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Ni-Fe layered double hydroxide derived catalysts for non-plasma and DBD plasma-assisted CO2 methanation
Laboratoire Procédés - Plasmas - Microsystèmes - D Moreno, MV Ognier, S Motak, Grzybek, T Da Costa, P Galvez
Catalys Science & Technology - 45 17 - DOI: 10.1016/j.ijhydene.2019.06.095 - 2020
A series of bi-metallic layered double hydroxide derived materials, containing a fixed amount of Ni promoted with various amounts of Fe were obtained by co-precipitation. The synthesized materials were characterized by X-ray diffraction (XRD), temperature-programmed reduction (H 2-TPR), temperature-programmed desorption of CO 2 (CO 2-TPD), elemental analysis and low temperature N 2 sorption and tested as catalysts in CO 2 methanation at atmospheric pressure. The obtained results confirmed the formation of mixed nano-oxides after thermal decomposition of the precursor and suggest successful introduction of both nickel and iron into the layers of Layered Double Hydroxides (LDHs). The introduction of Fe into the layered double hydroxides changed the interaction between Ni and supports matrix as proven by temperature programmed reduction (H 2-TPR). The introduction of low amount of iron influenced positively the catalytic activity in CO 2 methanation at 250 C, with CO 2 conversion increasing from 21% to 72% with CH 4 selec-tivity ranging from 97 to 99% at 250 C. No other products, except CH 4 and CO were registered during the experiments. In order to enhance the catalytic activity a non-thermal plasma created by dielectric barrier discharge was applied. The obtained results prove that * Corresponding author.
Plasma-Induced Polymerizations: A New Synthetic Entry in Liquid Crystal Elastomer Actuators
Laboratoire Procédés - Plasmas - Microsystèmes - Bin Ni Mengxue Zhang Cédric Guyon Patrick Keller Michael Tatoulian Min
First published - 41 19 - https://doi.org/10.1002/marc.202000385 - 2020
The research on soft actuators including liquid crystal elastomers (LCEs) becomes more and more appealing at a time when the expansion of artificial systems is blooming. Among the various LCE actuators, the bending deformation is often in the origin of many actuation modes. Here, a new strategy with plasma technology is developed to prepare single‐layer main‐chain LCEs with thermally actuated bending and contraction deformations. Two distinct reactions, plasma polymerization and plasma‐induced photopolymerization, are used to polymerize in one step the nematic monomer mixture aligned by magnetic field. The plasma polymerization forms cross‐linked but disoriented structures at the surface of the LCE film, while the plasma‐induced photopolymerization produces aligned LCE structure in the bulk. The actuation behaviors (bending and/or contraction) of LCE films can be adjusted by plasma power, reaction time, and sample thickness. Soft robots like crawling walker and flower mimic are built by LCE films with bending actuation.
Fast carbonylation reaction from CO2 using plasma gas/liquid microreactors for radiolabeling applications
Laboratoire Procédés - Plasmas - Microsystèmes - Marion Gaudeau, Mengxue Zhang, Michaël Tatoulian, Camille Lescot and Stéphanie Ognier
Reaction Chemistry & Engineering - 5 1981-1991 - doi.org/10.1039/D0RE00289E. - 2020
Carbon-11 is undoubtedly an attractive PET radiolabeling synthon because carbon is present in all biological molecules. It is mainly found under 11CO2, but the latter being not very reactive, it is necessary to convert it into a secondary precursor. 11CO is an attractive precursor for labeling the carbonyl position through transition-metal mediated carbonylation because of its access to a wide range of functional groups (e.g., amides, ureas, ketones, esters, and carboxylic acids) present in most PET tracer molecules. However, the main limitations of 11CO labeling are the very short half-life of the radioisotope carbon-11 and its low concentration, and the low reactivity and poor solubility of 11CO in commonly used organic solvents. In this work, we show that a possible solution to these limitations is to use microfluidic reactor technology to perform carbonylation reactions, whilst a novel approach to generate CO from CO2 by plasma is described. The methodology consists of the decomposition of CO2 into CO by non-thermal DBD plasma at room temperature and atmospheric pressure, followed by the total incorporation of CO thus formed in the gas phase by carbonylation reaction, in less than 2 min of residence time. This “proof of principle” developed in carbon-12 would be further applied in carbon-11. Although considerable advances in 11CO chemistry have been reported in recent years, its application in PET tracer development is still an area of work in progress, because of the lack of commercially available synthesis instruments designed for 11C-carbonylations. To the best of our knowledge, such an innovative and efficient process, combining microfluidics and plasma, allowing the very fast organic synthesis of carbonyl molecules from CO2 with high yield, in mild conditions, has never been studied.
