Biology subjects

Delalande, O.

Publications and source records attributed to Delalande, O..

3 recordsLinked to original sources

bioRxiv2025

Combining FRET and super-resolution microscopy reveals kinase activation and mitochondrial activity at the nanoscale

Protein kinases are key regulators of intracellular signaling, showing activities in specific subcellular compartments and in micro- or nano-domains. Genetically encoded biosensors based on Forsters Resonance Energy Transfer (FRET) are powerful tools to track kinase dynamics. Yet, they are typically limited by spatial resolution and remain challenging to implement with super-resolution microscopy approaches that require specific fluorophores or optical setups. Aurora kinase A (AURKA), a multifunctional serine/threonine kinase, has recently emerged as a critical regulator of mitochondrial physiology. However, visualising AURKA activation and activity in living cells with sub-diffraction precision remains a challenge. Here, we introduce BioSenSRRF, a versatile approach that combines conventional FRET biosensors with Super-Resolution Radial Fluctuations (SRRF) microscopy. BioSenSRRF requires no modification of existing probes, can be implemented using standard microscopy setups, and is supported by a publicly available ImageJ framework to streamline image processing and data analysis. Using this pipeline, we imaged AURKA activation at mitochondria and its activity in regulating mitochondrial ATP levels. We uncover that AURKA activation and activity are compartmentalised into distinct mitochondrial domains containing the mitochondrial ATP synthase. These subdomains depend on the catalytic activity of AURKA, and they can be altered using previously validated AURKA inhibitors. Finally, we demonstrate that the cancer-associated polymorphism F31I enhances AURKA activation and ATP production on ATP synthase-enriched subdomains, underscoring its pathological relevance. Altogether, BioSenSRRF provides a broadly accessible framework to enhance the spatial resolution of genetically encoded biosensors. This strategy opens new avenues for dissecting the subcellular organisation of protein complexes and their contribution to physiology and disease states.

cell biology↗

bioRxiv2025

Structural characterization of hibernating ribosomes in four Gram-negative pathogenic bacteria

Ribosomes are universal molecular machines that translate messenger RNA into proteins. Depending on what the cells needs for protein synthesis, ribosomes exist in various functional states, and when cells face stressful conditions, their ribosomes may enter into an inactive hibernation state. In bacteria, this ability to become dormant through ribosomal hibernation plays a crucial role in their survival, especially when the cells are targeted by ribosome-targeting antibiotics. The phenomenon occurs when ribosomes bind with hibernation factors. In this work, we examine how these special proteins interact with ribosomes from four different Gram-negative pathogens, and how this leads to translation inhibition, reporting the single-particle cryo-electron microscopy structures of hibernating ribosomes from P. aeruginosa, E. hormaechei, K. quasipneumoniae, and A. baumannii. We show that these ribosomal complexes contain either short HPF or YfiA tightly bound to the 30S A-site, P-site, and even occasionally to the E-site. Our results provide valuable insights into the ribosomal hibernation mechanism, paving the way to both a better understanding of bacterial dormancy and the possibility of developing antibiotics which will target hibernating ribosomes rather than just active ones.

biochemistry↗

bioRxiv2020

Heterodimeric insecticidal peptide provides new insights into the molecular and functional diversity of ant venoms

Ants use venom for predation, defence and communication, however, the molecular diversity, function and potential applications of ant venom remains understudied compared to other venomous lineages such as arachnids, snakes and cone snails. In this work, we used a multidisciplinary approach that encompassed field work, proteomics, sequencing, chemical synthesis, structural analysis, molecular modelling, stability studies, and a series of in vitro and in vivo bioassays to investigate the molecular diversity of the venom of the Amazonian Pseudomyrmex penetrator ants. We isolated a potent insecticidal heterodimeric peptide {Delta}-pseudomyrmecitoxin-Pp1a ({Delta}-PSDTX-Pp1a) composed of a 27-residue long A-chain and a 33-residue long B-chain crosslinked by two disulfide bonds in an antiparallel orientation. We chemically synthesised {Delta}-PSDTX-Pp1a, its corresponding parallel AA and BB homodimers, and its monomeric chains and demonstrated that {Delta}-PSDTX-Pp1a had the most potent insecticidal effects in blow fly assays (LD50 = 3 nM). Molecular modelling and circular dichroism studies revealed strong alpha-helical features, indicating its cytotoxic effects could derive from membrane disruption, which was further supported by insect cell calcium assays. The native heterodimer was also substantially more stable against proteolytic degradation (t1/2 =13 h) than its homodimers or monomers (t1/2 <20 min), indicating an evolutionary advantage of the more complex structure. The proteomic analysis of Pseudomyrmex penetrator venom and in-depth characterisation of {Delta}-PSDTX-Pp1a provide novel insights in the structural complexity of ant venom, and further exemplifies how nature exploits disulfide-bond formation and dimerization to gain an evolutionary advantage via improved stability; a concept that is also highly relevant for the design and development of peptide therapeutics, molecular probes and bioinsecticides.

pharmacology and toxicology↗