title: Localization and abundance analysis of human lncRNAs at single-cell and single-molecule resolution.
abstract: BACKGROUND
  Long non-coding RNAs (lncRNAs) have been implicated in diverse biological processes. In contrast to extensive genomic annotation of lncRNA transcripts, far fewer have been characterized for subcellular localization and cell-to-cell variability. Addressing this requires systematic, direct visualization of lncRNAs in single cells at single-molecule resolution.
  
  
  RESULTS
  We use single-molecule RNA-FISH to systematically quantify and categorize the subcellular localization patterns of a representative set of 61 lncRNAs in three different cell types. Our survey yields high-resolution quantification and stringent validation of the number and spatial positions of these lncRNA, with an mRNA set for comparison. Using this highly quantitative image-based dataset, we observe a variety of subcellular localization patterns, ranging from bright sub-nuclear foci to almost exclusively cytoplasmic localization. We also find that the low abundance of lncRNAs observed from cell population measurements cannot be explained by high expression in a small subset of 'jackpot' cells. Additionally, nuclear lncRNA foci dissolve during mitosis and become widely dispersed, suggesting these lncRNAs are not mitotic bookmarking factors. Moreover, we see that divergently transcribed lncRNAs do not always correlate with their cognate mRNA, nor do they have a characteristic localization pattern.
  
  
  CONCLUSIONS
  Our systematic, high-resolution survey of lncRNA localization reveals aspects of lncRNAs that are similar to mRNAs, such as cell-to-cell variability, but also several distinct properties. These characteristics may correspond to particular functional roles. Our study also provides a quantitative description of lncRNAs at the single-cell level and a universally applicable framework for future study and validation of lncRNAs.
authors: Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA, 02142, USA. nmcabili@broadinstitute.org.|Department of Systems Biology, Harvard Medical School, Boston, MA, 02115, USA. nmcabili@broadinstitute.org.|Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA, 02138, USA. nmcabili@broadinstitute.org.
authors: Moran N
authors: MN
authors: Cabili
authors: School of Engineering and Applied Science, University of Pennsylvania, Philadelphia, PA, 19104, USA. dunagin@seas.upenn.edu.
authors: Margaret C
authors: MC
authors: Dunagin
authors: School of Engineering and Applied Science, University of Pennsylvania, Philadelphia, PA, 19104, USA. pmccl@seas.upenn.edu.
authors: Patrick D
authors: PD
authors: McClanahan
authors: School of Engineering and Applied Science, University of Pennsylvania, Philadelphia, PA, 19104, USA. biaescha@gmail.com.
authors: Andrew
authors: A
authors: Biaesch
authors: School of Engineering and Applied Science, University of Pennsylvania, Philadelphia, PA, 19104, USA. opadovan@sas.upenn.edu.
authors: Olivia
authors: O
authors: Padovan-Merhar
authors: Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA, 02142, USA. aregev@broadinstitute.org.|Howard Hughes Medical Institute, Department of Biology, Massachusetts Institute of Technology, Cambridge, MA, 02140, USA. aregev@broadinstitute.org.
authors: Aviv
authors: A
authors: Regev
authors: Broad Institute of MIT and Harvard, 7 Cambridge Center, Cambridge, MA, 02142, USA. john_rinn@harvard.edu.|Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA, 02138, USA. john_rinn@harvard.edu.
authors: John L
authors: JL
authors: Rinn
authors: School of Engineering and Applied Science, University of Pennsylvania, Philadelphia, PA, 19104, USA. arjunraj@seas.upenn.edu.
authors: Arjun
authors: A
authors: Raj
mesh_terms: D000818:Animals; D002467:Cell Nucleus / Q000378:metabolism; D005786:Gene Expression Regulation; D006367:HeLa Cells; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D008297:Male; D051379:Mice; D008938:Mitosis; D034443:RNA Transport / Q000235:genetics; D062085:RNA, Long Noncoding / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D059010:Single-Cell Analysis; D014158:Transcription, Genetic
chemical_list: D062085:RNA, Long Noncoding; D012333:RNA, Messenger
references: Nature. 2013 Jul 18;499(7458):360-3
references: 23792564
references: Nat Protoc. 2013 Sep;8(9):1743-58
references: 23949380
references: Science. 2013 Aug 16;341(6147):789-92
references: 23907535
references: Nat Methods. 2013 Sep;10(9):865-7
references: 23913259
references: J Mol Biol. 2013 Oct 9;425(19):3698-706
references: 23816838
references: Cell. 2013 Nov 21;155(5):990-6
references: 24267885
references: Mol Biol Cell. 2004 Jan;15(1):81-90
references: 14528011
references: Cell Cycle. 2004 Aug;3(8):1057-61