Performance evaluation of a MIP for the MISPE-LC determination of p-[18F]MPPF and a potential metabolite in human plasma
Laboratoire Spectrométrie de masse biologique et protéomique - F.Lecomte J.Aerts Plenevaux .Defraiteur. Chapuis-Hugonc. Rozetd. Chiape. Luxen. Pichon, Ph.Huberta C.Huberta
ELSEVIER - 180 113015 - https://doi.org/10.1016/j.jpba.2019.113015 - 2020
Within the family of serotonin (5-HT) receptors, the 5-HT1A subtype is particularly interesting as it may be involved in various physiological processes or psychological disorders. The p-[18F]MPPF, a highly selective 5-HT1A antagonist, is used for in vivo studies in human or animal by means of positron emission tomography (PET) [1].
In order to selectively extract p-[18F]MPPF and its main metabolites from plasma, molecularly imprinted polymer (MIP) was prepared against these compounds by using the p-MPPF as template. For the control of the selectivity, non-imprinted polymer (NIP) was also synthesized without template. The MIP sorbent, packed in disposable extraction cartridges (DECs), was then evaluated as molecularly imprinted solid-phase extraction (MISPE) prior to the LC determination. The conditions of extraction were evaluated in order to obtain the highest selective retention of the p-[18F]MPPF and its metabolites on this MIP. The MIP selectivity was exploited in the loading and washing steps by adjusting the pH of plasma samples at a suitable value and by selecting mixtures for the washing step to limit the contribution of non-specific interactions. Other important parameters involved in the conditioning and elution steps were also studied. Finally, a pre-validation was carried out with optimal extraction conditions to demonstrate the performance of this MISPE-LC method as a generic method in the context of evaluation of new MISPE for p-[18F]MPPF and its potential for metabolites extraction from human plasma.
Identification and semi-relative quantification of intact glycoforms by nano-LC–(Orbitrap)MS: application to the α-subunit of human chorionic gonadotropin and follicle-stimulating hormone
Laboratoire Spectrométrie de masse biologique et protéomique - Amira Al Matari, Audrey Combès, Julien Camperi, Thierry Fournier, Valérie Pichon & Nathalie Delaunay
ELSEVIER - 412 5729–5741 - , 10.1007/s00216-020-02794-3 - 2020
Human chorionic gonadotropin (hCG) and follicle-stimulating hormone (FSH) belong to the family of glycoprotein polypeptide hormones called gonadotropins. They are heterodimers sharing the α-subunit structure that has 2 N-glycosylation sites. A method based on nano-reversed-phase liquid chromatography coupled to high-resolution mass spectrometry with an Orbitrap analyzer was developed for the first time to characterize the glycosylation state of the α-subunit at the intact level. A recombinant hCG-based drug, Ovitrelle®, was analyzed. This method combined with an appropriate data treatment allowed the detection of not only the major isoforms but also the minority ones with a high mass accuracy. More than 30 hCGα glycoforms were detected without overlapping of the isotopic patterns. The figures of merit of the method were assessed. The relative standard deviations (RSDs) of the retention time ranged between 0.1 and 6.08% (n = 3), with an average of 0.4%. The RSDs of the peak area measured on the extracted ion chromatogram of each glycoform are below 38% (n = 3), with an average of 16%, thus allowing semi-relative quantification. The ability to accurately profile glycosylated variants of hCGα was next demonstrated by comparing qualitatively and semi-quantitatively 3 batches of Ovitrelle®. The method was also used to analyze 3 batches of a recombinant FSH-based drug, Puregon®, and 30 FSHα glycoforms were detected and semi-quantified. This demonstrates the high potential of this method for fast quality control or comparison of the glycosylation of glycoprotein-based pharmaceutical preparations.
