title: Three-dimensional architecture of tick-borne encephalitis virus replication sites and trafficking of the replicated RNA.
abstract: Flavivirus replication is accompanied by the rearrangement of cellular membranes that may facilitate viral genome replication and protect viral components from host cell responses. The topological organization of viral replication sites and the fate of replicated viral RNA are not fully understood. We exploited electron microscopy to map the organization of tick-borne encephalitis virus (TBEV) replication compartments in infected cells and in cells transfected with a replicon. Under both conditions, 80-nm vesicles were seen within the lumen of the endoplasmic reticulum (ER) that in infected cells also contained virions. By electron tomography, the vesicles appeared as invaginations of the ER membrane, displaying a pore that could enable release of newly synthesized viral RNA into the cytoplasm. To track the fate of TBEV RNA, we took advantage of our recently developed method of viral RNA fluorescent tagging for live-cell imaging combined with bleaching techniques. TBEV RNA was found outside virus-induced vesicles either associated to ER membranes or free to move within a defined area of juxtaposed ER cisternae. From our results, we propose a biologically relevant model of the possible topological organization of flavivirus replication compartments composed of replication vesicles and a confined extravesicular space where replicated viral RNA is retained. Hence, TBEV modifies the ER membrane architecture to provide a protected environment for viral replication and for the maintenance of newly replicated RNA available for subsequent steps of the virus life cycle.
authors.affiliation: Laboratory of Molecular Virology, The International Center for Genetic Engineering and Biotechnology (ICGEB), Padriciano, Trieste, Italy.
authors.forename: Lisa
authors.initials: L
authors.lastname: Miorin
authors.forename: Inés
authors.initials: I
authors.lastname: Romero-Brey
authors.forename: Paolo
authors.initials: P
authors.lastname: Maiuri
authors.forename: Simone
authors.initials: S
authors.lastname: Hoppe
authors.forename: Jacomine
authors.initials: J
authors.lastname: Krijnse-Locker
authors.forename: Ralf
authors.initials: R
authors.lastname: Bartenschlager
authors.forename: Alessandro
authors.initials: A
authors.lastname: Marcello
mesh_terms: D000818:Animals; D058948:Cell Tracking; D002522:Chlorocebus aethiops; D006224:Cricetinae; D004669:Encephalitis Viruses, Tick-Borne / Q000235:genetics / Q000502:physiology* / Q000648:ultrastructure*; D004675:Encephalitis, Tick-Borne / Q000821:virology*; D004721:Endoplasmic Reticulum / Q000648:ultrastructure / Q000821:virology*; D057809:HEK293 Cells; D006801:Humans; D008854:Microscopy, Electron; D012367:RNA, Viral / Q000235:genetics* / Q000378:metabolism; D014709:Vero Cells; D014779:Virus Replication
chemical_list: D012367:RNA, Viral
references.citation: EMBO Rep. 2011 Dec 01;12(12):1280-5
references.pmid: 22015688
references.citation: Cell Host Microbe. 2010 Jul 22;8(1):77-85
references.pmid: 20638644
references.citation: J Virol. 1997 Dec;71(12):9608-17
references.pmid: 9371625
references.citation: Virology. 2008 Aug 1;377(2):419-30
references.pmid: 18533218
references.citation: J Virol. 1987 Nov;61(11):3641-4
references.pmid: 2959795
references.citation: Retrovirology. 2007 May 30;4:36
references.pmid: 17537237
references.citation: Methods. 2011 Jan;53(1):62-7
references.pmid: 20600934
references.citation: J Virol. 2006 Jun;80(11):5308-20
references.pmid: 16699011
references.citation: J Biol Chem. 2007 Mar 23;282(12):8873-82
