Audrey Hessel

Institut de Pharmacologie et de Biologie Structurale

Author Statistics

Papers

Citation

H-Index

i-10 index

Research Trends

Author Order

Document Type

Co-Authors

Paul Schanda

Institute of Science and Technology Austria

Daniel Ladant

Institut Pasteur

Orso Subrini

Institut Pasteur

Alexandre Chenal

Centre National de la Recherche Scientifique

Miriam Pinilla

Institut de Pharmacologie et de Biologie Structurale

Karin Santoni

Institut de Pharmacologie et de Biologie Structurale

Katharina Weinhäupl

Institut de Biologie Structurale

Beate Bersch

Institut de Biologie Structurale

Céline Cougoule

Institut de Pharmacologie et de Biologie Structurale

ÉM

Étienne Meunier

Institut de Pharmacologie et de Biologie Structurale

Cooperative Institutions

Centre National de la Recherche Scientifique

161

Inserm

103

Université Paris Cité

Institut Pasteur

Assistance Publique – Hôpitaux de Paris

Université de Toulouse

Université Toulouse III - Paul Sabatier

Institut de Pharmacologie et de Biologie Structurale

Sorbonne Université

Commissariat à l'Énergie Atomique et aux Énergies Alternatives

Author Statistics

Papers

Citation

H-Index

i-10 index

Research Field

Human NLRP1 Is a Sensor of 3CL Proteases from Pathogenic Coronaviruses in Lung Epithelial Cells

SSRN Electronic Journal (2021)

Rémi Planès Miriam Pinilla Karin Santoni Audrey Hessel Kenneth Lay

Inflammation observed in SARS-CoV-2-infected patients suggests that inflammasomes, proinflammatory intracellular complexes, regulate various steps of infection. Lung epithelial cells express inflammasome-forming sensors and constitute the primary entry door of SARS-CoV-2. Here, we describe that the NLRP1 inflammasome detects SARS-CoV-2 infection in human lung epithelial cells. Specifically, human NLRP1 is cleaved at the Q333 site by multiple coronavirus 3CL proteases, which triggers inflammasome assembly, cell death and limits the production of infectious viral particles. Analysis of NLRP1-associated pathways unveils that 3CL proteases also cleave and inactivate the pyroptosis executioner Gasdermin (GSDM)-D. Consequently, Caspase-3 and GSDM-E promote alternative cell pyroptosis, a process exacerbated in cells exhibiting imparied type I interferon production. Finally, analysis of pyroptosis markers in plasma from COVID-19 patients with characterized severe pneumonia due to Interferon alterations identify GSDM-E/Caspase-3 as biological markers of disease severity. Overall, our findings identify NLRP1 as a key sensor of SARS-CoV-2 infection in lung epithelia.Funding Information: This project has been funded on lab own funds from unrelated grants from the Fondation pour la Recherche Médicale (FRM) and ERC StG (INFLAME) to EM, from ERC StG (ANTIViR) to CG, by the French Ministry of Health with the participation of the Groupement Interrégional de Recherche Clinique et d'Innovation Sud-Ouest Outre-Mer (PHRCI 2020 IMMUNOMARK-COV) to G-M.B. The ASB3 structure is supported by LABEX, Investissement d'Avenir and foundation Bettencourt grants to ON. MP and RP were respectively funded by a CIFRE PhD fellowship and a research grant from Invivogen. SB is supported by a PhD fellowship from Mali ministry of education and from the FRM (FDT 12794). SALC is supported by a Vaincre La Mucoviscidose (VLM) PhD fellowship.Declaration of Interests: Authors declare no conflict of interest. Ethics Approval Statement: All donors had given written informed consent and the study was approved by the ethical review board "Comité de Protection des Personnes Est-III" (ID-RCB 2020-A01292-37).

