Abstract
MicroRNAs (miRNAs) are single stranded noncoding RNAs of approximately 22 nucleotides that act as posttranscriptional gene regulators by binding partially complementary sequences in the 3′ untranslated region (3′-UTR) of target messenger RNAs (mRNAs). MicroRNAs regulate many biological processes including embryonal development, differentiation, apoptosis, and proliferation and the targets of miRNAs range from signalling proteins and transcription factors to RNA binding proteins. Recently, variations in the expression of certain miRNAs have been linked to a variety of human diseases including cancer and viral infections, validating miRNAs as potential targets for drug discovery. Several tools have been developed to control the function of individual miRNAs and have been applied to study their biological role and therapeutic potential; however, common methods lack a precise level of control that allows for the study of miRNA function with high spatial and temporal resolution. Toward this goal, a light-activated miRNA antagomir for mature miR-21 was developed through the site-specific installation of caging groups on the bases of selected nucleotides. Installation of caged nucleotides led to complete inhibition of the antagomir–miRNA hybridization and inactivation of antagomir function. The miRNA-inhibitory activity of the caged antagomirs was fully restored upon decaging through a brief UV irradiation. The synthesized antagomir was applied to the photochemical regulation of miR-21 function in mammalian cells. Moreover, spatial and temporal control over antagomir activity and thus miR-21 function was obtained in mammalian cells. The presented approach enables the precise regulation of miRNA function with unprecedented spatial and temporal resolution using UV irradiation and can be readily extended to any miRNA of interest.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
Carthew R (2006) Gene regulation by microRNAs. Curr Opin Genet Dev 16:203–208
Djuranovic S, Nahvi A, Green R (2012) miRNA-mediated gene silencing by translational repression followed by mRNA deadenylation and decay. Science 336:237–240
Pillai RS (2005) MicroRNA function: multiple mechanisms for a tiny RNA? RNA 11: 1753–1761
Lewis BP, Burge CB, Bartel DP (2005) Conserved seed pairing, often flanked by adenosines, indicates that thousands of human genes are microRNA targets. Cell 120:15–20
Appasani K (2008) MicroRNAs: from basic science to disease biology. Cambridge University Press, Cambridge
Janga SC, Vallabhaneni S (2011) MicroRNAs as post-transcriptional machines and their interplay with cellular networks. Adv Exp Med Biol 722:59–74
Tong AW, Nemunaitis J (2008) Modulation of miRNA activity in human cancer: a new paradigm for cancer gene therapy? Cancer Gene Ther 15:341–355
Sevignani C, Calin G, Siracusa L, Croce C (2006) Mammalian microRNAs: a small world for fine-tuning gene expression. Mamm Genome 17:189–202
Port JD, Sucharov C (2010) Role of microRNAs in cardiovascular disease: therapeutic challenges and potentials. J Cardiovasc Pharmacol 56:444–453
Lindsay MA (2008) microRNAs and the immune response. Trends Immunol 29:343–351
Cullen BR (2011) Viruses and microRNAs: RISCy interactions with serious consequences. Genes Dev 25:1881–1894
Esquela-Kerscher A, Slack F (2006) Oncomirs – microRNAs with a role in cancer. Nat Rev Cancer 6:259–269
Esau C (2008) Inhibition of microRNA with antisense oligonucleotides. Methods 44: 55–60
Veedu R, Wengel J (2010) Locked nucleic acids: promising nucleic acid analogs for therapeutic applications. Chem Biodiver 7:536–542
Brown B, Naldini L (2009) Exploiting and antagonizing microRNA regulation for therapeutic and experimental applications. Nat Rev Genet 10:578–585
Deiters A (2010) Small molecule modifiers of the microRNA and RNA interference pathway. AAPS J 12:51–60
Meister G, Landthaler M, Dorsett Y, Tuschl T (2004) Sequence-specific inhibition of microRNA- and siRNA-induced RNA silencing. RNA 10:544–550
Krützfeldt J, Rajewsky N, Braich R, Rajeev K, Tuschl T, Manoharan M et al (2005) Silencing of microRNAs in vivo with 'antagomirs'. Nature 438:685–689
Ebert MS, Neilson JR, Sharp PA (2007) MicroRNA sponges: competitive inhibitors of small RNAs in mammalian cells. Nat Methods 4:721–726
