Exosomes: Fit to deliver small RNA

Abstract

Exosomes are specialized membranous nano-sized vesicles derived from endocytic compartments that are released by many cell types. Microvesicles are distinctive from exosomes in that they are produced by shedding of the plasmamembrane and usually larger in size (>1 µm). Exosome biogenesis involves the tightly controlled process of inward budding from the limiting membrane of multivesicular bodies (MVBs). This results in numerous intraluminal vesicles in the lumen of MVBs that contain distinct protein repertoires. It has been suggested that microvesicles shed by certain tumor cells hold functional messenger RNA (mRNA) that may promote tumor progression. We discovered that purified exosomes contain functional microRNAs (miRNAs) and small RNA, but detected little mRNA. Although a clear and decisive distinction between microvesicles and exosomes cannot be made and different subsets of exosomes exist, we speculate that exosomes are specialized in carrying small RNA including the class 22-25 nucleotide regulatory miRNAs. To demonstrate this we developed a co-culture system and found that exosomes are continuously secreted and transferred from Epstein Barr virus (EBV)-infected cells to uninfected neighboring cells. Throughout exosome transfer, the exogenous EBV-encoded miRNAs were delivered to subcellular sites of miRNA-mediated gene repression. Additionally, we found evidence that mature miRNAs are transferred between circulating cells in humans, since we detected EBV-miRNAs in non-infected cells in the peripheral blood of patients that include monocytes and T cells. In this addendum we discuss these findings in the context of recently published papers that advanced our current knowledge of exosome physiology, (mi)RNA function and intercellular RNA transfer. Based on this information we propose that an intercellular (miRNA-based) mode of signal transmission may be well suited in controlling space-confined processes such as the initiation of immune responses in the secondary (peripheral) lymphoid tissues or in a tumor microenvironment. Deciphering the molecular mechanism(s) that control small RNA loading into exosomes and transfer to recipient cells in vitro will provide new evidence for the physiological relevance of vesicle-mediated intercellular communication in vivo.

Keywords: exosomes; intercellular communication; microvesicles; small RNA; virus.

Figure 1

Exosome-mediated small RNA transfer in a co-culture system. A co-culture transwell device with 5,00,000 EBV positive exosome producer cells (LCL) and 1,00,000 dendritic cells (immature monocyte-derived dendritic cells) as recipients, were co-cultured for the indicated times and separated by a semi-permeable membrane with either a 1.0 µm (black bars) or 0.4 µm pore-size (grey bar). The bars represent the relative amount of EBV-encoded small RNA (EBERs), detected by semi-quantitative RT-PCR in the recipient cells as an indication for small RNA transfer.

Figure 2

EM image of purified LCL exosomes with typical cup-shaped morphology. Exosomes may operate together or alone (arrow). Bar indicates 100 nm.