Pyroptosis

Proinflammatory cytokine

NLRP1

10.2139/ssrn.3948202

Cite

Citations (0)

Immunogenicity of the Plasmodium falciparum PfEMP1-VarO Adhesin: Induction of Surface-Reactive and Rosette-Disrupting Antibodies to VarO Infected Erythrocytes

PLoS ONE (2015)

Micheline Guillotte Alexandre Juillerat Sébastien Igonet Audrey Hessel Stéphane Pêtres

Adhesion of Plasmodium falciparum-infected red blood cells (iRBC) to human erythrocytes (i.e. rosetting) is associated with severe malaria. Rosetting results from interactions between a subset of variant PfEMP1 (Plasmodium falciparum erythrocyte membrane protein 1) adhesins and specific erythrocyte receptors. Interfering with such interactions is considered a promising intervention against severe malaria. To evaluate the feasibility of a vaccine strategy targetting rosetting, we have used here the Palo Alto 89F5 VarO rosetting model. PfEMP1-VarO consists of five Duffy-Binding Like domains (DBL1-5) and one Cysteine-rich Interdomain Region (CIDR1). The binding domain has been mapped to DBL1 and the ABO blood group was identified as the erythrocyte receptor. Here, we study the immunogenicity of all six recombinant PfEMP1-VarO domains and the DBL1- CIDR1 Head domain in BALB/c and outbred OF1 mice. Five readouts of antibody responses are explored: ELISA titres on the recombinant antigen, VarO-iRBC immunoblot reactivity, VarO-iRBC surface-reactivity, capacity to disrupt VarO rosettes and the capacity to prevent VarO rosette formation. For three domains, we explore influence of the expression system on antigenicity and immunogenicity. We show that correctly folded PfEMP1 domains elicit high antibody titres and induce a homogeneous response in outbred and BALB/c mice after three injections. High levels of rosette-disrupting and rosette-preventing antibodies are induced by DBL1 and the Head domain. Reduced-alkylated or denatured proteins fail to induce surface-reacting and rosette-disrupting antibodies, indicating that surface epitopes are conformational. We also report limited cross-reactivity between some PfEMP1 VarO domains. These results highlight the high immunogenicity of the individual domains in outbred animals and provide a strong basis for a rational vaccination strategy targeting rosetting.

Antigenicity

Rosette (schizont appearance)

10.1371/journal.pone.0134292

Cite

Citations (4)

Biophysical investigations of the adenylate cyclase (CyaA) toxin from Bordetella pertussis

Toxicon (2016)

Sara E. Cannella Véronique Yvette Ntsogo Enguéné Johanna C. Karst Audrey Hessel Orso Subrini

Adenylate Cyclase Toxin

Bordetella

10.1016/j.toxicon.2016.01.030

Cite

Citations (0)

Publisher Correction: Functional control of a 0.5 MDa TET aminopeptidase by a flexible loop revealed by MAS NMR

Nature Communications (2022)

Diego F. Gauto Pavel Macek Duccio Malinverni Hugo Fraga Matteo Paloni

10.1038/s41467-022-31243-1

Cite

Citations (0)

EEF2-inactivating toxins engage the NLRP1 inflammasome and promote epithelial barrier disruption uponPseudomonasinfection

bioRxiv (Cold Spring Harbor Laboratory) (2023)

Miriam Pinilla Raoul Mazars Romain Vergé Léana Gorse Karin Santoni

ABSTRACT The intracellular inflammasome complex have been implicated in the maladaptive tissue damage and inflammation observed in chronic Pseudomonas aeruginosa infection. Human airway and corneal epithelial cells, which are critically altered during chronic infections mediated by P. aeruginosa , specifically express the inflammasome sensor NLRP1. Here, together with a companion study, we report that the NLRP1 inflammasome detects Exotoxin A (EXOA), a ribotoxin released by P. aeruginosa Type 2 Secretion System (T2SS) during chronic infection. Mechanistically, EXOA-driven Eukaryotic Elongation Factor 2 (EEF2) ribosylation and covalent inactivation promotes ribotoxic stress and subsequent NLRP1 inflammasome activation, a process shared with other EEF2-inactivating toxins, Diphtheria Toxin and Cholix Toxin. Biochemically, irreversible EEF2 inactivation triggers ribosome stress-associated kinases ZAKα- and P38-dependent NLRP1 phosphorylation and subsequent proteasome-driven functional degradation. Finally, Cystic Fibrosis cells from patients exhibit exacerbated P38 activity and hypersensitivity to EXOA-induced ribotoxic stress-dependent NLRP1 inflammasome activation, a process inhibited by the use of ZAKα inhibitors. Altogether, our results show the importance of P. aeruginosa virulence factor EXOA at promoting NLRP1-dependent epithelial damage and identify ZAKα as a critical sensor of virulence-inactivated EEF2. KEY POINTS P. aeruginosa induces NLRP1-dependent pyroptosis in human corneal and nasal epithelial cells P. aeruginosa Exotoxin A (EXOA) and other EEF2-inactivating bacterial exotoxins activate the human NLRP1 inflammasome EEF2 inactivation promotes ribotoxic stress response and ZAKα kinase-dependent NLRP1 inflammasome activation. Bronchial epithelial cells from Cystic Fibrosis patients show extreme sensitivity to ribotoxic stress-dependent NLRP1 inflammasome activation in response to Exotoxin A P38 and ZAKα inhibition protects Cystic Fibrosis epithelial cell from EXOA-induced pyroptosis