Identification and semi-relative quantification of intact glycoforms by nano-LC–(Orbitrap)MS: application to the α-subunit of human chorionic gonadotropin and follicle-stimulating hormone
Laboratoire Spectrométrie de masse biologique et protéomique - Julien Camperi, Audrey Combès, Thierry Fournier, Valerie Pichon & Nathalie Delaunay
Research Paper - 412 4423–4432 - 10.1007/s00216-020-02684-8 - 2020
In the present work, the human chorionic gonadotropin (hCG) hormone was characterized for the first time by hydrophilic interaction liquid chromatography (HILIC) coupled to high-resolution (HR) quadrupole/time-of-flight (qTOF) mass spectrometry (MS) at the intact level. This heterodimeric protein, consisting of two subunits (hCGα and hCGβ), possesses 8 potential glycosylation sites leading to a high number of glycoforms and has a molecular weight of about 35 kDa. The HILIC conditions optimized in a first paper but using UV detection were applied here with MS for the analysis of two hCG-based drugs, a recombinant hCG and a hCG isolated from the urine of pregnant women. An amide column (150 × 2.1 mm, 2.6 μm, 150 Å), a mobile phase composed of acetonitrile and water both containing 0.1% of trifluoroacetic acid, and a temperature of 60 °C were used. The gradient was from 85 to 40% ACN in 30 min. The use of TFA that had been shown to be necessary for the separation of glycoforms caused, as expected, an ion suppression effect in MS that was partially overcome by increasing the amount of protein injected (2 μL at 1 mg mL−1) and reducing the detection m/z range (from 1500 to 300). These conditions allowed the detection of different glycoforms of hCGα. The performance of the HILIC-HRMS method was compared with that previously obtained in RPLC-HRMS in terms of the number of detected glycoforms, selectivity, and sensitivity. The complementarity and orthogonality of the HILIC and RP modes for the analysis of hCG at the intact level were demonstrated.
S-nitrosylation affects TRAP1 structure and ATPase activity and modulates cell response to apoptotic stimuli
Laboratoire Spectrométrie de masse biologique et protéomique - Fiorella Faienza, Matteo Lambrughi, Salvatore Rizza, Chiara Pecorari, Paola Giglio, Juan Salamanca Viloria, Maria Francesca Allega, Giovanni Chiappetta, Joëlle Vinh, Francesca Pacello, Andrea Battistoni, Andrea Rasola, Elena Papaleo, Giuseppe Filomeni
bioRxiv - - doi: 10.1016/j.bcp.2020.113869 - 2020
The mitochondrial chaperone TRAP1 has been involved in several mitochondrial functions, and modulation of its expression/activity has been suggested to play a role in the metabolic reprogramming distinctive of cancer cells. TRAP1 posttranslational modifications, i.e. phosphorylation, can modify its capability to bind to different client proteins and modulate its oncogenic activity. Recently, it has been also demonstrated that TRAP1 is S-nitrosylated at Cys501, a redox modification associated with its degradation via the proteasome. Here we report molecular dynamics simulations of TRAP1, together with analysis of long-range structural communication, providing a model according to which Cys501 S-nitrosylation induces conformational changes to distal sites in the structure of the protein. The modification is also predicted to alter open and closing motions for the chaperone function. By means of colorimetric assays and site directed mutagenesis aimed at generating C501S variant, we also experimentally confirmed that selective S-nitrosylation of Cys501 decreases ATPase activity of recombinant TRAP1. Coherently, C501S mutant was more active and conferred protection to cell death induced by staurosporine. Overall, our results provide the first in silico, in vitro and cellular evidence of the relevance of Cys501 S-nitrosylation in TRAP1 biology.
