title: Quantitative imaging of single mRNA splice variants in living cells.
abstract: Alternative messenger RNA (mRNA) splicing is a fundamental process of gene regulation, and errors in RNA splicing are known to be associated with a variety of different diseases. However, there is currently a lack of quantitative technologies for monitoring mRNA splice variants in cells. Here, we show that a combination of plasmonic dimer probes and hyperspectral imaging can be used to detect and quantify mRNA splice variants in living cells. The probes are made from gold nanoparticles functionalized with oligonucleotides and can hybridize to specific mRNA sequences, forming nanoparticle dimers that exhibit distinct spectral shifts due to plasmonic coupling. With this approach, we show that the spatial and temporal distribution of three selected splice variants of the breast cancer susceptibility gene, BRCA1, can be monitored at single-copy resolution by measuring the hybridization dynamics of the nanoplasmonic dimers. Our study provides insights into RNA and its transport in living cells, which could improve our understanding of cellular protein complexes, pharmacogenomics, genetic diagnosis and gene therapies.
authors.affiliation: 1] Department of Bioengineering, Department of Electrical Engineering and Computer Science, University of California Berkeley, Berkeley, California 94720, USA [2] Department of Agricultural and Biological Engineering, Bindley Bioscience Center, Purdue University, 225 South University Street, West Lafayette, Indiana 47907, USA.
authors.forename: Kyuwan
authors.initials: K
authors.lastname: Lee
authors.affiliation: Department of Agricultural and Biological Engineering, Bindley Bioscience Center, Purdue University, 225 South University Street, West Lafayette, Indiana 47907, USA.
authors.forename: Yi
authors.initials: Y
authors.lastname: Cui
authors.affiliation: Department of Bioengineering, Department of Electrical Engineering and Computer Science, University of California Berkeley, Berkeley, California 94720, USA.
authors.forename: Luke P
authors.initials: LP
authors.lastname: Lee
authors.affiliation: Department of Agricultural and Biological Engineering, Bindley Bioscience Center, Purdue University, 225 South University Street, West Lafayette, Indiana 47907, USA.
authors.forename: Joseph
authors.initials: J
authors.lastname: Irudayaraj
mesh_terms: D017398:Alternative Splicing / Q000502:physiology*; D019313:BRCA1 Protein / Q000096:biosynthesis*; D045744:Cell Line, Tumor; D006801:Humans; D062048:Narrow Band Imaging / Q000295:instrumentation / Q000379:methods*; D012333:RNA, Messenger / Q000737:chemistry / Q000502:physiology*; D020349:Surface Plasmon Resonance / Q000295:instrumentation / Q000379:methods*
chemical_list: D019313:BRCA1 Protein; C492913:BRCA1 protein, human; D012333:RNA, Messenger
references.citation: Science. 2004 Jun 18;304(5678):1797-800
references.pmid: 15205532
references.citation: Biochem Biophys Res Commun. 2001 Jan 12;280(1):32-8
references.pmid: 11162473
references.citation: Nat Nanotechnol. 2008 Sep;3(9):548-51
references.pmid: 18772916
references.citation: Nat Biotechnol. 2004 Jul;22(7):883-7
references.pmid: 15170215
references.citation: Science. 2011 Apr 22;332(6028):475-8
references.pmid: 21512033
references.citation: Ann Biomed Eng. 2006 Jan;34(1):39-50
references.pmid: 16463087
references.citation: Genes Dev. 2009 Mar 15;23(6):740-54
references.pmid: 19261746
references.citation: J Phys Chem B. 2009 Oct 22;113(42):14021-5
references.pmid: 19780515
references.citation: Mol Pathol. 2003 Aug;56(4):191-7
references.pmid: 12890739
references.citation: Biophys J. 2000 Jun;78(6):3260-74
references.pmid: 10828002
references.citation: Nat Genet. 1999 May;22(1):37-43
references.pmid: 10319859
references.citation: Genes Dev. 2009 Mar 15;23(6):719-28
references.pmid: 19261748
references.citation: Nature. 2010 Sep 30;467(7315):604-7
references.pmid: 20844488
references.citation: Angew Chem Int Ed Engl. 2006 Dec 4;45(47):8022-5
references.pmid: 17091512
references.citation: Appl Opt. 1996 Jun 20;35(18):3193-9
references.pmid: 21102699
references.citation: Science. 1997 Aug 22;277(5329):1078-81
references.pmid: 9262471
references.citation: Oncogene. 1997 Jan 9;14(1):1-16
references.pmid: 9010228
references.citation: Proc Natl Acad Sci U S A. 2003 Sep 30;100(20):11350-5
references.pmid: 13679586
references.citation: Small. 2013 Apr 8;9(7):1106-15
references.pmid: 23281179
references.citation: Nano Lett. 2009 Feb;9(2):562-70
references.pmid: 19128006
references.citation: Proc Natl Acad Sci U S A. 2007 Feb 20;104(8):2667-72
references.pmid: 17307879
references.citation: Nature. 1996 Aug 15;382(6592):607-9
references.pmid: 8757129
references.citation: Nat Methods. 2008 Oct;5(10):877-9
references.pmid: 18806792
references.citation: Nano Lett. 2007 Aug;7(8):2300-6
references.pmid: 17602538
references.citation: Anal Chem. 2008 May 1;80(9):3342-9
references.pmid: 18341356
references.citation: Science. 2000 Sep 8;289(5485):1757-60
references.pmid: 10976070
references.citation: Nat Nanotechnol. 2012 Dec;7(12):821-4
references.pmid: 23103935
references.citation: Biophys J. 2009 Jun 3;96(11):4709-16
references.pmid: 19486693
references.citation: Cell. 1980 Nov;22(2 Pt 2):479-88
references.pmid: 6256082
references.citation: Cell. 2011 Dec 23;147(7):1484-97
references.pmid: 22196726
references.citation: Science. 2002 Aug 2;297(5582):836-40
references.pmid: 12161654
references.citation: Nature. 2014 Jan 30;505(7485):701-5
references.pmid: 24336214
references.citation: Genes Dev. 2009 Mar 15;23(6):729-39
references.pmid: 19261749
references.citation: ACS Nano. 2009 Aug 25;3(8):2147-52
references.pmid: 19702321
references.citation: J Am Chem Soc. 2007 Dec 19;129(50):15477-9
references.pmid: 18034495
references.citation: Nature. 1996 Aug 15;382(6592):609-11
references.pmid: 8757130
references.citation: Nat Nanotechnol. 2006 Oct;1(1):47-52
references.pmid: 18654141
references.citation: Oncogene. 1999 Nov 18;18(48):6684-91
references.pmid: 10597274
references.citation: Proc Natl Acad Sci U S A. 2003 Nov 11;100(23):13308-13
references.pmid: 14583593
references.citation: Nat Biotechnol. 2005 Jun;23(6):741-5
references.pmid: 15908940
references.citation: Exp Cell Res. 1999 Feb 25;247(1):148-59
references.pmid: 10047457
references.citation: ACS Nano. 2011 Mar 22;5(3):2109-17
references.pmid: 21314177
references.citation: Anal Biochem. 1998 Sep 10;262(2):157-76
references.pmid: 9750129
references.citation: Proc Natl Acad Sci U S A. 2009 Oct 20;106(42):17735-40
references.pmid: 19805121