Articles

Deciphering RNA G-quadruplex function during the early steps of HIV-1 infection
- Amrane Samir
- Jaubert Chloé
- Bedrat Amina
- Rundstadler Tiffany
- Recordon-Pinson Patricia
- Aknin Cindy
- Guédin Aurore
- de Rache Aurore
- Bartolucci Laura
- Diene Ibra
- Éric Lemoine Fréd
- Gascuel Olivier
- Pratviel Geneviève
- Mergny Jean-Louis
- Andreola Marie-Line
Nucleic Acids Research, Oxford University Press , 2022, 50 (21), pp.12328-12343 . G-quadruplexes (G4s) are four-stranded nucleic acid structures formed by the stacking of G-tetrads. Here we investigated their formation and function during HIV-1 infection. Using bioinformatics and biophysics analyses we first searched for evolutionary conserved G4-forming sequences in HIV-1 genome. We identified 10 G4s with conservation rates higher than those of HIV-1 regulatory sequences such as RRE and TAR. We then used porphyrin-based G4binders to probe the formation of the G4s during infection of human cells by native HIV-1. The G4binders efficiently inhibited HIV-1 infectivity, which is attributed to the formation of G4 structures during HIV-1 replication. Using a qRT-PCR approach, we showed that the formation of viral G4s occurs during the first 2 h post-infection and their stabilization by the G4-binders prevents initiation of reverse transcription. We also used a G4-RNA pull-down approach, based on a G4-specific biotinylated probe, to allow the direct detection and identification of viral G4-RNA in infected cells. Most of the detected G4-RNAs contain crucial regulatory elements such as the PPT and cPPT sequences as well as the U3 region. Hence, these G4s would function in the early stages of infection when the viral RNA genome is being processed for the reverse transcription step. (10.1093/nar/gkac1030)
DOI : 10.1093/nar/gkac1030
Telomeres expand sphere of influence: emerging molecular impact of telomeres in non-telomeric functions
- Vinayagamurthy Soujanya
- Bagri Sulochana
- Mergny Jean-Louis
- Chowdhury Shantanu
Trends in Genetics, Elsevier , 2022, 39 (1), pp.59-73 . Although the impact of telomeres on physiology stands well established, a question remains: how do telomeres impact cellular functions at a molecular level? This is because current understanding limits the influence of telomeres to adjacent subtelomeric regions despite the wide-ranging impact of telomeres. Emerging work in two distinct aspects offers opportunities to bridge this gap. First, telomere-binding factors were found with non-telomeric functions. Second, locally induced DNA secondary structures called G-quadruplexes are notably abundant in telomeres, and gene regulatory regions genome wide. Many telomeric factors bind to G-quadruplexes for non-telomeric functions. Here we discuss a more general model of how telomeres impact the non-telomeric genome – through factors that associate at telomeres and genome wide – and influence cell-intrinsic functions, particularly aging, cancer, and pluripotency. (10.1016/j.tig.2022.10.002)
DOI : 10.1016/j.tig.2022.10.002
Gallic Acid-Triethylene Glycol Aptadendrimers Synthesis, Biophysical Characterization and Cellular Evaluation
- Miranda André
- Lopez-Blanco Roi
- Lope-Nunes Jéssica
- Mel Ana M
- Cabral Campello Maria Pau
- Paulo António
- Oliveira Maria Cristina
- Mergny Je-Louis
- Oliveira Paula A
- Fernandez-Megia Eduardo
- Cruz Carla
Pharmaceutics, MDPI , 2022, 14 (11), pp.2456 . Herein, we describe the synthesis of an aptadendrimer by covalent bioconjugation of a gallic acid–triethylene glycol (GATG) dendrimer with the G-quadruplex (G4) AT11 aptamer (a modified version of AS1411) at the surface. We evaluated the loading and interaction of an acridine orange ligand, termed C8, that acts as an anticancer drug and binder/stabilizer of the G4 structure of AT11. Dynamic light scattering experiments demonstrated that the aptadendrimer was approximately 3.1 nm in diameter. Both steady-state and time-resolved fluorescence anisotropy evidenced the interaction between the aptadendrimer and C8. Additionally, we demonstrated that the iodine atom of the C8 ligand acts as an effective intramolecular quencher in solution, while upon complexation with the aptadendrimer, it adopts a more extended conformation. Docking studies support this conclusion. Release experiments show a delivery of C8 after 4 h. The aptadendrimers tend to localize in the cytoplasm of various cell lines studied as demonstrated by confocal microscopy. The internalization of the aptadendrimers is not nucleolin-mediated or by passive diffusion, but via endocytosis. MTT studies with prostate cancer cells and non-malignant cells evidenced high cytotoxicity mainly due to the C8 ligand. The rapid internalization of the aptadendrimers and the fluorescence properties make them attractive for the development of potential nanocarriers. (10.3390/pharmaceutics14112456)
