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Page 54 Extracell Vesicles Circ Nucleic Acids 2020;1:20-56 I http://dx.doi.org/10.20517/evcna.2020.10

model systems to study exosome dynamics and function by expressing CD63-pHluorin in cells and
zebrafish embryos [1-3] . This work represents a comprehensive and complimentary approach to study
endogenous EV regulation and function in vitro and in vivo at high spatiotemporal accuracy.

REFERENCES
1. Verweij FJ, Bebelman MP, Jimenez CR, et al. Quantifying exosome secretion from single cells reveals a modulatory role for GPCR
signaling. J Cell Biol 2018; 217:1129-42.
2. Verweij FJ, Revenu C, Arras G, et al. Live Tracking of Inter-organ Communication by Endogenous Exosomes In Vivo. Dev Cell
2019;48:573-589.e4.
3. Bebelman, M.P., Bun, P., Huveneers, S. et al. Real-time imaging of multivesicular body–plasma membrane fusion to quantify exosome
release from single cells. Nat Protoc 2020;15:102-21.

45. A CRISPR-Cas9-based reporter system for single-cell detection of extracellular vesicle-
mediated functional transfer of RNA

1
1,2
2
2,3
Authors: Olivier G. de Jong , Daniel E. Murphy , Imre Mäger , Eduard Willms , Antonio Garcia-
2,4
6
7
1
5
Guerra , Jerney J. Gitz-Francois , Juliet Lefferts , Dhanu Gupta , Sander C. Steenbeek , Jacco van
2
1
6
7
Rheenen , Samir El Andaloussi , Raymond M. Schiffelers , Matthew J. A. Wood , Pieter Vader 1,8
E-mail: pvader@umcutrecht.nl
Affiliations:
1 Laboratory of Clinical Chemistry and Hematology, University Medical Center Utrecht, Utrecht, The
Netherlands.
2 Department of Paediatrics, University of Oxford, Oxford, United Kingdom.
3 Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, United Kingdom.
4 Department of Physics, University of Oxford, Oxford, United Kingdom.
5 Pediatric Pulmonology and Regenerative Medicine Center, University Medical Center Utrecht, Utrecht, The
Netherlands.
6 Department of Laboratory Medicine, Clinical Research Center, Karolinska Institutet, Huddinge, Sweden.
7 Department of Molecular Pathology, Oncode Institute, Netherlands Cancer Institute, Amsterdam, The
Netherlands.
8 Department of Experimental Cardiology, University Medical Center Utrecht, Utrecht, The Netherlands.

Abstracts: Extracellular vesicles (EVs) play an important role in communication between cells through
functional transfer of bioactive cargo, including RNA. Despite increasing interest in EV-mediated RNA
transfer, in-depth knowledge on mechanisms underlying EV-mediated RNA delivery and processing is
limited due to a lack of suitable readout systems. Here, we report a novel CRISPR/Cas9-based reporter
system that allows for studying EV-mediated RNA transfer at a single-cell level. After validation of this
system by studying the role of known targets involved in EV uptake and intracellular membrane trafficking,
we have employed this system to uncover various novel genes that play a regulatory role in functional
RNA transfer. Thus, this novel system may be used for the study of specific genetic targets and pathways
underlying EV-mediated functional RNA delivery.

46. Exploitation of the Leishmania Exosomal Pathway by Leishmania RNA virus 1

Authors: Martin Olivier
E-mail: martin.olivier@mcgill.ca