Abstract

From Seeing to Believing: Visualization and Tracking of Extracellular Vesicles

Charles Lai, Assistant Research Fellow, Institute of Atomic and Molecular Sciences, Academia Sinica

Extracellular vesicles (EVs), including exosomes and microvesicles, are nanosized membrane- bound vesicles released in abundance by cancer cells. These EVs are capable of delivering a select subset of proteins and RNAs designed to promote tumor growth and metastases. However, the exact time, location (i.e. spatiotemporal properties) and cargo delivery capacity of EVs remained unknown due to a lack of available methods to detect them. Here we engineered bioluminescent and fluorescent reporter systems which allow EV visualization in real-time both in vitro and in vivo. The multimodal bioluminescent reporter labels EVs with a membrane-bound variant of Gaussia luciferase fused to a biotin acceptor peptide, thereby enabling whole-animal multimodal imaging of nanoscale EVs, as well as pharmacokinetics analysis of EVs. We next developed the bifunctional fluorescent EV/EV-RNA reporters to study multiple EV populations with subcellular image resolution. By multiplexing the fluorescent and bioluminescent EV reporters, we further elucidated the rapid dynamics of tumor EV uptake and translation of EV-delivered cargo mRNAs in cancer cells. These novel molecular imaging platforms revealed that EV mediates a dynamic and multidirectional communication between cells with delivery of functional mRNA, thereby manipulating its recipient cells at neighboring and distal sites.