DOI : 10.3390/pharmaceutics14112456
Design, Synthesis, and Antiprotozoal Evaluation of New Promising 2,9-Bis[(substituted-aminomethyl)]-4,7-phenyl-1,10phenanthroline Derivatives, a Potential Alternative Scaffold to Drug Efflux
- Guillon Jean
- Cohen Anita
- Boudot Clotilde
- Monic Sarah
- Savrimoutou Solène
- Moreau Stéphane
- Albenque-Rubio Sandra
- Lafon-Schmaltz Camille
- Dassonville-Klimpt Alexandra
- Mergny Jean-Louis
- Ronga Luisa
- Bernabeu de Maria Mikel
- Lamarche Jérémy
- Dal Lago Cristina
- Largy Eric
- Gabelica Valérie
- Moukha Serge
- Dozolme Pascale
- Agnamey Patrice
- Azas Nadine
- Mullié Catherine
- Courtioux Bertrand
- Sonnet Pascal
Pathogens, MDPI , 2022, 11 (11), pp.1339 . A series of novel 2,9-bis[(substituted-aminomethyl)]-4,7-phenyl-1,10-phenanthroline derivatives was designed, synthesized, and evaluated in vitro against three protozoan parasites (Plasmodium falciparum, Leishmania donovani and Trypanosoma brucei brucei). Pharmacological results showed antiprotozoal activity with IC50 values in the sub and μM range. In addition, the in vitro cytotoxicity of these original molecules was assessed with human HepG2 cells. The substituted diphenylphenanthroline 1l was identified as the most potent antimalarial derivative with a ratio of cytotoxic to antiparasitic activities of 505.7 against the P. falciparum CQ-resistant strain W2. Against the promastigote forms of L. donovani, the phenanthrolines 1h, 1j, 1n and 1o were the most active with IC50 from 2.52 to 4.50 μM. The phenanthroline derivative 1o was also identified as the most potent trypanosomal candidate with a selectivity index (SI) of 91 on T. brucei brucei strain. FRET melting and native mass spectrometry experiments evidenced that the nitrogen heterocyclic derivatives bind the telomeric G-quadruplexes of P. falciparum and Trypanosoma. Moreover, as the telomeres of the parasites P. falciparum and Trypanosoma could be considered to be possible targets of this kind of nitrogen heterocyclic derivatives, their potential ability to stabilize the parasitic telomeric G-quadruplexes have been determined through the FRET melting assay and by native mass spectrometry. (10.3390/pathogens11111339)
DOI : 10.3390/pathogens11111339
Synthesis and evaluation of 2,9-disubstituted-1,10-phenanthroline derivatives as G-quadruplex binders
- Figueiredo Joana
- Carreira-Barral Israel
- Quesada Roberto
- Mergny Jean-Louis
- Cruz Carla
Bioorganic and Medicinal Chemistry Letters, Elsevier , 2022, 73, pp.116971 . G-quadruplex (G4) structures are non-canonical DNA/RNA secondary structures able to form within guanine rich nucleic acids sequences. They are present in several regions of the human genome including gene promoters, untranslated sequences, and telomeres. Due to their biological relevance G4 structures are considered important drug targets, in particular for anticancer therapies, leading to the development of G4 stabilizing small molecules. Telomeric regions have received special attention in this field since they can fold into several distinct intramolecular G-quadruplexes topologies. Herein, we report the synthesis of 2,9-disubstituted-1,10-phenanthroline derivatives and their ability to stabilize different intramolecular telomeric G4 sequences. We evaluated ligand-induced stabilization, selectivity and specificity of ligands using Förster Resonance Energy Transfer (FRET) melting experiments and circular dichroism (CD). In addition, we assessed the cytotoxicity of ligands against two cancer cell lines (A549 and H1299) and one healthy cell line (NHDF). (10.1016/j.bmc.2022.116971)
DOI : 10.1016/j.bmc.2022.116971
Full-Field Optical Coherence Microscopy for Histology-Like Analysis of Stromal Features in Corneal Grafts
- Irsch Kristina
- Grieve Kate
- Borderie Marie
- Vilbert Maëlle
- Plamann Karsten
- Ghoubay Djida
- Georgeon Cristina
- Borderie Vincent