The zoonotic pathogen Leptospira interrogans mitigates environmental stress through cyclic-di-GMP-controlled biofilm production
Laboratoire Spectrométrie de masse biologique et protéomique - Thibeaux R, Soupé-Gilbert ME, Kainiu M, Girault D, Bierque E, Fernandes J, Bähre H, Douyère A, Eskenazi N, Vinh J, Picardeau M, Goarant C
NPJ Biofilms Microbiomes - 6(1) 24 - DOI [DOI] – 10.1038/s41522-020-0134-1 - 2020
The zoonotic bacterium Leptospira interrogans is the aetiological agent of leptospirosis, a re-emerging infectious disease that is a growing public health concern. Most human cases of leptospirosis result from environmental infection. Biofilm formation and its contribution to the persistence of virulent leptospires in the environment or in the host have scarcely been addressed. Here, we examined spatial and time-domain changes in biofilm production by L. interrogans. Our observations showed that biofilm formation in L. interrogans is a highly dynamic process and leads to a polarized architecture. We notably found that the biofilm matrix is composed of extracellular DNA, which enhances the biofilm’s cohesiveness. By studying L. interrogans mutants with defective diguanylate cyclase and phosphodiesterase genes, we show that biofilm production is regulated by intracellular levels of bis-(3′-5′)-cyclic dimeric guanosine monophosphate (c-di-GMP) and underpins the bacterium’s ability to withstand a wide variety of simulated environmental stresses. Our present results show how the c-di-GMP pathway regulates biofilm formation by L. interrogans, provide insights into the environmental persistence of L. interrogans and, more generally, highlight leptospirosis as an environment-borne threat to human health.
On-Chip Sample Preparation Using a ChipFilter Coupled to NanoLC-MS/MS for Bottom-Up Proteomics
Laboratoire Spectrométrie de masse biologique et protéomique - Massamba M Ndiaye, Ha Phuong Ta, Giovanni Chiappetta, Joëlle Vinh
J Proteome Res - 19(7) 2654-2663 - DOI: 10.1021/acs.jproteome.9b00832 - 2020
Sample preparation is a crucial step in bottom-up proteomics. Analytical performances of bottom-up proteomics can be improved by the miniaturization of sample preparation. Many microfluidic devices have been designed in the field of proteomics, but many of them are not capable of handling complex samples and do not integrate the processing and digestion steps. We propose a ChipFilter Proteolysis (CFP) microfluidic device as a proteomics reactor for the miniaturization of protein sample processing and digestion steps, whose design is closely related to the experimental setup of filter-aided sample processing, even if no denaturing surfactant is required. The microchip has two reaction chambers of 0.6 μL volume separated by a protein filtration membrane in regenerated cellulose (10kD cutoff) that will concentrate or retain large polypeptides and will release small molecules. Cell lysis, protein concentration, and rapid chemical or enzymatic treatment can be performed in the ChipFilter. Complex proteomic samples like yeast protein extract or whole human cells proteome have been successfully analyzed with our microchip. Compared with the membrane-based commercial ultracentrifugation cartridge, our microfluidic device offered a better proteome coverage with 10 times less starting material and 8 times faster protocol duration.
