Introduction to Extracellular Vesicles Biogenesis

Authors: Erik R Abels Xandra O Breakefield

Publish Date: 2016/04/06

Volume: 36, Issue: 3, Pages: 301-312

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Abstract

Extracellular vesicles are a heterogeneous group of membranelimited vesicles loaded with various proteins lipids and nucleic acids Release of extracellular vesicles from its cell of origin occurs either through the outward budding of the plasma membrane or through the inward budding of the endosomal membrane resulting in the formation of multivesicular bodies which release vesicles upon fusion with the plasma membrane The release of vesicles can facilitate intercellular communication by contact with or by internalization of contents either by fusion with the plasma membrane or by endocytosis into “recipient” cells Although the interest in extracellular vesicle research is increasing there are still no real standards in place to separate or classify the different types of vesicles This review provides an introduction into this expanding and complex field of research focusing on the biogenesis nucleic acid cargo loading content release and uptake of extracellular vesiclesExtracellular vesicles EVs are a heterogeneous family of membranelimited vesicles originating from the endosome or plasma membrane Pan and Johnstone 1983 were among the first to describe EVs Initially it was shown that the release of EVs was part of a disposal mechanism to discard unwanted materials from cells Subsequent research has shown that the release of EVs is also an important mediator of intercellular communication that is involved in normal physiological process as well as in pathological progression Frühbeis et al 2012 2013 Marcilla et al 2012 Luga et al 2012 RegevRudzki et al 2013 Barteneva et al 2013EVs are currently classified based on their mode of release or size EVs can be released by “donor” cells either through the outward budding of the plasma membrane termed shedding microvesicles MVs or ectosomes Minciacchi et al 2015 Another release process involves the inward budding of the endosomal membrane resulting in the formation of multivesicular bodies MVBs with exosomes released by fusion of the outer MVB membrane to the plasma membrane Théry et al 2009 Denzer et al 2000 Vesicles may also be released from nanotubular structures extending from the plasma membrane Rilla et al 2013 2014 In addition to the differences in the mode of release the size of the vesicles is also used for characterization Although different scales are used MVs range from 50 to 10000 nm and exosomes are smaller with a diameter of 30 to 150 nm György et al 2011 Baietti et al 2012 Colombo et al 2013 Overall EVs comprise a wide variety of vesicles ranging from 30 to 1000 nm in size with a variety of cargos and the different types of vesicles overlap in their size distribution It must be emphasized that there is some controversy on nomenclature and sizes of the different types of vesicles Gould and Raposo 2013 Witwer et al 2013 however basic requirements of criteria for EVs have been established Lötvall et al 2014 So far no real standards have been set to classify the different types of vesicles so one should be careful with the use of size alone in defining different types of vesicles In the future the mode of biogenesis means of isolation and cargo may turn out to be far more important criteria Given how the different isolation methods may influence the nature of EVs methods should be compared in order to develop a gold standard for the different protocols and measurements MomenHeravi et al 2012 To be able to compare results it must be stressed that publications on EVs need to clarify their isolation methods in detail and in general term EVs should be used unless there are specific markers defined to classify the different types of vesiclesSo far extensive evidence on all these different types of vesicles indicates that EVs are a key player in the intercellular communication between cells along with secretion of small soluble molecules the secretome and cell–cell contact Raposo and Stoorvogel 2013 Cocucci et al 2009 Once released the EVs can be internalized via endocytosis or membrane fusion releasing their contents into “recipient” cells Mulcahy et al 2014 Recent studies have shown that these EVs contain various proteins sugars lipids and a wide variety of genetic materials such as DNA mRNA and noncoding ncRNAs with the content protected from proteases and nucleases in the extracellular space by the limiting membrane Henderson and Azorsa 2012 Théry et al 2002 EVs have the potential to deliver combinatorial information to multiple cells in their tissue microenvironment and throughout the body BajKrzyworzeka et al 2006 Ratajczak et al 2006 Skog et al 2008This review provides an introduction into the world of EVs focusing primarily subtypes labeled as exosomes and MVs and discusses basics of the biogenesis nucleic acid cargo loading content release and uptake of these vesicles Thus it provides the necessary background for interpretation of the articles in this Special Issue on the role of EVs in the neurobiology and diseases of the nervous systemAs EVs have traditionally been classified based on differences in biogenesis we will focus on the different molecular mechanisms resulting in either the release of vesicles upon the fusion of the MVBs with the plasma membrane or the release via the outward budding and fission of the plasma membrane Akers et al 2013Exosomes are derived from the endosomal system and are formed as intraluminal vesicles ILVs in the MVBs This network of ILVs is used to degrade recycle or exocytose proteins lipids and nucleic acids Within the endosomal system or endocytic pathway the endosomes are divided into different compartments—early endosomes late endosomes and recycling endosomes Grant and Donaldson 2009 Endosomes form by invagination of the plasma membrane The early endosomes can fuse with endocytic vesicles at which point the content is destined for degradation recycling or secretion Contents to be recycled are sorted into recycling endosomes Morelli et al 2004 The remaining early endosomes transform into late endosomes Stoorvogel et al 1991 The late endosomes accumulate ILVs formed by inward budding of the endosomal membrane During this process cytosolic proteins nucleic acids and lipids are sorted into these small vesicles Late endosomes containing a multitude of small vesicles are termed MVBs These MVBs can either fuse with the lysosome if the content is fated for degradation or fuse with the cellular membrane releasing the ILVs as exosomes into the extracellular space Grant and Donaldson 2009Molecular mechanisms of ESCRTdependent and independent MVB biogenesis Multiple biogenesis machineries have been described for generating ILVs in MVBs a ESCRTdependent MVB biogenesis requires the ESCRT protein and ESCRTassociated proteins ALIX TSG101 Chmp4 and SKD1 to form MVBs containing CD63 MHC II ubiquitinated proteins and KFERQcontaining proteins b Three ESCRTindependent pathways are controlled by different proteins 1 heparanase and ARF6/PLD2 associated with the presence of syntenin1 syndecan and CD63 in exosomes 2 nSMase in which the exosomes are enriched with PLP CD63 CD81 and TSG101 Components in image derived from Servier Medical Art Powerpoint image bank Servier 2016

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