Pseudomonas exotoxin

Pyroptosis

Exotoxin

NLRP1

10.1101/2023.01.16.524164

Cite

Citations (2)

Molecular Crowding Effects on the CyaA Toxin RTX Domain: Implication for Toxin Secretion

The FASEB Journal (2015)

Alexandre Chenal Ana Cristina Sotomayor Pérez Orso Subrini Audrey Hessel Daniel Ladant

Ligand‐induced disorder‐to‐order transition plays a key role in the biological functions of many proteins that contain intrinsically disordered domains. Here, we present data on an RTX (« Repeat in ToXin ») protein, RC L , an intrinsically disordered protein (IDP) that folds upon calcium binding. RTX motifs are calcium‐binding nonapeptide sequences that are found in more than 250 virulence factors secreted by Gram‐negative pathogenic bacteria. The RC L protein is a RTX region of the CyaA toxin, one of the major virulence factors secreted by Bordetella pertussis, the causative agent of the whooping cough. Using a combination of biophysical approaches, we showed that RC L exhibits the hallmarks of IDP in the absence of calcium. Calcium binding triggers a strong reduction of the mean net charge, dehydration and compaction, folding and stabilization of secondary and tertiary structures of RC L . We further showed that the crowding agent Ficoll70 did not affect the structural content of the apo‐state and holo‐state of RC L but increased protein affinity for calcium and stabilizes both states of RC L , increasing their half‐melting temperature (ΔTm), without affecting enthalpy changes. The power law dependence of the ΔTm increase on the volume fraction allowed the estimation of the Flory exponents of the thermally unfolded states. Altogether, our data suggest that, in the apo‐state as found in the crowded bacterial cytosol, RTX proteins adopt extended unfolded conformations that may facilitate protein export by the secretion machinery. Subsequently, both calcium gradient across bacterial cell wall and molecular crowding enhance the calcium‐dependent folding and stability of RTX proteins once secreted in the extracellular milieu.

10.1096/fasebj.29.1_supplement.lb214

Cite

Citations (0)

Molecular Crowding Stabilizes Both the Intrinsically Disordered Calcium-Free State and the Folded Calcium-Bound State of a Repeat in Toxin (RTX) Protein

Journal of the American Chemical Society (2013)

Ana-Cristina Sotomayor-Pérez Orso Subrini Audrey Hessel Daniel Ladant Alexandre Chenal

Macromolecular crowding affects most chemical equilibria in living cells, as the presence of high concentrations of macromolecules sterically restricts the available space. Here, we characterized the influence of crowding on a prototypical RTX protein, RC(L). RTX (Repeat in ToXin) motifs are calcium-binding nonapeptide sequences that are found in many virulence factors produced by Gram-negative bacteria and secreted by dedicated type 1 secretion systems. RC(L) is an attractive model to investigate the effect of molecular crowding on ligand-induced protein folding, as it shifts from intrinsically disordered conformations (apo-form) to a stable structure upon calcium binding (holo-form). It thus offers the rare opportunity to characterize the crowding effects on the same polypeptide chain under two drastically distinct folding states. We showed that the crowding agent Ficoll70 did not affect the structural content of the apo-state and holo-state of RC(L) but increased the protein affinity for calcium. Moreover, Ficoll70 strongly stabilized both states of RC(L), increasing their half-melting temperature, without affecting enthalpy changes. The power law dependence of the melting temperature increase (ΔT(m)) on the volume fraction (φ) followed theoretical excluded volume predictions and allowed the estimation of the Flory exponent (ν) of the thermally unfolded polypeptide chain in both states. Altogether, our data suggest that, in the apo-state as found in the crowded bacterial cytosol, RTX proteins adopt extended unfolded conformations that may facilitate protein export by the type I secretion machinery. Subsequently, crowding also enhances the calcium-dependent folding and stability of RTX proteins once secreted in the extracellular milieu.