Journal of visualized experiments : JoVE, JoVE , 2022, 18 (188), pp.e0291613 . Corneal transparency is essential to provide a clear view into and out of the eye, yet clinical means to assess such transparency are extremely limited and usually involve a subjective grading of visible opacities by means of slit-lamp biomicroscopy. Here, we describe an automated algorithm allowing extraction of quantitative corneal transparency parameters with standard clinical spectral-domain optical coherence tomography (SD-OCT). Our algorithm employs a novel pre-processing procedure to standardize SD-OCT image analysis and to numerically correct common instrumental artifacts before extracting mean intensity stromal-depth ( z ) profiles over a 6-mm-wide corneal area. The z -profiles are analyzed using our previously developed objective method that derives quantitative transparency parameters directly related to the physics of light propagation in tissues. Tissular heterogeneity is quantified by the Birge ratio B r and the photon mean-free path ( l s ) is determined for homogeneous tissues (i.e., B r ~1 ). SD-OCT images of 83 normal corneas (ages 22–50 years) from a standard SD-OCT device (RTVue-XR Avanti, Optovue Inc.) were processed to establish a normative dataset of transparency values. After confirming stromal homogeneity ( B r <10), we measured a median l s of 570 μm (interdecile range: 270–2400 μm). By also considering corneal thicknesses, this may be translated into a median fraction of transmitted (coherent) light T coh(stroma) of 51% (interdecile range: 22–83%). Excluding images with central saturation artifact raised our median T coh(stroma) to 73% (interdecile range: 34–84%). These transparency values are slightly lower than those previously reported, which we attribute to the detection configuration of SD-OCT with a relatively small and selective acceptance angle. No statistically significant correlation between transparency and age or thickness was found. In conclusion, our algorithm provides robust and quantitative measurements of corneal transparency from standard SD-OCT images with sufficient quality (such as ‘Line’ and ‘CrossLine’ B-scan modes without central saturation artifact) and addresses the demand for such an objective means in the clinical setting. (10.3791/57104)
DOI : 10.3791/57104
G-Quadruplex Aptamer-Ligand Characterization
- Moreira David
- Leitão Daniela
- Lopes-Nunes Jéssica
- Santos Tiago
- Figueiredo Joana
- Miranda André
- Alexandre Daniela
- Tomaz Cândida
- Mergny Jean-Louis
- Cruz Carla
Molecules, MDPI , 2022, 27 (20), pp.6781 . In this work we explore the structure of a G-rich DNA aptamer termed AT11-L2 (TGGTGGTGGTTGTTGTTGGTGGTGGTGGT; derivative of AT11) by evaluating the formation and stability of G-quadruplex (G4) conformation under different experimental conditions such as KCl concentration, temperature, and upon binding with a variety of G4 ligands (360A, BRACO-19, PDS, PhenDC3, TMPyP4). We also determined whether nucleolin (NCL) can be a target of AT11-L2 G4. Firstly, we assessed by circular dichroism, UV and NMR spectroscopies the formation of G4 by AT11-L2. We observed that, for KCl concentrations of 65 mM or less, AT11-L2 adopts hybrid or multiple topologies. In contrast, a parallel topology predominates for buffer containing 100 mM of KCl. The Tm of AT11-L2 in 100 mM of KCl is 38.9 °C, proving the weak stability of this sequence. We also found that upon titration with two molar equivalents of 360A, BRACO-19 and PhenDC3, the G4 is strongly stabilized and its topology is maintained, while the addition of 3.5 molar equivalents of TMPyP4 promotes the disruption of G4. The KD values between AT11-L2 G4, ligands and NCL were obtained by fluorescence titrations and are in the range of µM for ligand complexes and nM when adding NCL. In silico studies suggest that four ligands bind to the AT11-L2 G4 structure by stacking interactions, while the RBD1,2 domains of NCL interact preferentially with the thymines of AT11-L2 G4. Finally, AT11-L2 G4 co-localized with NCL in NCL-positive tongue squamous cell carcinoma cell line. (10.3390/molecules27206781)
DOI : 10.3390/molecules27206781
Coordinated metabolic transitions and gene expression by NAD+ during adipogenesis
- Sánchez-Ramírez Edgar
- Ung Thi Phuong Lien
- Alarcón del Carmen Alejandro
- del Toro-Ríos Ximena
- Fajardo-Orduña Guadalupe R
- Noriega Lilia G
- Cortés-Morales Victor A
- Tovar Armando R
- Montesinos Juan José
- Orozco-Solís Ricardo
- Stringari Chiara
- Aguilar-Arnal Lorena