references.pmid: 17276984
references.citation: J Gen Virol. 2009 Aug;90(Pt 8):1781-1794
references.pmid: 19420159
references.citation: Nature. 2000 Apr 6;404(6778):604-9
references.pmid: 10766243
references.citation: J Virol. 2006 Dec;80(24):12197-208
references.pmid: 17035331
references.citation: Cell Host Microbe. 2009 Apr 23;5(4):318-28
references.pmid: 19380111
references.citation: J Virol. 2004 Jul;78(14):7400-9
references.pmid: 15220413
references.citation: Virology. 2008 Sep 15;379(1):64-77
references.pmid: 18667221
references.citation: Virus Res. 2012 Feb;163(2):660-6
references.pmid: 22155022
references.citation: Biol Cell. 2005 Feb;97(2):147-72
references.pmid: 15656780
references.citation: Curr Protoc Cell Biol. 2003 May;Chapter 13:Unit 13.5
references.pmid: 18228420
references.citation: Traffic. 2005 Nov;6(11):967-77
references.pmid: 16190978
references.citation: J Virol. 2012 Jul;86(13):7360-71
references.pmid: 22553322
references.citation: Virology. 1996 Jun 1;220(1):232-40
references.pmid: 8659120
references.citation: Lancet. 2008 May 31;371(9627):1861-71
references.pmid: 18514730
references.citation: PLoS One. 2012;7(10):e47912
references.pmid: 23112871
references.citation: Virology. 1987 Apr;157(2):330-7
references.pmid: 3029975
references.citation: J Virol. 2003 Apr;77(7):4370-82
references.pmid: 12634393
references.citation: J Virol. 1997 Feb;71(2):1497-505
references.pmid: 8995675
references.citation: Methods Enzymol. 1983;96:466-85
references.pmid: 6656640
references.citation: Virology. 1999 May 25;258(1):108-17
references.pmid: 10329573
references.citation: Virus Res. 1996 Aug;43(2):125-36
references.pmid: 8864202
references.citation: J Virol. 2001 Nov;75(22):10787-99
references.pmid: 11602720
references.citation: Biophys J. 1976 Sep;16(9):1055-69
references.pmid: 786399
references.citation: Cell Host Microbe. 2009 Apr 23;5(4):365-75
references.pmid: 19380115
references.citation: Nat Rev Microbiol. 2008 May;6(5):363-74
references.pmid: 18414501
references.citation: J Virol. 2006 May;80(9):4623-32
references.pmid: 16611922
references.citation: Virology. 1998 Jun 5;245(2):203-15
references.pmid: 9636360
references.citation: J Virol. 2010 Sep;84(17):8470-83
references.pmid: 20554782
references.citation: J Cell Biol. 2007 Oct 22;179(2):291-304
references.pmid: 17954611
references.citation: Antiviral Res. 2003 Jan;57(1-2):129-46
references.pmid: 12615309
references.citation: J Gen Virol. 2007 Feb;88(Pt 2):470-475
references.pmid: 17251564
references.citation: J Virol. 1999 Nov;73(11):9555-67
references.pmid: 10516064
references.citation: J Gen Virol. 2007 Nov;88(Pt 11):3013-3017
references.pmid: 17947524
references.citation: J Gen Virol. 1997 May;78 ( Pt 5):1049-57
references.pmid: 9152422
references.citation: J Gen Virol. 2005 May;86(Pt 5):1415-1421
references.pmid: 15831953
references.citation: J Virol. 1999 Dec;73(12):10272-80
references.pmid: 10559344
references.citation: Nat Rev Microbiol. 2006 May;4(5):371-82
references.pmid: 16582931
references.citation: Virology. 2005 Jan 20;331(2):247-59
references.pmid: 15629769
references.citation: J Virol. 2010 Oct;84(20):10438-47
references.pmid: 20686019
references.citation: Virology. 2001 Jan 5;279(1):161-72
references.pmid: 11145899
references.citation: PLoS Biol. 2007 Sep;5(9):e220
references.pmid: 17696647
references.citation: J Virol. 1997 Sep;71(9):6650-61
references.pmid: 9261387
references.citation: J Struct Biol. 1996 Jan-Feb;116(1):71-6
references.pmid: 8742726
references.citation: Nat Rev Mol Cell Biol. 2001 Jun;2(6):444-56
references.pmid: 11389468
references.citation: J Virol. 2008 Nov;82(21):10519-31
references.pmid: 18715913