Exosome-mediated transfer of mRNAs and microRNAs is a novel mechanism of genetic exchange between cells
TL;DR: It is shown that exosomes contain both mRNA and microRNA, which can be delivered to another cell, and can be functional in this new location, and it is proposed that this RNA is called “exosomal shuttle RNA” (esRNA).
Abstract: Exosomes are vesicles of endocytic origin released by many cells. These vesicles can mediate communication between cells, facilitating processes such as antigen presentation. Here, we show that exosomes from a mouse and a human mast cell line (MC/9 and HMC-1, respectively), as well as primary bone marrow-derived mouse mast cells, contain RNA. Microarray assessments revealed the presence of mRNA from approximately 1300 genes, many of which are not present in the cytoplasm of the donor cell. In vitro translation proved that the exosome mRNAs were functional. Quality control RNA analysis of total RNA derived from exosomes also revealed presence of small RNAs, including microRNAs. The RNA from mast cell exosomes is transferable to other mouse and human mast cells. After transfer of mouse exosomal RNA to human mast cells, new mouse proteins were found in the recipient cells, indicating that transferred exosomal mRNA can be translated after entering another cell. In summary, we show that exosomes contain both mRNA and microRNA, which can be delivered to another cell, and can be functional in this new location. We propose that this RNA is called "exosomal shuttle RNA" (esRNA).
Citations
More filters
TL;DR: Exosomes transfer not only membrane components but also nucleic acid between different cells, emphasizing their role in intercellular communication.
1,979 citations
Cornell University1, University of Pennsylvania2, NewYork–Presbyterian Hospital3, Hannover Medical School4, Rockefeller University5, National Taiwan University6, Oslo University Hospital7, University of Oslo8, Memorial Sloan Kettering Cancer Center9, Eppley Institute for Research in Cancer and Allied Diseases10, University of Nebraska Medical Center11
TL;DR: It is shown that PDAC-derived exosomes induce liver pre-metastatic niche formation in naive mice and consequently increase liver metastatic burden and suggests that exosomal MIF primes the liver for metastasis and may be a prognostic marker for the development of PDAC liver metastasis.
Abstract: Lyden and colleagues report that pancreatic cancer-derived exosomes induce a pre-metastatic niche in the liver by promoting TGFβ secretion from Kupffer cells, leading to fibronectin production in hepatic stellate cells and macrophage recruitment.
1,973 citations
TL;DR: The maturation programme entails a dramatic transformation of these dynamic organelles disconnecting them functionally and spatially from early endosomes and preparing them for their unidirectional role as a feeder pathway to lysosomes.
Abstract: Being deeply connected to signalling, cell dynamics, growth, regulation, and defence, endocytic processes are linked to almost all aspects of cell life and disease. In this review, we focus on endosomes in the classical endocytic pathway, and on the programme of changes that lead to the formation and maturation of late endosomes/multivesicular bodies. The maturation programme entails a dramatic transformation of these dynamic organelles disconnecting them functionally and spatially from early endosomes and preparing them for their unidirectional role as a feeder pathway to lysosomes.
1,859 citations
Cites background from "Exosome-mediated transfer of mRNAs ..."
...A similar process has, interestingly, been proposed for the inclusion of certain RNAs into ILVs (Valadi et al, 2007; Gibbings and Voinnet, 2010)....
[...]
TL;DR: The need for standardization of specimen handling, appropriate normative controls, and isolation and analysis techniques to facilitate comparison of results is emphasized, and it is recognized that continual development and evaluation of techniques will be necessary as new knowledge is amassed.
Abstract: The emergence of publications on extracellular RNA (exRNA) and extracellular vesicles (EV) has highlighted the potential of these molecules and vehicles as biomarkers of disease and therapeutic targets. These findings have created a paradigm shift, most prominently in the field of oncology, prompting expanded interest in the field and dedication of funds for EV research. At the same time, understanding of EV subtypes, biogenesis, cargo and mechanisms of shuttling remains incomplete. The techniques that can be harnessed to address the many gaps in our current knowledge were the subject of a special workshop of the International Society for Extracellular Vesicles (ISEV) in New York City in October 2012. As part of the “ISEV Research Seminar: Analysis and Function of RNA in Extracellular Vesicles (evRNA)”, 6 round-table discussions were held to provide an evidence-based framework for isolation and analysis of EV, purification and analysis of associated RNA molecules, and molecular engineering of EV for therapeutic intervention. This article arises from the discussion of EV isolation and analysis at that meeting. The conclusions of the round table are supplemented with a review of published materials and our experience. Controversies and outstanding questions are identified that may inform future research and funding priorities. While we emphasize the need for standardization of specimen handling, appropriate normative controls, and isolation and analysis techniques to facilitate comparison of results, we also recognize that continual development and evaluation of techniques will be necessary as new knowledge is amassed. On many points, consensus has not yet been achieved and must be built through the reporting of well-controlled experiments. Keywords: extracellular vesicle; exosome; microvesicle; standardization; isolation (Published: 27 May 2013) Citation: Journal of Extracellular Vesicles 2013, 2 : 20360 - http://dx.doi.org/10.3402/jev.v2i0.20360
1,840 citations
Cites background or methods from "Exosome-mediated transfer of mRNAs ..."