Journal of Cell Biology, Rockefeller University Press , 2022, 221 (12) . Adipocytes are the main cell type in adipose tissue, which is a critical regulator of metabolism, highly specialized in storing energy as fat. Adipocytes differentiate from multipotent mesenchymal stromal cells (hMSCs) through adipogenesis, a tightly controlled differentiation process involving close interplay between metabolic transitions and sequential programs of gene expression. However, the specific gears driving this interplay remain largely obscure. Additionally, the metabolite nicotinamide adenine dinucleotide (NAD +) is becoming increasingly recognized as a regulator of lipid metabolism, and a promising therapeutic target for dyslipidemia and obesity. Here, we explored how NAD + bioavailability controls adipogenic differentiation from hMSC. We found a previously unappreciated repressive role for NAD + on adipocyte commitment, while a functional NAD +-dependent deacetylase SIRT1 appeared crucial for terminal differentiation of pre-adipocytes. Repressing NAD + biosynthesis during adipogenesis promoted the adipogenic transcriptional program, while two-photon microscopy and extracellular flux analyses suggest that SIRT1 activity mostly relies on the metabolic switch. Interestingly, SIRT1 controls subcellular compartmentalization of redox metabolism during adipogenesis. (10.1083/jcb.202111137)
DOI : 10.1083/jcb.202111137
Synthesis and structure–activity relationship studies of pyrido [1,2-e]purine-2,4(1H,3H)-dione derivatives targeting Flavin-Dependent $Thymidylate\ Synthase$ in $Mycobacterium\ tuberculosis$
- Biteau Nicolas G.
- Roy Vincent
- Nicolas Cyril
- Becker Hubert F
- Lambry Jean-Christophe
- Myllykallio Hannu
- Agrofoglio Luigi A.
Molecules, MDPI , 2022, 27, pp.6216 . In 2002, a new class of thymidylate synthase (TS) involved in the de novo synthesis of dTMP named Flavin-Dependent Thymidylate Synthase (FDTS) encoded by the thyX gene was discovered; FDTS is present only in 30% of prokaryote pathogens and not in human pathogens, which makes it an attractive target for the development of new antibacterial agents, especially against multi-resistant pathogens. We report herein the synthesis and structure-activity relationship of a novel series of hitherto unknown pyrido[1,2-E]purine-2,4(1H,3H)-dione analogues. Several synthetics efforts were done to optimize regioselective N1-alkylation through organopalladium cross-coupling. Modelling of potential hits were performed to generate a model of interaction into the active pocket of FDTS to understand and guide further synthetic modification. All those compounds were evaluated on an in-house in vitro NADPH oxidase assays screening as well as against Mycobacterium tuberculosis ThyX. The highest inhibition was obtained for compound 23a with 84.3% at 200 μM without significant cytotoxicity (CC50 > 100 micromolar) on PBM cells. (10.3390/molecules27196216)
DOI : 10.3390/molecules27196216
Parametrization of Force Field Bonded Terms under Structural Inconsistency
- Croitoru Anastasia
- Aleksandrov Alexey
Journal of Chemical Information and Modeling, American Chemical Society , 2022, pp.4471-4482 . (10.1021/acs.jcim.2c00950)
DOI : 10.1021/acs.jcim.2c00950
Iso-FRET: an isothermal competition assay to analyze quadruplex formation in vitro
- Luo Yu
- Verga Daniela
- Mergny Jean-Louis
Nucleic Acids Research, Oxford University Press , 2022, 50 (16), pp.e93-e93 . Algorithms have been widely used to predict G-quadruplexes (G4s)-prone sequences. However, an experimental validation of these predictions is generally required. We previously reported a high-throughput technique to evidence G4 formation in vitro called FRET-MC. This method, while convenient and reproducible, has one known weakness: its inability to pin point G4 motifs of low thermal stability. As such quadruplexes may still be biologically relevant if formed at physiological temperature, we wanted to develop an independent assay to overcome this limitation. To this aim, we introduced an isothermal version of the competition assay, called iso-FRET, based on a duplex-quadruplex competition and a well-characterized bis-quinolinium G4 ligand, PhenDC3. G4-forming competitors act as decoys for PhenDC3, lowering its ability to stabilize the G4-forming motif reporter oligonucleotide conjugated to a fluorescence quencher (37Q). The decrease in available G4 ligand concentration restores the ability of 37Q to hybridize to its FAM-labeled short complementary C-rich strand (F22), leading to a decrease in fluorescence signal. In contrast, when no G4-forming competitor is present, PhenDC3 remains available to stabilize the 37Q quadruplex, preventing the formation of the F22 + 37Q complex. Iso-FRET was first applied to a reference panel of 70 sequences, and then used to investigate 23 different viral sequences. (10.1093/nar/gkac465)