Peptides Derived From Insulin Granule Proteins Are Targeted by CD8+ T Cells Across MHC Class I Restrictions in Humans and NOD Mice
Laboratoire Spectrométrie de masse biologique et protéomique - Marie Eliane Azoury, Mahmoud Tarayrah, Georgia Afonso, Aurore Pais, Maikel L. Colli, Claire Maillard, Cassandra Lavaud, Laure Alexandre-Heymann, Sergio Gonzalez-Duque, Yann Verdier4, Joelle Vinh, Sheena Pinto, Soren Buus, Danièle Dubois-L
Diabetes - 69(12) 2678-2690 - https://doi.org/10.2337/db20-0013 - 2020
The antigenic peptides processed by β-cells and presented through surface HLA class I molecules are poorly characterized. Each HLA variant (e.g., the most common being HLA-A2 and HLA-A3) carries some peptide-binding specificity. Hence, features that, despite these specificities, remain shared across variants may reveal factors favoring β-cell immunogenicity. Building on our previous description of the HLA-A2/A3 peptidome of β-cells, we analyzed the HLA-A3–restricted peptides targeted by circulating CD8+ T cells. Several peptides were recognized by CD8+ T cells within a narrow frequency (1–50/106), which was similar in donors with and without type 1 diabetes and harbored variable effector/memory fractions. These epitopes could be classified as conventional peptides or neoepitopes, generated either via peptide cis-splicing or mRNA splicing (e.g., secretogranin-5 [SCG5]–009). As reported for HLA-A2–restricted peptides, several epitopes originated from β-cell granule proteins (e.g., SCG3, SCG5, and urocortin-3). Similarly, H-2Kd–restricted CD8+ T cells recognizing the murine orthologs of SCG5, urocortin-3, and proconvertase-2 infiltrated the islets of NOD mice and transferred diabetes into NOD/scid recipients. The finding of granule proteins targeted in both humans and NOD mice supports their disease relevance and identifies the insulin granule as a rich source of epitopes, possibly reflecting its impaired processing in type 1 diabetes
Deciphering shell proteome within different Baltic populations of mytilid mussels illustrates important local variability and potential consequences in the context of changing marine conditions
Laboratoire Spectrométrie de masse biologique et protéomique - JaisonArivalagan, Benjamin Marie, Giovanni Chiappetta, Joëlle Vinh, Xavier Gallet, Matthieu Lebon Saloua M'Zoudi Philippe Dubois Sophie Berland Arul Marie
ELSEVIER - 745 140878 - doi.org/10.1016/j.scitotenv.2020.140878 - 2020
Molluscs defend themselves against predation and environmental stressors through the possession of mineralized shells. Mussels are widely used to predict the effects of abiotic factors such as salinity and pH on marine calcifiers in the context of changing ocean conditions. Shell matrix proteins are part of the molecular control regulating the biomineralization processes underpinning shell production. Under changing environmental conditions, differential expression of these proteins leads to the phenotypic plasticity of shells seen in many mollusc species. Low salinity decreases the availability of calcium and inorganic carbon in seawater and consequently energetic constraints often lead to thin, small and fragile shells in Mytilid mussels inhabiting Baltic Sea. To understand how the modulation of shell matrix proteins alters biomineralization, we compared the shell proteomes of mussels living under full marine conditions in the North Sea to those living in the low saline Baltic Sea. Modulation of proteins comprising the Mytilus biomineralization tool kit is observed. These data showed a relative increase in chitin related proteins, decrease in SD-rich, GA-rich shell matrix proteins indicating that altered protein scaffolding and mineral nucleation lead to impaired shell microstructures influencing shell resistance in Baltic Mytilid mussels. Interestingly, proteins with immunity domains in the shell matrix are also found to be modulated. Shell traits such as periostracum thickness, organic content and fracture resistance qualitatively correlates with the modulation of SMPs in Mytilid mussels providing key insights into control of biomineralization at molecular level in the context of changing marine conditions.