...growing evidence for enrichment of certain RNA species in specific vesicle types (1,17,18)....
[...]
...EV are generally purified from medium conditioned by cells for 24 48 h, but shorter conditioning times have been used to analyze specific release of EV-associated markers after a specific stimulus: for instance, surface receptormediated stimulation (214,215) or fMLP, PMA or calcium ionophore treatment for 15 60 min (1,216,217)....
[...]
TL;DR: The novel role of exosomes highlights a new perspective into intercellular mediation of tissue injury and repair, and engenders novel approaches to the development of biologics for tissue repair.
1,816 citations
Cites background from "Exosome-mediated transfer of mRNAs ..."
..., 2009; Taylor and Gercel-Taylor, 2008), peripheral blood cells (Hunter et al., 2008; Valadi et al., 2007), endothelial progenitor cells (Deregibus et al....
[...]
...More importantly, these microvesicular RNA could be transferred to other cells and translated in the recipient cells (Deregibus et al., 2007; Ratajczak et al., 2006; Valadi et al., 2007)....
[...]
...Several tumor cell types (Rosell et al., 2009; Taylor and Gercel-Taylor, 2008), peripheral blood cells (Hunter et al., 2008; Valadi et al., 2007), endothelial progenitor cells (Deregibus et al., 2007), and embryonic stem cells (Ratajczak et al., 2006) have been shown to secrete RNA-containing…...
[...]
References
More filters
TL;DR: PicTar, a computational method for identifying common targets of micro RNAs, is presented and widespread coordinate control executed by microRNAs is suggested, thus providing evidence for coordinate microRNA control in mammals.
Abstract: MicroRNAs are small noncoding RNAs that recognize and bind to partially complementary sites in the 3' untranslated regions of target genes in animals and, by unknown mechanisms, regulate protein production of the target transcript. Different combinations of microRNAs are expressed in different cell types and may coordinately regulate cell-specific target genes. Here, we present PicTar, a computational method for identifying common targets of microRNAs. Statistical tests using genome-wide alignments of eight vertebrate genomes, PicTar's ability to specifically recover published microRNA targets, and experimental validation of seven predicted targets suggest that PicTar has an excellent success rate in predicting targets for single microRNAs and for combinations of microRNAs. We find that vertebrate microRNAs target, on average, roughly 200 transcripts each. Furthermore, our results suggest widespread coordinate control executed by microRNAs. In particular, we experimentally validate common regulation of Mtpn by miR-375, miR-124 and let-7b and thus provide evidence for coordinate microRNA control in mammals.
4,660 citations
"Exosome-mediated transfer of mRNAs ..." refers background in this paper
...Hypothetically, the 121 miRNAs that we have found in the mast cell exosomes may interfere with 24,000 mRNAs as it has been suggested that each species may interact with up to 200 mRNA...
[...]
TL;DR: It is demonstrated by immunoelectron microscopy that the limiting membrane of MIICs can fuse directly with the plasma membrane, resulting in release from the cells of internal MHC class II-containing vesicles, suggesting a role for exosomes in antigen presentation in vivo.
Abstract: Antigen-presenting cells contain a specialized late endocytic compartment, MIIC (major histocompatibility complex [MHC] class II-enriched compartment), that harbors newly synthesized MHC class II molecules in transit to the plasma membrane. MIICs have a limiting membrane enclosing characteristic internal membrane vesicles. Both the limiting membrane and the internal vesicles contain MHC class II. In this study on B lymphoblastoid cells, we demonstrate by immunoelectron microscopy that the limiting membrane of MIICs can fuse directly with the plasma membrane, resulting in release from the cells of internal MHC class II-containing vesicles. These secreted vesicles, named exosomes, were isolated from the cell culture media by differential centrifugation followed by flotation on sucrose density gradients. The overall surface protein composition of exosomes differed significantly from that of the plasma membrane. Exosome-bound MHC class II was in a compact, peptide-bound conformation. Metabolically labeled MHC class II was released into the extracellular medium with relatively slow kinetics, 10 +/- 4% in 24 h, indicating that direct fusion of MIICs with the plasma membrane is not the major pathway by which MHC class II reaches the plasma membrane. Exosomes derived from both human and murine B lymphocytes induced antigen-specific MHC class II-restricted T cell responses. These data suggest a role for exosomes in antigen presentation in vivo.