DOI : 10.1093/nar/gkac465
Sol-gel transition induced by alumina nanoparticles in a model pulmonary surfactant
- Berret Jean-François
- Mousseau Fanny
- Le Borgne Rémi
- Oikonomou Evdokia K
Colloids and Surfaces A: Physicochemical and Engineering Aspects, Elsevier , 2022, 646, pp.128974 . Inhaled airborne particles smaller than 100 nm entering the airways have been shown to deposit in significant amount in the alveolar region of the lungs. The interior of the alveoli is covered with a ~ 1 µm thick lining fluid, called pulmonary surfactant. Inhaled nanoparticles are susceptible to interact with the lung fluid and modify pulmonary functions. Here we evaluate the structural and rheological properties of the pulmonary surfactant substitute Curosurf® which is administered to premature babies for the treatment of respiratory distress syndrome. Curosurf® is considered a reliable model of endogenous pulmonary surfactant in terms of composition, structure and function. Using active microrheology based on magnetically actuated wires, we find that Curosurf® dispersions exhibit a Newtonian behavior at lipid concentration from 0 to 80 g L-1 , and that the viscosity follows the Krieger-Dougherty law observed for a wide variety of colloids. Upon addition of 40 nm alumina nanoplatelets, a significant change of the Curosurf® rheology is noticed. The dispersions then enter a soft solid phase characterized by an infinite viscosity and a non-zero equilibrium elastic modulus. The sol-gel transition induced by the nanoparticles is interpreted as the result of the alumina/vesicle interaction, which are illustrated by transmission electron microscopy. It also suggests a potential toxicity associated with the modification of the lung fluid structural and dynamical properties. (10.1016/j.colsurfa.2022.128974)
DOI : 10.1016/j.colsurfa.2022.128974
The Newly Sequenced Genome of Pisum sativum Is Replete with Potential G-Quadruplex-Forming Sequences—Implications for Evolution and Biological Regulation
- Dobrovolná Michaela
- Bohálová Natália
- Peška Vratislav
- Wang Jiawei
- Luo Yu
- Bartas Martin
- Volná Adriana
- Mergny Jean-Louis
- Brázda Václav
International Journal of Molecular Sciences, MDPI , 2022, 23 (15), pp.8482 . G-quadruplexes (G4s) have been long considered rare and physiologically unimportant in vitro curiosities, but recent methodological advances have proved their presence and functions in vivo. Moreover, in addition to their functional relevance in bacteria and animals, including humans, their importance has been recently demonstrated in evolutionarily distinct plant species. In this study, we analyzed the genome of Pisum sativum (garden pea, or the so-called green pea), a unique member of the Fabaceae family. Our results showed that this genome contained putative G4 sequences (PQSs). Interestingly, these PQSs were located nonrandomly in the nuclear genome. We also found PQSs in mitochondrial (mt) and chloroplast (cp) DNA, and we experimentally confirmed G4 formation for sequences found in these two organelles. The frequency of PQSs for nuclear DNA was 0.42 PQSs per thousand base pairs (kbp), in the same range as for cpDNA (0.53/kbp), but significantly lower than what was found for mitochondrial DNA (1.58/kbp). In the nuclear genome, PQSs were mainly associated with regulatory regions, including 5 UTRs, and upstream of the rRNA region. In contrast to genomic DNA, PQSs were located around RNA genes in cpDNA and mtDNA. Interestingly, PQSs were also associated with specific transposable elements such as TIR and LTR and around them, pointing to their role in their spreading in nuclear DNA. The nonrandom localization of PQSs uncovered their evolutionary and functional significance in the Pisum sativum genome. (10.3390/ijms23158482)
DOI : 10.3390/ijms23158482
The Newly Sequenced Genome of Pisum sativum Is Replete with Potential G-Quadruplex-Forming Sequences—Implications for Evolution and Biological Regulation
- Dobrovolná Michaela
- Bohálová Natália
- Peška Vratislav
- Wang Jiawei
- Luo Yu
- Bartas Martin
- Volná Adriana
- Mergny Jean-Louis
- Brázda Václav
International Journal of Molecular Sciences, MDPI , 2022, 23 (15), pp.8482 .
