Abstract
Healthy brain function is mediated by several complementary signalling pathways, many of which are driven by extracellular vesicles (EVs). EVs are heterogeneous in both size and cargo and are constitutively released from cells into the extracellular milieu. They are subsequently trafficked to recipient cells, whereupon their entry can modify the cellular phenotype. Here, in order to further analyse the mRNA and protein cargo of neuronal EVs, we isolated EVs by size exclusion chromatography from human induced pluripotent stem cell (iPSC)-derived neurons. Electron microscopy and dynamic light scattering revealed that the isolated EVs had a diameter of 30–100 nm. Transcriptomic and proteomics analyses of the EVs and neurons identified key molecules enriched in the EVs involved in cell surface interaction (integrins and collagens), internalisation pathways (clathrin- and caveolin-dependent), downstream signalling pathways (phospholipases, integrin-linked kinase and MAPKs), and long-term impacts on cellular development and maintenance. Overall, we show that key signalling networks and mechanisms are enriched in EVs isolated from human iPSC-derived neurons.
Highlights
Extracellular vesicles (EVs) represent a major contact-independent mechanism for intercellular communication in the brain [1]
A Subgroup of Cellular mRNA Transcripts are Selectively Incorporated into Neuronal EVs
In order to characterize the mRNA cargo of neuronal sEVs, RNA-seq was performed on the induced pluripotent stem cell (iPSC)-derived neurons and their EVs
Summary
Extracellular vesicles (EVs) represent a major contact-independent mechanism for intercellular communication in the brain [1]. EVs comprise a wide range of sizes, cargoes and subcellular origins [2, 3]. Small EVs, defined as having a diameter of approximately 50–150 nm, include exosomes which are of endosomal origin [4, 5]. EVs are released from the cell and contain a heterogeneous population of macromolecules, including lipids, proteins and various classes of RNA [2]. The mRNA cargo can be translationally competent as demonstrated by microarray analysis in recipient cells [7] and translation of Arc protein in recipient cells [8], which may modulate synaptic function. EVs have been implicated as a conduit for the cell to cell transfer of several misfolded proteins in neurodegenerative diseases, which may be a mechanism for the spread of pathogenic protein conformers in neurodegeneration [9]. A range of cellular stresses, including hypoxia and hypoglycaemia have been shown to modify EV cargo, including mRNA [10,11,12,13,14,15]
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