references: 15254409
references: FEBS Lett. 1973 May 15;32(1):109-12
references: 4123728
references: Proc Natl Acad Sci U S A. 1982 Dec;79(23):7331-5
references: 6961411
references: Nature. 1991 Jan 3;349(6304):82-4
references: 1985270
references: Cell. 1992 Oct 30;71(3):515-26
references: 1423610
references: Cell. 1992 Oct 30;71(3):527-42
references: 1423611
references: J Cell Biol. 1996 Feb;132(3):259-75
references: 8636206
references: Science. 1998 Apr 24;280(5363):585-90
references: 9554849
references: Proc Natl Acad Sci U S A. 2005 Nov 22;102(47):17008-13
references: 16284251
references: PLoS Biol. 2006 Oct;4(10):e309
references: 17048983
references: BMC Genomics. 2007;8:39
references: 17270048
references: Genome Res. 2007 May;17(5):556-65
references: 17387145
references: J Cell Sci. 2007 Aug 1;120(Pt 15):2498-506
references: 17623775
references: J Biol Chem. 2007 Aug 24;282(34):24731-42
references: 17569660
references: Proc Natl Acad Sci U S A. 2008 Jan 15;105(2):716-21
references: 18184812
references: Mol Cell Biol. 2008 Jun;28(11):3713-28
references: 18299392
references: Mol Biol Cell. 2008 Sep;19(9):3793-800
references: 18562689
references: Nat Methods. 2008 Oct;5(10):877-9
references: 18806792
references: Dev Cell. 2008 Nov;15(5):668-79
references: 18848501
references: Science. 2008 Dec 12;322(5908):1717-20
references: 18988810
references: Science. 2008 Dec 19;322(5909):1851-4
references: 19056938
references: Science. 2008 Dec 19;322(5909):1849-51
references: 19056940
references: Science. 2008 Dec 19;322(5909):1845-8
references: 19056941
references: Nat Rev Genet. 2009 Mar;10(3):155-9
references: 19188922
references: Proc Natl Acad Sci U S A. 2009 Feb 24;106(8):2525-30
references: 19188602
references: Nature. 2009 Mar 12;458(7235):223-7
references: 19182780
references: Mol Cell. 2009 Mar 27;33(6):717-26
references: 19217333
references: Proc Natl Acad Sci U S A. 2009 Jul 14;106(28):11667-72
references: 19571010
references: Nucleic Acids Res. 2009 Aug;37(14):e100
references: 19515934
references: Elife. 2013;2:e01749
references: 24381249
references: BMC Genomics. 2013;14:914
references: 24365181
references: Nat Struct Mol Biol. 2014 Feb;21(2):198-206
references: 24463464
references: Mol Cell Endocrinol. 2014 Apr 5;386(1-2):16-33
references: 24035864
references: PLoS One. 2014;9(3):e93488
references: 24675777
references: Curr Biol. 2014 Jun 2;24(11):1283-8
references: 24856210
references: Genome Biol. 2013;14(9):R104
references: 24070194
references: Genome Biol. 2012;13(11):R107
references: 23181609
references: Cell Cycle. 2009 Aug 15;8(16):2557-64
references: 19597342
references: Brief Funct Genomic Proteomic. 2009 Nov;8(6):407-23
references: 19770204
references: Sci Signal. 2010;3(107):ra8
references: 20124551
references: Nature. 2010 Feb 18;463(7283):913-8
references: 20164922
references: Nat Biotechnol. 2010 May;28(5):503-10
references: 20436462
references: Mol Cell. 2010 Jun 11;38(5):662-74
references: 20541999
references: Mol Cell. 2010 Sep 24;39(6):925-38
references: 20797886
references: Cell. 2010 Oct 29;143(3):390-403
references: 21029862
references: Genes Dev. 2011 Jan 1;25(1):11-6
references: 21205863
references: Nature. 2011 Apr 7;472(7341):120-4
references: 21423168
references: Genes Cells. 2011 May;16(5):479-90
references: 21463453
references: Trends Genet. 2011 Aug;27(8):295-306
references: 21680045
references: Genes Dev. 2011 Sep 15;25(18):1915-27
references: 21890647
references: Annu Rev Genet. 2011;45:379-403
references: 21942369
references: Cell. 2011 Nov 11;147(4):773-88
references: 22078878
references: Nat Cell Biol. 2012 Jan;14(1):106-14
references: 22119784
references: Cell. 2011 Dec 23;147(7):1537-50
references: 22196729
references: Mol Cell. 2012 Feb 24;45(4):470-82
references: 22264825
references: Genome Res. 2012 Mar;22(3):577-91
references: 22110045
references: J Mol Endocrinol. 2012 Jun;48(3):R45-53
references: 22393162
references: Annu Rev Biochem. 2012;81:145-66
references: 22663078
references: Nat Methods. 2012 Jul;9(7):743-8
references: 22660740
references: Genome Res. 2012 Sep;22(9):1775-89
references: 22955988
references: Nature. 2013 Jan 10;493(7431):231-5
references: 23201690
references: Dev Cell. 2013 Jan 28;24(2):206-14
references: 23369715
references: Proc Natl Acad Sci U S A. 2013 Feb 19;110(8):2876-81
references: 23382218
references: Nat Methods. 2013 Mar;10(3):246-8
references: 23416756
references: Genetics. 2013 Mar;193(3):651-69
references: 23463798
references: Dev Growth Differ. 2012 Jan;54(1):44-54
references: 22070123
references: Genes Dev. 2013 Jun 1;27(11):1260-71
references: 23723417
references: Nature. 2013 Jun 13;498(7453):236-40
references: 23685454
references: Cell. 2013 Jul 3;154(1):26-46
references: 23827673