nAdder: A scale-space approach for the 3D analysis of neuronal traces
- Phan Minh Son
- Matho Katherine
- Beaurepaire Emmanuel
- Livet Jean
- Chessel Anatole
PLoS Computational Biology, PLOS , 2022, 18 (7), pp.e1010211 . Tridimensional microscopy and algorithms for automated segmentation and tracing are revolutionizing neuroscience through the generation of growing libraries of neuron reconstructions. Innovative computational methods are needed to analyze these neuronal traces. In particular, means to characterize the geometric properties of traced neurites along their trajectory have been lacking. Here, we propose a local tridimensional (3D) scale metric derived from differential geometry, measuring for each point of a curve the characteristic length where it is fully 3D as opposed to being embedded in a 2D plane or 1D line. The larger this metric is and the more complex the local 3D loops and turns of the curve are. Available through the GeNePy3D open-source Python quantitative geometry library ( https://genepy3d.gitlab.io ), this approach termed nAdder offers new means of describing and comparing axonal and dendritic arbors. We validate this metric on simulated and real traces. By reanalysing a published zebrafish larva whole brain dataset, we show its ability to characterize different population of commissural axons, distinguish afferent connections to a target region and differentiate portions of axons and dendrites according to their behavior, shedding new light on the stereotypical nature of neurites’ local geometry. (10.1371/journal.pcbi.1010211)
DOI : 10.1371/journal.pcbi.1010211
Construire un système d’imagerie 3D de tomographie par cohérence optique (OCT)
- Latour Gaël
Photoniques, EDP Sciences , 2022 (114), pp.21-25 . Les étudiants ont la fierté de construire - de toute pièce - un microscope produisant des images tridimensionnelles (3D) d’objets transparents ou faiblement diffusants. Le projet laisse le temps aux étudiants de progresser en toute autonomie et de surmonter les difficultés théoriques et expérimentales rencontrées. Leur rigueur et leur ténacité sont récompensées par la beauté des films 3D qu’ils réalisent avec leur propre microscope. (10.1051/photon/202111421)
DOI : 10.1051/photon/202111421
G-quadruplex structure of the C. elegans telomeric repeat: a two tetrads basket type conformation stabilized by a non-canonical C–T base-pair
- Marquevielle Julien
- de Rache Aurore
- Vialet Brune
- Morvan Estelle
- Mergny Jean-Louis
- Amrane Samir
Nucleic Acids Research, Oxford University Press , 2022, 50 (12), pp.7134-7146 . The Caenorhabditis elegans model has greatly contributed to the understanding of the role of G-quadruplexes in genomic instability. The GGCTTA repeats of the C. elegans telomeres resemble the GGGTTA repeats of the human telomeres. However, the comparison of telomeric sequences (Homo sapiens, Tetrahymena, Oxytricha, Bombyx mori and Giardia) revealed that small changes in these repeats can drastically change the topology of the folded G-quadruplex. In the present work we determined the structure adopted by the C. elegans telomeric sequence d[GG(CTTAGG)3]. The investigated C. elegans telomeric sequence is shown to fold into an intramolecular two G-tetrads basket type G-quadruplex structure that includes a C–T base pair in the diagonal loop. This work sheds light on the telomeric structure of the widely used C. elegans animal model. (10.1093/nar/gkac523)
DOI : 10.1093/nar/gkac523
Photoswitching Behavior of Flavin–Inhibitor Complex in a Nonphotocatalytic Flavoenzyme
- Zhuang Bo
- Vos Marten
Journal of the American Chemical Society, American Chemical Society , 2022, 144, pp.11569-11573 . (10.1021/jacs.2c04763)
DOI : 10.1021/jacs.2c04763
Spatial distribution of air bubbles created by an imping water jet into a free water surface
- Bense Alexandre
- Lizee Mathieu
- Guillet Thibault
- Gallot Guilhem
Emergent Scientist, EDP Sciences , 2022, 6 (3), pp.7 . In this paper we study the rise to the surface, radial motion and disappearance of the bubbles created by a water jet pouring into a container, in particular their density at the surface as a function of the impact velocity. We first focus on their emergence radius at the surface which follows a log-normal distribution. Next, we establish experimentally a law relating the bubble velocity to the distance to the jet. We also investigate their disappearance, caused at low density mainly by explosion, and at high density predominantly by coalescence. Finally, we build an accurate model for the density of bubbles. (10.1051/emsci/2022001)
