely its C-terminal region (CTR), self-assembles into an amyloid-like structure in vitro. In the present work, we present evidence that Hfq unambiguously for...

Propagation of structural information through conformational changes in host‐encoded amyloid proteins is at the root of many neurodegenerative disorders. Although important breakthroughs have been made in the field, fundamental issues like the 3D‐structures of the fibrils involved in some of those disorders are still to be elucidated. To better cha...

Hfq is a pleiotropic regulator that has key roles in the control of genetic expression. The protein noticeably regulates translation efficiency and RNA decay in Gram-negative bacteria, due to the Hfq-mediated interaction between small regulatory noncoding RNA and mRNA. This property is of primary importance for bacterial adaptation and virulence. W...

Hfq is a pleiotropic regulator that mediates several aspects of bacterial RNA metabolism. The protein notably regulates translation efficiency and RNA decay in Gram-negative bacteria, usually via its interaction with small regulatory RNA. Besides these RNA-related functions, Hfq has also been described as one of the nucleoid associated proteins sha...

Post-transcriptional control of gene expression by small regulatory noncoding RNA (sRNA) needs protein accomplices to occur. Past research mainly focused on the RNA chaperone Hfq as cofactor. Nevertheless, recent studies indicated that other proteins might be involved in sRNA-based regulations. As some of these proteins have been shown to self-asse...

In recent years, single-molecule fluorescence resonance energy transfer (smFRET) has emerged as a powerful technique to study macromolecular interactions. The chief advantages of smFRET analysis compared to bulk measurements include the possibility to detect sample heterogeneities within a large population of molecules and the facility to measure k...

Regulation of RNA turnover is of utmost importance for controlling the concentration of transcripts and consequently cellular protein levels. Among the processes controlling RNA decay, small noncoding regulatory RNAs (sRNAs) have recently emerged as major new players. In this chapter, we describe and discuss protocols that can be used to measure sR...

The effect of the heat unstable nucleoid structuring protein HU on the conformation of single DNA molecules confined in a nanochannel was investigated with fluorescence microscopy. Pre-incubated DNA molecules contract in the longitudinal direction of the channel with increasing concentration of HU. This contraction is mainly due to HU-mediated brid...

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Hfq is a bacterial protein that is involved in several aspects of nucleic acids metabolism. It has been described as one of the nucleoid associated proteins shaping the bacterial chromosome, although it is better known to influence translation and turnover of cellular RNAs. Here, we explore the role of Escherichia coli Hfq's C-terminal domain in th...

Nucleoid Associated Proteins (NAPs) play a key role in the compaction and expression of the prokaryotic genome. Here we report the organization of a major NAP, the protein H-NS on a double stranded DNA fragment. For this we have carried out a small angle neutron scattering study in conjunction with contrast variation to obtain the contributions to...

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Hfq is a bacterial pleiotropic regulator that mediates several aspects of nucleic acids metabolism. The protein notably influences translation and turnover of cellular RNAs. Although most previous contributions concentrated on Hfq's interaction with RNA, its association to DNA has also been observed in vitro and in vivo. Here, we focus on DNA-compa...

RNAs are flexible molecules involved in a multitude of roles in the cell. Specifically, noncoding RNAs (i.e., RNAs that do not encode a protein) have important functions in the regulation of biological processes such as RNA decay, translation, or protein translocation. In bacteria, most of those noncoding RNAs have been shown to be critical for pos...

Accumulating evidence indicates that RNA metabolism components assemble into supramolecular cellular structures to mediate functional compartmentalization within the cytoplasmic membrane of the bacterial cell. This cellular compartmentalization could play important roles in the processes of RNA degradation and maturation. These components include H...

The ability to study the localization and organization of proteins within the tiny cells of bacteria, such as Escherichia coli, has paved the way for a new and exciting era of prokaryotic cellular biology. Previously unrecognized levels of spatiotemporal and supramolecular organization of proteins have been revealed within the prokaryotic cell that...

The bacterial actin-homolog MreB is a key player in bacterial cell-wall biosynthesis and is required for the maintenance of the rod-like morphology of Escherichia coli. However, how MreB cellular levels are adjusted to growth conditions is poorly understood. Here, we show that DsrA, an E. coli small noncoding RNA (sRNA), is involved in the post-tra...

DsrA is an Escherichia coli small noncoding RNA that acts by base pairing to some mRNAs in order to control their translation and turnover. It was recently shown that DsrA is able to self-associate in a way similar to DNA and to build nanostructures. Although functional consequence of this RNA self-assembly in vivo is not yet understood, the format...

Recent discoveries of noncoding regulatory RNAs have led to further understanding of the elements controlling genetic expression. In E. coli, most of those ncRNAs for which functional knowledge is available were shown to be dependent on the Hfq RNA chaperone and to act as inhibitors of translation by base pairing with their mRNA target. Nevertheles...

The primacy and power of RNA in governing many processes of life has begun to be more fully appreciated in both the discovery and inventive sciences. A variety of RNA interactions regulate gene expression, and structural self-assembly underlies many of these processes. The understanding sparked by these discoveries has inspired and informed the eng...

In Escherichia coli, the essential motor protein Rho promotes transcription termination in a tightly controlled manner that is not fully understood. Here, we show that the general post-transcriptional regulatory protein Hfq associates with Rho to regulate Rho function. The Hfq:Rho complex can be further stabilized by RNA bridging both factors in a...

Hfq is a key regulator involved in multiple aspects of stress tolerance and virulence of bacteria. There has been an intriguing question as to how this RNA chaperone achieves two completely opposite functions—annealing and unwinding—for different RNA substrates. To address this question, we studied the Hfq-mediated interaction of fragments of a non...

