MYO1C stabilizes actin and facilitates the arrival of transport carriers at the Golgi complex

In this study, we aimed to identify the myosin motor proteins that control trafficking at the Golgi complex. In addition to the known Golgi-associated myosins MYO6, MYO18A and MYH9 (myosin IIA), we identified MYO1C as a novel player at the Golgi in a human cell line. We demonstrate that depletion of MYO1C induces Golgi complex fragmentation and decompaction. MYO1C accumulates at dynamic structures around the Golgi complex that colocalize with Golgi-associated actin dots. MYO1C depletion leads to loss of cellular F-actin, and Golgi complex decompaction is also observed after inhibition or loss of the actin-related protein 2/3 complex, Arp2/3 (also known as ARPC). We show that the functional consequence of MYO1C depletion is a delay in the arrival of incoming transport carriers, both from the anterograde and retrograde routes. We propose that MYO1C stabilizes actin at the Golgi complex, facilitating the arrival of incoming transport carriers at the Golgi.This article has an associated First Person interview with the first author of the paper.Fil: Capmany, Anahi. Institute Curie; Francia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mendoza; Argentina. Universidad Nacional de Cuyo. Facultad de Ciencias Medicas. Instituto de Inmunologia; ArgentinaFil: Yoshimura, Azumi. Institute Curie; FranciaFil: Kerdous, Rachid. Institute Curie; FranciaFil: Caorsi, Valentina. Abbelight; FranciaFil: Lescure, Aurianne. Institute Curie; FranciaFil: Nery, Elaine Del. Institute Curie; FranciaFil: Coudrier, Evelyne. Institute Curie; FranciaFil: Goud, Bruno. Institute Curie; FranciaFil: Schauer, Kristine. Institute Curie; Franci

Permeability of DOPC bilayers under photoinduced oxidation: Sensitivity to photosensitizer.

The modification of lipid bilayer permeability is one of the most striking yet poorly understood physical transformations that follow photoinduced lipid oxidation. We have recently proposed that the increase of permeability of photooxidized 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) bilayers is controlled by the time required by the oxidized lipid species to diffuse and aggregate into pores. Here we further probe this mechanism by studying photosensitization of DOPC membranes by methylene blue (MB) and DO15, a more hydrophobic phenothiazinium photosensitizer, under different irradiation powers. Our results not only reveal the interplay between the production rate and the diffusion of the oxidized lipids, but highlight also the importance of photosensitizer localization in the kinetics of oxidized membrane permeability

Utilisation de nanoparticules copolymériques pour le ciblage cellulaire de photosensibilisateurs (étude sur des membranes modèles et corrélation avec l internalisation cellulaire)

