Barcoding of small extracellular vesicles with CRISPR-gRNA enables comprehensive, subpopulation-specific analysis of their biogenesis and release regulators.
basic_science · Level V
Where this comes from
- Record sourced from PubMed, PMID 39562573.
- Also identified by DOI 10.1038/s41467-024-53736-x and PMC identifier 11577021.
- Licence recorded as CC BY.
- The licence permits redistribution, so the abstract is shown in full and the full text is available from the publisher.
Abstract
Small extracellular vesicles (sEVs) are important intercellular information transmitters in various biological contexts, but their release processes remain poorly understood. Herein, we describe a high-throughput assay platform, CRISPR-assisted individually barcoded sEV-based release regulator (CIBER) screening, for identifying key players in sEV release. CIBER screening employs sEVs barcoded with CRISPR-gRNA through the interaction of gRNA and dead Cas9 fused with an sEV marker. Barcode quantification enables the estimation of the sEV amount released from each cell in a massively parallel manner. Barcoding sEVs with different sEV markers in a CRISPR pooled-screening format allows genome-wide exploration of sEV release regulators in a subpopulation-specific manner, successfully identifying previously unknown sEV release regulators and uncovering the exosomal/ectosomal nature of CD63<sup>+</sup>/CD9<sup>+</sup> sEVs, respectively, as well as the synchronization of CD9<sup>+</sup> sEV release with the cell cycle. CIBER should be a valuable tool for detailed studies on the biogenesis, release, and heterogeneity of sEVs.
Medical subject headings
- Extracellular Vesicles
- CRISPR-Cas Systems
- RNA, Guide, CRISPR-Cas Systems
- Tetraspanin 29
- Tetraspanin 30