Hfq is a bacterial protein involved in RNA metabolism. Besides this, Hfq's role in DNA restructuring has also been suggested. Since this mechanism remains unclear, we examined the DNA conformation upon Hfq binding by combining vibrational spectroscopy and neutron scattering. Our analysis reveals that Hfq, which preferentially interacts with deoxyad...

RNA molecules are important factors involved in different cellular processes and have a multitude of roles in the cell. These roles include serving as a temporary copy of genes used for protein synthesis or functions in translational machinery. Interestingly, RNA is so far the only biological molecule that serves both as a catalyst (like proteins)...

Background Hfq is a bacterial protein involved in several aspects of nucleic acid transactions, but one of its best-characterized functions is to affect the post-transcriptional regulation of mRNA by virtue of its interactions with stress-related small regulatory (sRNA). Methodology and Principal Finding By using cellular imaging based on the meta...

RNA molecules are important factors involved in different cellular processes and have a multitude of roles in the cell. These roles include serving as a temporary copy of genes used for protein synthesis or functions in translational machinery. Interestingly, RNA is so far the only biological molecule that serves both as a catalyst (like proteins)...

The core light-harvesting LH1 protein from Rhodospirillum rubrum can dissociate reversibly in the presence of n-octyl-beta-D-glucopyranoside into smaller subunit forms, exhibiting a dramatic blue-shift in absorption. During this process, two main species are observed: a dimer that absorbs at 820 nm (B820) and a monomer absorbing at 777 nm (B777). I...

Sm-like proteins are ubiquitous ring-shaped oligomers that exhibit a variety of nucleic acid-binding activities. They have been linked functionally to various cellular events involving RNA, and it is generally believed that their activity is exerted via the passive binding of nucleic acids. Our earlier studies of the Sm-like Escherichia coli protei...

Hfq protein is vital for the function of many non-coding small (s)RNAs in bacteria but the mechanism by which Hfq facilitates the function of sRNA is still debated. We developed a fluorescence resonance energy transfer assay to probe how Hfq modulates the interaction between a sRNA, DsrA, and its regulatory target mRNA, rpoS. The relevant RNA fragm...

Hfq is a nucleic acid-binding protein that functions as a global regulator of gene expression by virtue of its interactions with several small, non-coding RNA species. Originally identified as an Escherichia coli host factor required for RNA phage Qbeta replication, Hfq is now known to post-transcriptionally regulate bacterial gene expression by mo...

In all living cells 3' ends of RNA are posttranscriptionally elongated or shortened by nucleotidyl transferases and ribonucleases. The detailed analysis of the rpsO mRNA of Escherichia coli presented here demonstrates that transcription terminates in vivo at two sites located seven and eight nucleotides downstream from the GC-rich hairpin of the in...

The Hfq (Host factor 1) polypeptide is a nucleic acid binding protein involved in the synthesis of many polypeptides. Hfq particularly affects the translation and the stability of several RNAs. In an earlier study, the use of fold recognition methods allowed us to detect a relationship between Escherichia coli Hfq and the Sm topology. This topology...

The effect of partial digestion by trypsin and GluC protease on the association of the membrane polypeptides of LH1 from Rhodospirillum (Rsp.) rubrum was studied. Trypsin and GluC protease treatments of LH1 result in the cleavage of the first three amino acids from the alpha polypeptide and of the first 18 amino acids from the beta polypeptide, res...

The Hfq protein, which shares sequence and structural homology with the Sm and Lsm proteins, binds to various RNAs, primarily recognizing AU‐rich single‐stranded regions. In this paper, we study the ability of the Escherichia coli Hfq protein to bind to a polyadenylated fragment of rpsO mRNA. Hfq exhibits a high specificity for a 100‐nucleotide RNA...

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...

The core light-harvesting protein from Rhodospirillum rubrum is of particular interest for studying membrane polypeptide association, as it can be reversibly dissociated in the presence of n-octyl-beta-D-glucopyranoside (betaOG) into smaller subunit forms, which exhibit dramatically blue-shifted absorption properties (Miller et al. (1987) Biochemis...

The Hfq polypeptide of Escherichia coli is a nucleic acid-binding protein involved in the expression of many proteins. Derivation of its three-dimensional structure is important for our understanding of its role in gene regulation at the molecular level. In this study, we combined computational and biophysical analysis to derive a possible structur...

We have studied the equilibrium between the dissociated B777 form (absorbing at 777 nm) of the light-harvesting complex of Rhodospirillum rubrum and the oligomeric B820 form. Analysis of the reaction order for the B820 dissociation reaction to form B777 shows that this reaction depends on the concentration of octylglucoside detergent (n-octyl-beta-...

Yeast RNA:pseudouridine synthetase Pus1 catalyzes the formation of pseudouridines in tRNAs. We report here the quaternary structure of purified recombinant Pus1 in solution. At low concentration, in the absence of tRNA, Pus1 oligomerizes while at high concentration it precipitates. This oligomerization/aggregation can be prevented by addition of do...

Pseudouridine synthetase Pus1 from Saccharomyces cerevisiae is a multisite-specific enzyme that catalyses the formation of pseudouridine residues at different positions in several tRNA transcripts. Recombinant Pus1, tagged with six histidine residues at its N terminus was expressed in Escherichia coli and purified. Transcripts of yeast tRNAValand i...

RNA:pseudouridine synthetase (Pus1) from Saccharomyces cerevisiae is a multisite specific enzyme that catalyzes the formation of pseudouridine at positions 34 and 36 of intron-containing precursor tRNAIle and at positions 27 and/or 28 of several yeast tRNAs. In this paper we demonstrate that the purified recombinant Pus1, expressed in Escherichia c...