Development of Immobilized Enzyme Reactors for the characterization of the glycosylation heterogeneity of a protein
Laboratoire Spectrométrie de masse biologique et protéomique - Stan Perchepied, Nicolas Eskenazi, Chiara Giangrande, Julien Camperi, Thierry Fournier, Joëlle Vinh, Nathalie Delaunay, Valérie Pichon
ELSEVIER - 209 120568 - doi.org/10.1016/j.talanta.2019.120171 - 2020
The mapping of post-translational modifications (PTMs) of proteins can be addressed by bottom-up proteomics strategy using proteases to achieve the enzymatic digestion of the biomolecule. Glycosylation is one of the most challenging PTM to characterize due to its large structural heterogeneity. In this work, two Immobilized Enzyme Reactors (IMERs) based on trypsin and pepsin protease were used for the first time to fasten and improve the reliability of the specific mapping of the N-glycosylation heterogeneity of glycoproteins. The performance of the supports was evaluated with the digestion of human Chorionic Gonadotropin hormone (hCG), a glycoprotein characterized by four N- and four O-glycosylation sites, prior to the analysis of the digests by nanoliquid chromatography coupled to tandem mass spectrometry (nanoLC-MS/MS). Firstly, the repeatability of the nanoLC-MS/MS was evaluated and a method to control the identification of the identified glycans was developed to validate them regarding the retention time of glycopeptides in reversed phase nanoLC separation. The repeatability of the digestion with trypsin-based IMER was evaluated on the same hCG batch and on three independent batches with common located glycans up to 75%. Then, the performance of the IMER digestions was compared to in-solution digestions to evaluate the qualitative mapping of the glycosylation. It has given rise to 42 out of 45 common glycans between both digestions modes. For the first time, the complementarity of trypsin and pepsin was illustrated for the glycosylation mapping as trypsin led to identifications on 2 out of 4 glycosylation site while pepsin was informative on the 4 glycosylation site. The potential of IMERs for the study of the glycosylation of a protein was illustrated with the comparison of two hCG-based drugs, Ovitrelle® and Pregnyl
Characterization of home-made graphite/PDMS microband electrodes for amperometric detection in an original reusable glass-NOA®-PDMS electrophoretic microdevice
Laboratoire Synthèse Electrochimie Imagerie et Systèmes Analytiques - J.Gouyon F.d’Orlyé S.Griveau F.Bedioui A.Varenne
ELSEVIER - 329 135164 - doi.org/10.1016/j.electacta.2019.135164 - 2020
A new dismountable and reusable microchip for electrophoretic separation coupled to amperometric detection was developed. For this purpose, a new home made three-microbands electrode system was developed and microfabricated based on screen-printing for the inclusion of graphite/polydimethylsiloxane (C-PDMS) composite in microchannels down to 30 μm width. The composition of the composite as well as the fabrication methodology were optimized for an easy handling and an optimized electrochemical behavior. The electrochemical characterization of this composite material was first performed in bulk format (disc-shaped electrode, 6 mm diameter). It was then transposed to the micrometric scale for its integration in an original glass-NOA81®-PDMS microfluidic device allowing for reversible sealing. The microband electrodes were characterized by scanning electron microcopy and cyclic voltammetry, illustrating a good control of the microelectrode width. Then, the analytical performances of the C-PDMS composite microelectrodes were evaluated using Ru(NH3)63+ and FcMeOH as model electroactive molecules. The electrophoretic separation and quantitation of Ru(NH3)63+ were then performed in a background electrolyte made of hydrochloric acid and sodium chloride, leading to a LOD and a LOQ of 3.4 μmol L−1 and 11.3 μmol L−1, respectively. The re-openable NOA-based microdevice permits to regenerate the electrode surface by simply repositioning the microband on a new spot, allowing for robust analysis in a reusable system.