Kota J, Chivukula RR, O'Donnell KA, Wentzel EA, Montgomery CL, Hwang HW et al (2009) Therapeutic microRNA delivery suppresses tumorigenesis in a murine liver cancer model. Cell 137:1005–1017
Gumireddy K, Young D, Xiong X, Hogenesch J, Huang Q, Deiters A (2008) Small-molecule inhibitors of microrna miR-21 function. Angew Chem Int Ed Engl 47:7482–7484
Young D, Connelly C, Grohmann C, Deiters A (2010) Small molecule modifiers of microRNA miR-122 function for the treatment of hepatitis C virus infection and hepatocellular carcinoma. J Am Chem Soc 132:7976–7981
Liu Z, Sall A, Yang D (2008) MicroRNA: an emerging therapeutic target and intervention tool. Int J Mol Sci 9:978–999
Grünweller A, Hartmann R (2007) Locked nucleic acid oligonucleotides: the next generation of antisense agents? BioDrugs 21:235–243
Riggsbee CW, Deiters A (2010) Recent advances in the photochemical control of protein function. Trends Biotechnol 28:468–475
Deiters A (2010) Principles and applications of the photochemical control of cellular processes. Chembiochem 11:47–53
Deiters A (2009) Light activation as a method of regulating and studying gene expression. Curr Opin Chem Biol 13:678–686
Young DD, Deiters A (2007) Photochemical control of biological processes. Org Biomol Chem 5:999–1005
Fenno L, Yizhar O, Deisseroth K (2011) The development and application of optogenetics. Annu Rev Neurosci 34:389–412
Prokup A, Hemphill J, Deiters A (2012) DNA computation: a photochemically controlled AND gate. J Am Chem Soc 134:3810–3815
Govan JM, Lively MO, Deiters A (2011) Photochemical control of DNA decoy function enables precise regulation of nuclear factor κB activity. J Am Chem Soc 133:13176–13182
Young D, Lively M, Deiters A (2010) Activation and deactivation of DNAzyme and antisense function with light for the photochemical regulation of gene expression in mammalian cells. J Am Chem Soc 132:6183–6193
Young DD, Lusic H, Lively MO, Yoder JA, Deiters A (2008) Gene silencing in mammalian cells with light-activated antisense agents. Chembiochem 9:2937–2940
Young DD, Edwards WF, Lusic H, Lively MO, Deiters A (2008) Light-triggered polymerase chain reaction. Chem Commun (Camb) 462–4
Joshi KB, Vlachos A, Mikat V, Deller T, Heckel A (2012) Light-activatable molecular beacons with a caged loop sequence. Chem Commun (Camb) 48:2746–2748
Mikat V, Heckel A (2007) Light-dependent RNA interference with nucleobase-caged siRNAs. RNA 13:2341–2347
Medina PP, Nolde M, Slack FJ (2010) OncomiR addiction in an in vivo model of microRNA-21-induced pre-B-cell lymphoma. Nature 467:86–90
Connelly CM, Uprety R, Hemphill J, Deiters A (2012) Spatiotemporal control of microRNA function using light-activated antagomirs. Mol Biosyst 8:2987–2993
Furusawa K, Ueno K, Katsura T (1990) Synthesis and restricted conformation of 3′,5′-O-(di-tert-butylsilanediyl) ribonucleosides. Chem Lett 97–100
Lusic H, Deiters A (2006) A new photocaging group for aromatic N-heterocycles. Synth Stutt 2147–50
Iyer R, Egan W, Regan J, Beaucage S (1990) 3H-1,2-benzodithiole-3-one 1,1-dioxide as an improved sulfurizing reagent in the solid-phase synthesis of oligodeoxyribonucleoside phosphorothioates. J Am Chem Soc 112:1253–1254
Govan JM, Uprety R, Hemphill J, Lively MO, Deiters A (2012) Regulation of transcription through light-activation and light-deactivation of triplex-forming oligonucleotides in mammalian cells. ACS Chem Biol 7:1247–1256
Lusic H, Lively MO, Deiters A (2008) Light-activated deoxyguanosine: photochemical regulation of peroxidase activity. Mol Biosyst 4:508–511
Akiyama T, Nishimoto H, Ozaki S (1990) The selective protection of uridine with a para-methoxybenzyl-chloride- a synthesis of 2′-O-methyluridine. Bull Chem Soc Jpn 63:3356–3357
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2014 Springer Science+Business Media New York
About this protocol
Cite this protocol
Connelly, C.M., Deiters, A. (2014). Control of Oncogenic miRNA Function by Light-Activated miRNA Antagomirs. In: Robles-Flores, M. (eds) Cancer Cell Signaling. Methods in Molecular Biology, vol 1165. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-0856-1_9
Download citation
DOI: https://doi.org/10.1007/978-1-4939-0856-1_9
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-0855-4
Online ISBN: 978-1-4939-0856-1
eBook Packages: Springer Protocols