DOI : 10.1051/emsci/2022001
Conflits environnementaux en Afrique sahélienne : regards croisés sur les aires protégées sahéliennes camerounaises
- Dzokom Alexis
Sahel Nature Consulting Revue, Sahel Nature Consulting Revue (March 2025) , 2022, 6 (1), pp.1-13 . <div><p>En Afrique sahélienne, les aires protégées jouent un rôle central dans la conservation de la biodiversité. Cependant, leur mise en oeuvre suscite de nombreux conflits entre les communautés locales et les gestionnaires de l'environnement. Cette étude analyse les types, causes et conséquences des conflits environnementaux liés aux aires protégées sahéliennes camerounaises, en particulier dans le Parc National de Waza, la Réserve de Kalamaloué et les zones de chasse de l'Extrême-Nord. À partir d'une méthodologie qualitative basée sur des enquêtes de terrain, des entretiens semi-directifs et une revue documentaire, nous montrons que les conflits intercommunautaires (5,2 ±1,2) sont souvent exacerbés par l'exclusion des populations des ressources vitales(8,7 ±1,0), à l'inadéquation des politiques de conservation (7,9±1,4), et à l'influence des dynamiques climatiques et sécuritaires. L'analyse révèle aussi l'émergence d'initiatives communautaires et de mécanismes de gestion participative (5,9±1,9) encore embryonnaires. Les résultats soulignent la nécessité d'une gouvernance environnementale inclusive, tenant compte des réalités locales et des enjeux géopolitiques sahéliens.</p></div>
High-throughput tuning of ovarian cancer spheroids for on-chip invasion assays
- Chen Changchong
- He Yong
- Lopez Elliot
- Carreiras Franck
- Yamada Ayako
- Schanne-Klein Marie-Claire
- Lambert Ambroise
- Chen Yong
- Aimé Carole
Micro and Nano Engineering, Elsevier , 2022, 15, pp.100138 . We developed an invasion assay by using microfabricated culture devices. First, ovarian tumor spheroids were generated with a culture patch device consisting of an agarose membrane formed with a honeycomb microframe – the patch – and gelatin nanofiber backbone. By changing the dimensions of the honeycomb compartments we were able to control the number of cells and size of the spheroids. When the spheroids were placed on a patch coated with a thin membrane of fibrillary type I collagen, spheroid disruption was observed due to substrate induced cell migration. This process is straightforward and should be applicable to other cancer types, as well as assays under microfluidic conditions, thereby holding the potential for use in tumor modeling and anti-cancer drug development. (10.1016/j.mne.2022.100138)
DOI : 10.1016/j.mne.2022.100138
Erratum: Electric field measurements in plasmas: how focusing strongly distorts the E-FISH signal (2020 Plasma Sources Sci. Technol. 29 125002)
- Chng Tat Loon
- Starikovskaia Svetlana
- Schanne-Klein Marie-Claire
Plasma Sources Science and Technology, IOP Publishing , 2022, 31 (5), pp.059601 . (10.1088/1361-6595/ac601d)
DOI : 10.1088/1361-6595/ac601d
Efficient Biocatalytic System for Biosensing by Combining Metal–Organic Framework (MOF)-Based Nanozymes and G-Quadruplex (G4)-DNAzymes
- Mao Xuanxiang
- He Fangni
- Qiu Dehui
- Wei Shijiong
- Luo Rengan
- Chen Yun
- Zhang Xiaobo
- Lei Jianping
- Monchaud David
- Mergny Jean-Louis
- Ju Huangxian
- Zhou Jun
Analytical Chemistry, American Chemical Society , 2022, 94 (20), pp.7295-7302 . A high catalytic efficiency associated with a robust chemical structure are among the ultimate goals when developing new biocatalytic systems for biosensing applications. To get ever closer to these goals, we report here on a combination of metal-organic framework (MOF)-based nanozymes and a G-quadruplex (G4)-based catalytic system known as G4-DNAzyme. This approach aims at combining the advantages of both partners (chiefly, the robustness of the former and the modularity of the latter). To this end, we used MIL-53(Fe) MOF and linked it covalently to a G4-forming sequence (F3TC), itself covalently linked to its cofactor hemin. The resulting complex (referred to as MIL-53(Fe)/G4-hemin) exhibited exquisite peroxidase-mimicking oxidation activity and an excellent robustness (being stored in water for weeks). These properties were exploited to devise a new biosensing system based on a cascade of reactions catalyzed by the nanozyme (ABTS oxidation) and an enzyme, the alkaline phosphatase (or ALP, ascorbic acid 2-phosphate dephosphorylation). The product of the latter poisoning the former, we thus designed a biosensor for ALP (a marker of bone diseases and cancers), with a very low limit of detection (LOD, 0.02 U L-1), which is operative in human plasma samples. (10.1021/acs.analchem.2c00600)