2,978 citations
TL;DR: The results indicate that exosome isolation may provide an efficient first step in biomarker discovery in urine and identify numerous protein components of MVBs and of the endosomal pathway in general.
Abstract: Urine provides an alternative to blood plasma as a potential source of disease biomarkers. One urinary biomarker already exploited in clinical studies is aquaporin-2. However, it remains a mystery how aquaporin-2 (an integral membrane protein) and other apical transporters are delivered to the urine. Here we address the hypothesis that these proteins reach the urine through the secretion of exosomes [membrane vesicles that originate as internal vesicles of multivesicular bodies (MVBs)]. Low-density urinary membrane vesicles from normal human subjects were isolated by differential centrifugation. ImmunoGold electron microscopy using antibodies directed to cytoplasmic or anticytoplasmic epitopes revealed that the vesicles are oriented "cytoplasmic-side inward," consistent with the unique orientation of exosomes. The vesicles were small (<100 nm), consistent with studies of MVBs and exosomes from other tissues. Proteomic analysis of urinary vesicles through nanospray liquid chromatography-tandem mass spectrometry identified numerous protein components of MVBs and of the endosomal pathway in general. Full liquid chromatography-tandem MS analysis revealed 295 proteins, including multiple protein products of genes already known to be responsible for renal and systemic diseases, including autosomal dominant polycystic kidney disease, Gitelman syndrome, Bartter syndrome, autosomal recessive syndrome of osteopetrosis with renal tubular acidosis, and familial renal hypomagnesemia. The results indicate that exosome isolation may provide an efficient first step in biomarker discovery in urine.
1,941 citations
TL;DR: The fate of the transferrin receptor during in vitro maturation of sheep reticulocytes has been followed using FITC- and 125I-labeled anti-transferrin-receptor antibodies and it can be shown that at 0 degree C or in phosphate-buffered saline the rate of vesicle release is less than that at 37 degrees C in culture medium.
1,543 citations
"Exosome-mediated transfer of mRNAs ..." refers background in this paper
...Many cells have the capacity to release exosomes, including reticulocyte...
[...]
TL;DR: ES-MV isolated from murine ES cells in serum-free cultures significantly enhanced survival and improved expansion of murine HPC, and upregulated the expression of early pluripotent and early hematopoietic stem cells in these cells.
Abstract: Membrane-derived vesicles (MV) are released from the surface of activated eucaryotic cells and exert pleiotropic effects on surrounding cells. Since the maintenance of pluripotency and undifferentiated propagation of embryonic stem (ES) cells in vitro requires tight cell to cell contacts and effective intercellular signaling, we hypothesize that MV derived from ES cells (ES-MV) express stem cell-specific molecules that may also support self-renewal and expansion of adult stem cells. To address this hypothesis, we employed expansion of hematopoietic progenitor cells (HPC) as a model. We found that ES-MV (10 microg/ml) isolated from murine ES cells (ES-D3) in serum-free cultures significantly (i) enhanced survival and improved expansion of murine HPC, (ii) upregulated the expression of early pluripotent (Oct-4, Nanog and Rex-1) and early hematopoietic stem cells (Scl, HoxB4 and GATA 2) markers in these cells, and (iii) induced phosphorylation of MAPK p42/44 and serine-threonine kinase AKT. Furthermore, molecular analysis revealed that ES-MV express Wnt-3 protein and are selectively highly enriched in mRNA for several pluripotent transcription factors as compared to parental ES cells. More important, this mRNA could be delivered by ES-MV to target cells and translated into the corresponding proteins. The biological effects of ES-MV were inhibited after heat inactivation or pretreatment with RNAse, indicating a major involvement of protein and mRNA components of ES-MV in the observed phenomena. We postulate that ES-MV may efficiently expand HPC by stimulating them with ES-MV expressed ligands (e.g., Wnt-3) as well as increase their pluripotency after horizontal transfer of ES-derived mRNA.
1,464 citations