title: In vivo delivery of cytoplasmic RNA virus-derived miRNAs.
abstract: The discovery of microRNAs (miRNAs) revealed an unappreciated level of post-transcriptional control used by the cell to maintain optimal protein levels. This process has represented an attractive strategy for therapeutics that is currently limited by in vivo delivery constraints. Here, we describe the generation of a single-stranded, cytoplasmic virus of negative polarity capable of producing functional miRNAs. Cytoplasmic RNA virus-derived miRNAs accumulated to high levels in vitro, generated significant amounts of miRNA star strand, associated with the RNA-induced silencing complex (RISC), and conferred post transcriptional gene silencing in a sequence-specific manner. Furthermore, we demonstrate that these vectors could deliver miRNAs to a wide range of tissues, and sustain prolonged expression capable of achieving measurable knockdown of physiological targets in vivo. Taken together, these results validate noncanonical processing of cytoplasmic-derived miRNAs and provide a novel platform for small RNA delivery.
authors.affiliation: Department of Microbiology, Mount Sinai School of Medicine, New York, New York 10029, USA.
authors.forename: Ryan A
authors.initials: RA
authors.lastname: Langlois
authors.forename: Jillian S
authors.initials: JS
authors.lastname: Shapiro
authors.forename: Alissa M
authors.initials: AM
authors.lastname: Pham
authors.forename: Benjamin R
authors.initials: BR
authors.lastname: tenOever
mesh_terms: D000818:Animals; D060565:Argonaute Proteins / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D003593:Cytoplasm / Q000378:metabolism*; D023061:Gene Order; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D018345:Mice, Knockout; D035683:MicroRNAs / Q000096:biosynthesis / Q000235:genetics* / Q000378:metabolism; D012328:RNA Viruses / Q000235:genetics*; D034743:RNA-Induced Silencing Complex / Q000378:metabolism
chemical_list: C526735:Ago2 protein, mouse; D060565:Argonaute Proteins; D035683:MicroRNAs; D034743:RNA-Induced Silencing Complex
references.citation: Nature. 2009 Jul 23;460(7254):479-86
references.pmid: 19536157
references.citation: Nat Rev Genet. 2000 Nov;1(2):91-9
references.pmid: 11253666
references.citation: Cancer Cell. 2003 Oct;4(4):263-75
references.pmid: 14585354
references.citation: Mol Cell. 2007 Aug 3;27(3):435-48
references.pmid: 17679093
references.citation: J Clin Invest. 2010 Sep;120(9):3106-19
references.pmid: 20697157
references.citation: Mol Cell. 2007 Oct 26;28(2):328-36
references.pmid: 17964270
references.citation: Nature. 2006 May 25;441(7092):537-41
references.pmid: 16724069
references.citation: Nature. 2005 Feb 17;433(7027):769-73
references.pmid: 15685193
references.citation: Nat Genet. 2007 Mar;39(3):380-5
references.pmid: 17259983
references.citation: Proc Natl Acad Sci U S A. 2010 Jun 22;107(25):11525-30
references.pmid: 20534471
references.citation: Nature. 2001 Jan 18;409(6818):363-6
references.pmid: 11201747
references.citation: J Am Vet Med Assoc. 1958 Jul 1;133(1):57-62
references.pmid: 13549332
references.citation: Nat Protoc. 2008;3(6):1077-84
references.pmid: 18536652
references.citation: Genes Dev. 2011 Sep 15;25(18):1881-94
references.pmid: 21896651
references.citation: Nucleic Acids Res. 2010 Dec;38(22):8328-37
references.pmid: 20705652
references.citation: Hum Gene Ther. 2010 Apr;21(4):451-62
references.pmid: 19911974
references.citation: Mol Cell. 2002 Jun;9(6):1327-33
references.pmid: 12086629
references.citation: Cell. 1992 Jun 12;69(6):1011-20
references.pmid: 1318785
references.citation: Nat Methods. 2005 Apr;2(4):269-76
references.pmid: 15782219
references.citation: Cell Host Microbe. 2008 Sep 11;4(3):239-48
references.pmid: 18779050
references.citation: J Virol. 2010 Feb;84(3):1550-62
references.pmid: 19906911
references.citation: Nat Biotechnol. 2011 Jun 07;29(6):499-500
references.pmid: 21654668
references.citation: Genes Dev. 2005 Feb 15;19(4):445-52
references.pmid: 15713840
references.citation: Mol Cell. 2011 Mar 18;41(6):733-46
references.pmid: 21353615
references.citation: Annu Rev Biochem. 1999;68:649-86
references.pmid: 10872463
references.citation: Cell. 2007 Jul 13;130(1):89-100
references.pmid: 17599402
references.citation: Nature. 2008 Sep 4;455(7209):58-63
references.pmid: 18668040
references.citation: PLoS Pathog. 2010 Mar 19;6(3):e1000820
references.pmid: 20333250
references.citation: Nat Biotechnol. 2007 Dec;25(12):1457-67
references.pmid: 18026085
references.citation: Curr Biol. 2011 Mar 8;21(5):369-76
references.pmid: 21353554
references.citation: Cell. 2009 Jan 23;136(2):215-33
references.pmid: 19167326
references.citation: Cell. 2003 Oct 17;115(2):209-16
references.pmid: 14567918
references.citation: Nature. 2003 Sep 25;425(6956):415-9
references.pmid: 14508493
references.citation: Nature. 2005 Aug 4;436(7051):740-4
references.pmid: 15973356
references.citation: Genes Dev. 2001 Oct 15;15(20):2654-9
references.pmid: 11641272
references.citation: Nat Med. 2008 Nov;14(11):1278-83
references.pmid: 18953352
references.citation: Mol Ther. 2008 Aug;16(8):1437-43
references.pmid: 18560417
references.citation: RNA. 2010 Nov;16(11):2068-74
references.pmid: 20841420
references.citation: Proc Natl Acad Sci U S A. 2010 Jun 22;107(25):11519-24
references.pmid: 20534531
references.citation: Genes Dev. 2007 Aug 15;21(16):1999-2004
references.pmid: 17626790
references.citation: Science. 1994 Jun 24;264(5167):1918-21
references.pmid: 8009221
references.citation: Cell. 2003 Oct 17;115(2):199-208
references.pmid: 14567917
references.citation: Nucleic Acids Res. 2007;35(15):5154-64
references.pmid: 17660190
references.citation: Nature. 2008 Sep 4;455(7209):64-71
references.pmid: 18668037
references.citation: Proc Natl Acad Sci U S A. 1995 May 9;92(10):4477-81
references.pmid: 7753828
references.citation: PLoS Pathog. 2010 Feb 12;6(2):e1000764
references.pmid: 20169186
references.citation: Trends Mol Med. 2004 May;10(5):210-6
references.pmid: 15121047
references.citation: J Virol. 2006 Jul;80(14):7028-36
references.pmid: 16809308
references.citation: RNA Biol. 2011 Mar-Apr;8(2):190-4
references.pmid: 21593582
references.citation: Nat Biotechnol. 2009 Jun;27(6):572-6
references.pmid: 19483680