DOI : 10.1021/acs.analchem.2c00600
Stabilization of a DNA aptamer by ligand binding
- Santos Tiago
- Lopes-Nunes Jéssica
- Alexandre Daniela
- Miranda André
- Figueiredo Joana
- Silva Micael
- Mergny Jean-Louis
- Cruz Carla
Biochimie, Elsevier , 2022, S0300-9084 (22), pp.00114-6 . G-rich aptamers such as AS1411 are small oligonucleotides that present several benefits comparatively to monoclonal antibodies, since they are easier to manufacture and store, have small size and do not stimulate an immune response. We analyzed AT11-B1, a modified sequence of AT11 (itself a modified version of AS1411), in which one thymine was removed from the bulge region. We studied G-quadruplex (G4) formation/stabilization using PhenDC3, PDS, BRACO-19, TMPyP4 and 360A ligands by different biophysical techniques, namely circular dichroism (CD), Förster resonance energy transfer (FRET-melting) and nuclear magnetic resonance (NMR). The CD spectra showed that AT11-B1 adopts a predominant G4 of parallel topology when the buffer contains KCl or when ligands are added. PhenDC3 induced a ΔTm of 30 °C or more of the G4 structure as shown by CD- and FRET-melting experiments. The ligands demonstrate high affinity for AT11-B1 G4 and the NMR studies revealed that the AT11-B1 G4 involves four G-tetrad layers. The in silico studies suggest that all ligands bind AT11-B1 G4, namely, by stacking interactions, except PDS that may bind to the loop/groove interface. In addition, molecular dynamics simulations revealed that nucleolin (NCL) interacts with the AT11-B1 G4 structure through the RNA binding domain (RBD) 2 and the 12-residue linker between RBD1,2. Moreover, AT11-B1 G4 was internalized into a NCL-positive tongue squamous cell carcinoma cell line. In a nutshell, this study may help the identification of the ligands scaffolds to bind and stabilize AT11-B1, improving the targeting towards NCL that is overexpressed in cancer cells. (10.1016/j.biochi.2022.05.002)
DOI : 10.1016/j.biochi.2022.05.002
Molecular insights into the role of heme in the transcriptional regulatory system AppA/PpsR.
- Kapetanaki Sofia
- Fekete Zsuzsanna
- Dorlet Pierre
- Vos Marten
- Liebl Ursula
- Lukacs Andras
Biophysical Journal, Biophysical Society , 2022, 121, pp.2135-2151 . Heme has been shown to have a crucial role in the signal transduction mechanism of the facultative photoheterotrophic bacterium Rhodobacter sphaeroides. It interacts with the transcriptional regulatory complex AppA/PpsR, in which AppA and PpsR function as the antirepressor and repressor, respectively, of photosynthesis gene expression. The mechanism, however, of this interaction remains incompletely understood. In this study, we combined electron paramagnetic resonance (EPR) spectroscopy and Förster resonance energy transfer (FRET) to demonstrate the ligation of heme in PpsR with a proposed cysteine residue. We show that heme binding in AppA affects the fluorescent properties of the dark-adapted state of the protein, suggesting a less constrained flavin environment compared with the absence of heme and the light-adapted state. We performed ultrafast transient absorption measurements in order to reveal potential differences in the dynamic processes in the full-length AppA and its heme-binding domain alone. Comparison of the CO-binding dynamics demonstrates a more open heme pocket in the holo-protein, qualitatively similar to what has been observed in the CO sensor RcoM-2, and suggests a communication path between the blue-light-using flavin (BLUF) and sensing containing heme instead of cobalamin (SCHIC) domains of AppA. We have also examined quantitatively the affinity of PpsR to bind to individual DNA fragments of the puc promoter using fluorescence anisotropy assays. We conclude that oligomerization of PpsR is initially triggered by binding of one of the two DNA fragments and observe a ∼10-fold increase in the dissociation constant Kd for DNA binding upon heme binding to PpsR. Our study provides significant new insight at the molecular level on the regulatory role of heme that modulates the complex transcriptional regulation in R. sphaeroides and supports the two levels of heme signaling, via its binding to AppA and PpsR and via the sensing of gases like oxygen. (10.1016/j.bpj.2022.04.031)
DOI : 10.1016/j.bpj.2022.04.031