Biology subjects

Henriquet, C.

Publications and source records attributed to Henriquet, C..

3 recordsLinked to original sources

bioRxiv2026

Characterization of the p38α MAPK allosteric inhibition by a single chain Fv antibody

Most of the available p38 kinase inhibitors target the highly conserved ATP-binding site, thus explaining their off-target effects and toxicity. Here, we report the use of single-chain fragment variable (scFv) antibodies to identify new highly specific inhibitors. Using phage display, we selected five anti-p38 scFv antibodies among which one fully inhibited p38 kinase activity in vitro. We showed that this scFv did not affect ATP binding, while still acting as a competitive inhibitor of ATP hydrolysis. When expressed as intrabody in the nucleus of the THP-1 human monocytic cell line, the scFv inhibited the function of endogenous p38 and induced a significant decrease in lipopolysaccharide-induced tumor necrosis factor production. To gain insight into the mechanisms by which the scFv induced p38 inhibition, the scFv epitope was mapped by deep mutational scanning of p38 displayed on yeast surface and high-throughput selection by flow cytometry of p38 mutants no longer bound by the scFv. By this approach, we showed that the inhibitory scFv interacts with two helices within the C-terminal lobe of the kinase distant from the ATP-binding pocket and the docking groove. Altogether, our data show that the scFv antibody behaves as an allosteric inhibitor, constraining and maintaining p38 in an inactive form.

biochemistry↗

bioRxiv2021

Design of a new effector recognition specificity in a plant NLR immune receptor by molecular engineering of its integrated decoy domain

Plant nucleotide-binding and leucine-rich repeat domain proteins (NLRs) are immune sensors that specifically recognize pathogen effectors and induce immune responses. Designing artificial NLRs with new effector recognition specificities is a promising prospect for sustainable, knowledge-driven crop protection. However, such strategies are hampered by the complexity of NLR function. Here, we tested whether molecular engineering of the integrated decoy domain (ID) of an NLR could extend its recognition spectrum to a new effector. To this aim, we relied on the detailed molecular knowledge of the recognition of distinct Magnaporthe oryzae MAX (Magnaporthe AVRs and ToxB-like) effectors by the rice NLRs RGA5 and Pikp-1. For both NLRs, effector recognition involves physical binding to their HMA (Heavy Metal-Associated) IDs. However, AVR-PikD, the effector recognized by Pikp-1, binds to a completely different surface of the HMA domain compared to AVR-Pia and AVR1-CO39, recognized by RGA5. By introducing into the HMA domain of RGA5 the residues of the Pikp-1 HMA domain involved in AVR-PikD binding, we created a high-affinity binding surface for this new effector. In the Nicotiana benthamiana heterologous system, RGA5 variants carrying this engineered binding surface still recognize AVR-Pia and AVR1-CO39, but also perceive the new ligand, AVR-PikD, resulting in the activation of immune responses. Therefore, our study provides a proof of concept for the design of new effector recognition specificities in NLRs through molecular engineering of IDs. However, it pinpoints significant knowledge gaps that limit the full deployment of this NLR-ID engineering strategy and provides hypotheses for future research on this topic.

plant biology↗

bioRxiv2020

Lactoferrin retargets adenoviruses to TLR4 to induce an abortive NLRP3-associated pyroptotic response in human dendritic cells

Despite decades of investigations, we still poorly grasp the immunogenicity of human adenovirus (HAdV)-based vaccines in humans. In this study, we explored the role of lactoferrin, which belong to the alarmin subset of antimicrobial peptides that provide immediate direct and indirect activity against a range of pathogens following a breach in tissue homeostasis. Lactoferrin is a globular, iron-sequestering, glycoprotein that can increase HAdV infection and maturation of antigen-presenting cells. However, the mechanism by which HAdV-lactoferrin complexes induce maturation is unknown. We show that lactoferrin redirects HAdVs from species B, C, and D to toll-like receptor 4 (TLR4) complexes on human mononuclear phagocyte. TLR4-mediated internalization induces an abortive NLRP3-associated pyroptotic response inducing pro-inflammatory cytokine release and disrupting plasma membrane integrity without cell death. These data impact our understanding of the immunogenicity of HAdV-based vaccines and may provide ways to increase their efficacy.

immunology↗