Macromolecular Crowding

Folding (DSP implementation)

Intrinsically Disordered Proteins

10.1021/ja404790f

Cite

Citations (46)

Architecture and Assembly Dynamics of the Essential Mitochondrial TIM Chaperone Systems

SSRN Electronic Journal (2021)

Katharina Weinhäupl Yong Wang Audrey Hessel Martha Brennich Kresten Lindorff‐Larsen

The mitochondrial Tim chaperones are responsible for the transport of membrane proteins across the inter-membrane space to the inner and outer mitochondrial membranes. TIM9·10, a hexameric 70 kDa protein complex formed by 3 copies of Tim9 and Tim10, guides its clients across the aque- ous compartment. The TIM9·10·12 complex is the anchor point at the inner-membrane insertase complex TIM22. The subunit composition of the TIM9·10·12 complex remains debated. Joint NMR, small-angle X-ray scattering and MD simulation data al- low us to derive a structural model of the TIM9·10·12 assembly, which has a 2:3:1 stoichiometry (Tim9:Tim10:Tim12). We find that both TIM9·10 and TIM9·10·12 hexamers are in a dynamic equilibrium with their constituent subunits, exchanging on a minutes time scale. Residue-resolved NMR data establish that the subunits exhibit large conformational dynamics: when the conserved cysteines of the CX 3 C-X n -CX 3 C motifs are formed, short marginally stable α -helices are formed, and these are fully stabilized only upon formation of the mature hexameric chap- erone. We propose that the continuous subunit exchange is a means of mitochondria to control their level of inter-membrane space chaperones, and thus rapidly adapt to the cellular state.

Chaperone (clinical)

Inner membrane

10.2139/ssrn.3796489

Cite

Citations (0)

Structural basis of client specificity in mitochondrial membrane-protein chaperones

Science Advances (2020)

Iva Sučec Yong Wang Ons Dakhlaoui Katharina Weinhäupl Tobias Jores

Chaperones are essential for assisting protein folding and for transferring poorly soluble proteins to their functional locations within cells. Hydrophobic interactions drive promiscuous chaperone-client binding, but our understanding of how additional interactions enable client specificity is sparse. Here, we decipher what determines binding of two chaperones (TIM8·13 and TIM9·10) to different integral membrane proteins, the all-transmembrane mitochondrial carrier Ggc1 and Tim23, which has an additional disordered hydrophilic domain. Combining NMR, SAXS, and molecular dynamics simulations, we determine the structures of Tim23/TIM8·13 and Tim23/TIM9·10 complexes. TIM8·13 uses transient salt bridges to interact with the hydrophilic part of its client, but its interactions to the transmembrane part are weaker than in TIM9·10. Consequently, TIM9·10 outcompetes TIM8·13 in binding hydrophobic clients, while TIM8·13 is tuned to few clients with both hydrophilic and hydrophobic parts. Our study exemplifies how chaperones fine-tune the balance of promiscuity versus specificity.

Basis (linear algebra)

10.1126/sciadv.abd0263

Cite

Citations (30)

Characterization of a Membrane-active Peptide from the Bordetella pertussis CyaA Toxin

Journal of Biological Chemistry (2013)

Orso Subrini Ana-Cristina Sotomayor-Pérez Audrey Hessel Johanna Spiaczka-Karst Edithe Selwa

Pertactin

Bordetella

10.1074/jbc.m113.508838

Cite

Citations (51)