Hydrogel Matrix-Grafted Impedimetric Aptasensors for the Detection of Diclofenac
Laboratoire Synthèse Electrochimie Imagerie et Systèmes Analytiques - G. S. Kassahun, S. Griveau, S. Juillard, J. Champavert, A. Ringuedé, B. Bresson, Y. Tran*, F. Bedioui, and C. Slim
Langmuir - 36(4) 827–836 - doi.org/10.1021/acs.langmuir.9b02031 - 2020
Driven by the growing concern about the release of untreated emerging pollutants and the need for determining small amounts of these pollutants present in the environment, novel biosensors dedicated to molecular recognition are developed. We have designed biosensors using a novel class of grafted polymers, surface-attached hydrogel thin films, on conductive transducers as a biocompatible matrix for biomolecule immobilization. We showed that they can be dedicated to the molecular recognition of diclofenac (DCL). The immobilization of the aptamer onto surface-attached hydrogel thin films by covalent attachment provides a biodegradable shelter, providing the aptamer with excellent environments to preserve its active and functional structure while allowing the detection of DCL. The grafting of the aptamer is obtained using the formation of amide bonds via the activation of carboxylic acid groups of the poly(acrylic acid) hydrogel thin film. For improved sensitivity and higher stability of the sensor, a high density of the immobilized aptamer is enabled. The aptamer-modified electrode was then incubated with DCL solutions at different concentrations. The performances of the aptasensor were investigated by electrochemical impedance spectroscopy. The change in charge-transfer resistance was found to be linear with DCL concentration in the 30 pM to 1 μM range. The detection limit was calculated to be 0.02 nM. The improvement of the limit of detection can be mainly attributed to the three-dimensional environment of the hydrogel matrix which improves the grafting density of the aptamer and the affinity of the aptamer to DCL.
Mineral surfaces select for longer RNA molecules
Laboratoire Biochimie - Ryo Mizuuchi, Alex Blokhuis,b, Lena Vincent, Philippe Nghe, Niles Lehman, and David Baumd
Chem. Comm. - 55(14) 2090–2093. - 10.1039/c8cc10319d - 2019
We report empirically and theoretically that multiple prebiotic minerals can selectively accumulate longer RNAs, with selectivity enhanced at higher temperatures. We further demonstrate that surfaces can be combined with a catalytic RNA to form longer RNA polymers, supporting the potential of minerals to develop genetic information on the early Earth.
Large work extraction and the Landauer limit in a continuous Maxwell demon
Laboratoire Biochimie - Marco Ribezzi Crivellari
Nature Physics - 15(7) 93 - DOI: 10.1038/s41567-019-0481-0 - 2019
The relation between entropy and information dates back to the classical Maxwell demon paradox¹, a thought experiment proposed in 1867 by James Clerk Maxwell to violate the second law of thermodynamics. A variant of the classical Maxwell demon is the Szilard engine, proposed by Leo Szilard in 1929¹. In it, at a given time, the demon observes the compartment occupied by a single molecule in a vessel and extracts work by operating a pulley device. Here, we introduce the continuous Maxwell demon, a device capable of extracting arbitrarily large amounts of work per cycle by repeated measurements of the state of a system, and experimentally test it in single DNA hairpin pulling experiments. In the continuous Maxwell demon, the demon monitors the state of the DNA hairpin (folded or unfolded) by observing it at equally spaced time intervals, but it extracts work only when the molecule changes state. We demonstrate that the average maximum work per cycle that can be extracted by the continuous Maxwell demon is limited by the information content of the stored sequences, in agreement with the second law. Work extraction efficiency is found to be maximal in the large information-content limit where work extraction is fuelled by rare events.
High-throughput single-cell ChIP-seq identifies heterogeneity of chromatin states in breast cancer
Laboratoire Biochimie - Grosselin K1,2,3, Durand A4,5, Marsolier J, Poitou A, Marangoni E, Nemati F, Dahmani A, Lameiras S, Reyal F, Frenoy O, Pousse Y, Reichen M, Woolfe A, Brenan C, Griffiths AD, Vallot C, Gérard A.i
Nat Genet. - 51(6) 1060-1066 - doi: 10.1038/s41588-019-0424-9. - 2019
Modulation of chromatin structure via histone modification is a major epigenetic mechanism and regulator of gene expression. However, the contribution of chromatin features to tumor heterogeneity and evolution remains unknown. Here we describe a high-throughput droplet microfluidics platform to profile chromatin landscapes of thousands of cells at single-cell resolution. Using patient-derived xenograft models of acquired resistance to chemotherapy and targeted therapy in breast cancer, we found that a subset of cells within untreated drug-sensitive tumors share a common chromatin signature with resistant cells, undetectable using bulk approaches. These cells, and cells from the resistant tumors, have lost chromatin marks-H3K27me3, which is associated with stable transcriptional repression-for genes known to promote resistance to treatment. This single-cell chromatin immunoprecipitation followed by sequencing approach paves the way to study the role of chromatin heterogeneity, not just in cancer but in other diseases and healthy systems, notably during cellular differentiation and development.
