Small cargoes pass through synthetically glued Golgi stacks.

How are proteins transported across the stacked cisternae of the Golgi apparatus? Do they stay within the cisterna while the latter matures and progresses in an anterograde manner, or do they navigate between the cisternae via vesicles? Using synthetic biology, we engineered new tools designed to stabilize intercisternal adhesion such that Golgi cisternae are literally glued together, thus preventing any possible cisternal progression. Using bulk secretory assays and single-cell live imaging, we observed that small cargoes (but not large aggregated cargoes including collagen) still transited through glued Golgi, although the rate of transport was moderately reduced. ARF1, whose membrane recruitment is required for budding COPI vesicles, continues to cycle on and off glued Golgi. Numerous COPI-size vesicles were intercalated among the glued Golgi cisternae. These results suggest that cisternal progression is not required for anterograde transport, but do not address the possibility of cisternal maturation in situ

Lack of involvement of CD63 and CD9 tetraspanins in the extracellular vesicle content delivery process

Extracellular vesicles (EVs) are thought to mediate intercellular communication by transferring cargoes from donor to acceptor cells. The EV content-delivery process within acceptor cells is still poorly characterized and debated. CD63 and CD9, members of the tetraspanin family, are highly enriched within EV membranes and are respectively enriched within multivesicular bodies/endosomes and at the plasma membrane of the cells. CD63 and CD9 have been suspected to regulate the EV uptake and delivery process. Here we used two independent assays and different cell models (HeLa, MDA-MB-231 and HEK293T cells) to assess the putative role of CD63 and CD9 in the EV delivery process that includes uptake and cargo delivery. Our results suggest that neither CD63, nor CD9 are required for this function

Monitoring Intra-Golgi Transport with Acute Spatiotemporal Control of a Synthetic Cargo

International audienc

Efficient cell death mediated by bioengineered killer extracellular vesicles

Abstract Extracellular vesicles (EVs) are biological vehicles that are thought to mediate cell–cell communication via the transfer of biomolecules from donor to acceptor cells. Repurposing those natural vesicles into therapeutics delivery vectors is a high priority challenge for translational science. Here we engineer donor cells to produce copious amount of fusogenic EVs loaded with the catalytic domain of the Diphteria Toxin, known to trigger cell death through protein synthesis inhibition. We show that, when incubated with cancer acceptor cells, these Killer EVs block protein synthesis and lead to cell death. This proof of concept establishes the efficacy of Killer EVs in vitro, and suggests that further development may lead to tumor ablation in vivo, expanding the existing cancer therapeutics arsenal

Combining feature selection and shape analysis uncovers precise rules for miRNA regulation in Huntington’s disease mice

International audienceBACKGROUND:MicroRNA (miRNA) regulation is associated with several diseases, including neurodegenerative diseases. Several approaches can be used for modeling miRNA regulation. However, their precision may be limited for analyzing multidimensional data. Here, we addressed this question by integrating shape analysis and feature selection into miRAMINT, a methodology that we used for analyzing multidimensional RNA-seq and proteomic data from a knock-in mouse model (Hdh mice) of Huntington's disease (HD), a disease caused by CAG repeat expansion in huntingtin (htt). This dataset covers 6 CAG repeat alleles and 3 age points in the striatum and cortex of Hdh mice.RESULTS:Remarkably, compared to previous analyzes of this multidimensional dataset, the miRAMINT approach retained only 31 explanatory striatal miRNA-mRNA pairs that are precisely associated with the shape of CAG repeat dependence over time, among which 5 pairs with a strong change of target expression levels. Several of these pairs were previously associated with neuronal homeostasis or HD pathogenesis, or both. Such miRNA-mRNA pairs were not detected in cortex.CONCLUSIONS:These data suggest that miRNA regulation has a limited global role in HD while providing accurately-selected miRNA-target pairs to study how the brain may compute molecular responses to HD over time. These data also provide a methodological framework for researchers to explore how shape analysis can enhance multidimensional data analytics in biology and disease

Lack of involvement of CD63 and CD9 tetraspanins in the extracellular vesicle content delivery process

International audienceExtracellular vesicles (EVs) are thought to mediate intercellular communication by transferring cargoes from donor to acceptor cells. The EV content-delivery process within acceptor cells is still poorly characterized and debated. CD63 and CD9, members of the tetraspanin family, are highly enriched within EV membranes and are respectively enriched within multivesicular bodies/endosomes and at the plasma membrane of the cells. CD63 and CD9 have been suspected to regulate the EV uptake and delivery process. Here we used two independent assays and different cell models (HeLa, MDA-MB-231 and HEK293T cells) to assess the putative role of CD63 and CD9 in the EV delivery process that includes uptake and cargo delivery. Our results suggest that neither CD63, nor CD9 are required for this function

Lack of involvement of CD63 and CD9 tetraspanins in the extracellular vesicle content delivery process.

Extracellular vesicles (EVs) are thought to mediate intercellular communication by transferring cargoes from donor to acceptor cells. The EV content-delivery process within acceptor cells is still poorly characterized and debated. CD63 and CD9, members of the tetraspanin family, are highly enriched within EV membranes and are respectively enriched within multivesicular bodies/endosomes and at the plasma membrane of the cells. CD63 and CD9 have been suspected to regulate the EV uptake and delivery process. Here we used two independent assays and different cell models (HeLa, MDA-MB-231 and HEK293T cells) to assess the putative role of CD63 and CD9 in the EV delivery process that includes uptake and cargo delivery. Our results suggest that neither CD63, nor CD9 are required for this function