La thérapie photodynamique (PDT) représente actuellement une technique alternative à la chimiothérapie traditionnelle pour le traitement des cancers. Elle est basée sur la capacité de certaines molécules appelées photosensibilisateurs à générer des espèces oxygénées radicalaires suite à une irradiation lumineuse. La plupart de ces molécules sont connues pour leur rétention préférentielle par les tissus cancéreux, mais un certains nombre d inconvénients limitent leur utilisation. Notamment, leur relative hydrophobie limite leur solubilité dans les milieux biologiques. De nouvelles formules de photosensibilisateurs sont en cours de développement, comme l utilisation de nanoparticules, pour les véhiculer jusqu aux tissus cibles, permettant de réduire leur toxicité et d augmenter leur spécificité. Parmi celles-ci, les nanoparticules polymériques amphiphiles de PEO-b-PCL constituent de bons candidats en raison de leur biodégradabilité et de la possibilité de générer des nano-objets de tailles contrôlées. En outre, ces systèmes permettent de contrôler le relargage du photosensibilisateur véhiculé une fois dans les cellules cibles. Le but du travail présenté dans ce manuscrit se situe dans ce contexte et vise à comprendre les mécanismes impliqués dans l internalisation cellulaire d un photosensibilisateur, le phéo-a, seul ou par l intermédiaire des nanoparticules. La capacité de ce photosensibilisateur à interagir avec les membranes modèles, mimant la partie lipidiques des membranes biologiques, a été étudiée, en nous attachant notamment aux effets du taux de cholestérol, qui est un composant membranaire important, gouvernant ses propriétés physiques. Nos résultats montrent qu il implique, entre autre, une réduction importante du flip-flop du phéo-a à travers la membrane. La capacité du phéophorbide-a à endommager et perméabiliser les membranes a été ensuite évaluée. Cet effet perméabilisant du phéo-a a été comparé à celui d une chlorine tri-carboxylique amphiphile, la chlorine e6. L interaction du phéo-a avec les nanoparticules et son transfert à partir de ces compartiments vers les membranes ont été étudiée. Aucun transfert de la phéo-a par la phase aqueuse n a lieu, mais le transfert nécessite une collision directe entre les nanoparticules avec les vésicules. Au niveau cellulaire, les nanoparticules ont montré une nette amélioration de la quantité de phéo-a internalisée par rapport à celle internalisée seule. En outre, les mécanismes de cette internalisation ont été mis en évidence. Avec les nanoparticules, l incorporation du phéophorbide est bi-phasique. La première phase rapide est attribuée au transfert direct du photosensibilisateur des nanoparticules vers les membranes, au sein desquelles il se redistribue. Elle est suivie d une seconde phase lente attribuée à l endocytose des nanoparticules.Photodynamic therapy (PDT) is currently an alternative technique to traditional chemotherapy for cancer treatment. It is based on the ability of certain molecules called photosensitizers to generate reactive oxygen species after light irradiation. Most of these molecules are known for their preferential retention in tumor tissues, but a number of drawbacks limit their use. Particularly, their relative hydrophobicity limits their solubility in biological media. New forms of photosensitizers are being developed, such as the use of nanoparticles to convey to the targeted tissues, to reduce toxicity and increase their specificity. Among these, the amphiphilic polymer nanoparticles PEO-b-PCL are good candidates because of their biodegradability and ability to generate controlled nano-sized objects. In addition, these systems are used to control the release of the photosensitizer once transported into target cells. The aim of the work presented in this manuscript aims to understand the mechanisms involved in the cellular internalization of a photosensitizer, the pheophorbide-a (pheo-a), alone or via nanoparticles. The interaction of this photosensitizer with model of membranes mimicking the lipidic part of the biological ones has been studied, in particular, by focusing on the effects of cholesterol, which is a major membrane component, governing its physical properties. Our results show that involves, among other mechanisms, a significant reduction of the flip-flop rate of the phéo-a through the membrane. The ability of this photosensitizer to damage and permeabilize the membranes was then evaluated in comparison to that of an amphiphilic, tri-carboxylic chlorine, the chlorin e6. The interaction of the pheo-a with nanoparticles and its transfer from these compartments to the membranes were studied. No transfer of the pheo-a via the aqueous phase takes place, but the transfer requires a direct collision between the nanoparticles with the vesicles. At the cellular level, the nanoparticles showed a marked improvement in the amount of internalized pheo-a relative to the one internalized without nanoparticles. In addition, the mechanisms of internalization have been highlighted. With nanoparticles, the incorporation of pheo-a is biphasic. The first phase is assigned to the fast direct transfer of the photosensitizer from the nanoparticles to the membranes, in which it redistributes. It is followed by a second phase, assigned to slow rating endocytosis of the nanoparticles.PARIS-BIUSJ-Physique recherche (751052113) / SudocSudocFranceF

Light-Triggered Sequence-Specific Cargo Release from DNA Block Copolymer-Lipid Vesicles

Versatile functionalized nanocontainers, based on the stable incorporation of 22 mer DNA-b-PPO block copolymers (DBCs) into lipid vesicles, are presented (see picture). The study shows effective and sequence-specific cargo release from the DBC–lipid vesicles. Hybridization of these vesicles with an oligonucleotide photosensitizer allows for singlet oxygen generation upon irradiation, which induces cargo release.

Photosensitized Degradation of Model Lipid Membranes based on 1-palmitoyl-2-oleyl-phosphatidylcholine (POPC)

In this work, we study the interaction of a well-known photosensitizer, MePha, with models of biological membrane (Langmuir monolayers and Langmuir–Schaeffer planar bilayers) based on one of the most important natural lipid, POPC, for the subsequent investigation of photodestruction processes in a context of photodynamic therapy treatment. Changes of macroscopic properties and morphology of POPC/MePha model membranes upon irradiation by visible light are recorded by means of contact angle measurements and atomic force microscopy, demonstrating clearly the possibility to use these methods for the study of photodestruction of artificial lipid membranes on solid substrates, but also for a comparative study of the efficiency of novel photosensitizers