Experimental evidence of symmetry breaking of transition-path times
Laboratoire Biochimie - J.Gladrow, M. Ribezzi-Crivellari, F. Ritort & U. F. Keyser
Nature Communications - 10 55 - doi.org/10.1038/s41467-018-07873-9 - 2019
While thermal rates of state transitions in classical systems have been studied for almost a century, associated transition-path times have only recently received attention. Uphill and downhill transition paths between states at different free energies should be statistically indistinguishable. Here, we systematically investigate transition-path-time symmetry and report evidence of its breakdown on the molecular- and meso-scale out of equilibrium. In automated Brownian dynamics experiments, we establish first-passage-time symmetries of colloids driven by femtoNewton forces in holographically-created optical landscapes confined within microchannels. Conversely, we show that transitions which couple in a path-dependent manner to fluctuating forces exhibit asymmetry. We reproduce this asymmetry in folding transitions of DNA-hairpins driven out of equilibrium and suggest a topological mechanism of symmetry breakdown. Our results are relevant to measurements that capture a single coordinate in a multidimensional free energy landscape, as encountered in electrophysiology and single-molecule fluorescence experiments.
Recent insights into the genotype–phenotype relationship from massively parallel genetic assays
Laboratoire Biochimie - Harry Kemble Philippe Nghe Olivier Tenaillon
Nature Physics - 9 12 - doi.org/10.1111/eva.12846 - 2019
With the molecular revolution in Biology, a mechanistic understanding of the genotype–phenotype relationship became possible. Recently, advances in DNA synthesis and sequencing have enabled the development of deep mutational scanning assays, capable of scoring comprehensive libraries of genotypes for fitness and a variety of phenotypes in massively parallel fashion. The resulting empirical genotype–fitness maps pave the way to predictive models, potentially accelerating our ability to anticipate the behaviour of pathogen and cancerous cell populations from sequencing data. Besides from cellular fitness, phenotypes of direct application in industry (e.g. enzyme activity) and medicine (e.g. antibody binding) can be quantified and even selected directly by these assays. This review discusses the technological basis of and recent developments in massively parallel genetics, along with the trends it is uncovering in the genotype–phenotype relationship (distribution of mutation effects, epistasis), their possible mechanistic bases and future directions for advancing towards the goal of predictive genetics.
Large scale control and programming of gene expression using CRISPR.
Laboratoire Biochimie - Deyell M, Ameta S, Nghe P
Semin Cell Dev Biol. - S1084-9521(18 30110-1 - doi: 10.1016/j.semcdb.2019.05.013 - 2019
The control of gene expression in cells and organisms allows to unveil gene to function relationships and to reprogram biological responses. Several systems, such as Zinc fingers, TALE (Transcription activator-like effectors), and siRNAs (small-interfering RNAs), have been exploited to achieve this. However, recent advances in Clustered Regularly Interspaced Short Palindromic Repeats and Cas9 (CRISPR-Cas9) have overshadowed them due to high specificity, compatibility with many different organisms, and design flexibility. In this review we summarize state-of-the art for CRISPR-Cas9 technology for large scale gene perturbation studies, including single gene and multiple genes knock-out, knock-down, knock-up libraries, and their associated screening assays. We feature in particular the combination of these methods with single-cell transcriptomics approaches. Finally, we highlight the application of CRISPR-Cas9 systems in building synthetic circuits that can be interfaced with gene networks to control cellular states.

583 publications.