Broccoli Fluorets: Split Aptamers as a User-Friendly Fluorescent Toolkit for Dynamic RNA Nanotechnology
Abstract
:1. Introduction
2. Results and Discussion
3. Materials and Methods
3.1. Design of Broccoli Fluorets
3.2. RNA Preparation
3.3. Broccoli Aptamer and Fluoret Assembly
3.4. Co-Transcriptional Assembly
3.5. Electrophoretic Mobility Shift Assays
3.6. EDTA Degradation and Mg2+ Formation
3.7. Nuclease-Driven Assembly/Degradation
3.8. Strand Displacement
3.9. Thermal Deactivation/Activation
3.10. Blood Stability
3.11. Statistics
4. Conclusions
Supplementary Materials
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
References
- Shukla, G.C.; Haque, F.; Tor, Y.; Wilhelmsson, L.M.; Toulme, J.J.; Isambert, H.; Guo, P.; Rossi, J.J.; Tenenbaum, S.A.; Shapiro, B.A. A boost for the emerging field of RNA nanotechnology. ACS Nano 2011, 5, 3405–3418. [Google Scholar] [CrossRef] [PubMed]
- Afonin, K.A.; Kasprzak, W.K.; Bindewald, E.; Kireeva, M.; Viard, M.; Kashlev, M.; Shapiro, B.A. In silico design and enzymatic synthesis of functional RNA nanoparticles. Acc. Chem. Res. 2014, 47, 1731–1741. [Google Scholar] [CrossRef] [PubMed]
- Pinheiro, A.V.; Han, D.; Shih, W.M.; Yan, H. Challenges and opportunities for structural DNA nanotechnology. Nat. Nanotechnol. 2011, 6, 763–772. [Google Scholar] [CrossRef] [PubMed][Green Version]
- Hong, E.; Halman, J.R.; Shah, A.B.; Khisamutdinov, E.F.; Dobrovolskaia, M.A.; Afonin, K.A. Structure and composition define immunorecognition of nucleic acid nanoparticles. Nano Lett. 2018, 18, 4309–4321. [Google Scholar] [CrossRef] [PubMed]
- Leontis, N.B.; Stombaugh, J.; Westhof, E. The non-watson-crick base pairs and their associated isostericity matrices. Nucleic Acids Res. 2002, 30, 3497–3531. [Google Scholar] [CrossRef] [PubMed]
- Jasinski, D.; Haque, F.; Binzel, D.W.; Guo, P. Advancement of the emerging field of RNA nanotechnology. ACS Nano 2017, 11, 1142–1164. [Google Scholar] [CrossRef] [PubMed]
- Afonin, K.A.; Viard, M.; Koyfman, A.Y.; Martins, A.N.; Kasprzak, W.K.; Panigaj, M.; Desai, R.; Santhanam, A.; Grabow, W.W.; Jaeger, L.; et al. Multifunctional RNA nanoparticles. Nano Lett. 2014, 14, 5662–5671. [Google Scholar] [CrossRef]
- Shibata, T.; Fujita, Y.; Ohno, H.; Suzuki, Y.; Hayashi, K.; Komatsu, K.R.; Kawasaki, S.; Hidaka, K.; Yonehara, S.; Sugiyama, H.; et al. Protein-driven RNA nanostructured devices that function in vitro and control mammalian cell fate. Nat. Commun. 2017, 8, 540. [Google Scholar] [CrossRef]
- Li, H.; Lee, T.; Dziubla, T.; Pi, F.; Guo, S.; Xu, J.; Li, C.; Haque, F.; Liang, X.J.; Guo, P. RNA as a stable polymer to build controllable and defined nanostructures for material and biomedical applications. Nano Today 2015, 10, 631–655. [Google Scholar] [CrossRef]
- Xie, Z.; Liu, S.J.; Bleris, L.; Benenson, Y. Logic integration of mRNA signals by an RNAi-based molecular computer. Nucleic Acids Res. 2010, 38, 2692–2701. [Google Scholar] [CrossRef][Green Version]
- Xie, Z.; Wroblewska, L.; Prochazka, L.; Weiss, R.; Benenson, Y. Multi-input RNAi-based logic circuit for identification of specific cancer cells. Science 2011, 333, 1307. [Google Scholar] [CrossRef] [PubMed]
- Rinaudo, K.; Bleris, L.; Maddamsetti, R.; Subramanian, S.; Weiss, R.; Benenson, Y. A universal RNAi-based logic evaluator that operates in mammalian cells. Nat. Biotechnol. 2007, 25, 795. [Google Scholar] [CrossRef] [PubMed]
- Penchovsky, R.; Breaker, R.R. Computational design and experimental validation of oligonucleotide-sensing allosteric ribozymes. Nat. Biotechnol. 2005, 23, 1424–1433. [Google Scholar] [CrossRef]
- Soukup, G.A.; Breaker, R.R. Nucleic acid molecular switches. Trends Biotechnol. 1999, 17, 469–476. [Google Scholar] [CrossRef]
- Roark, B.K.; Tan, L.A.; Ivanina, A.; Chandler, M.; Castaneda, J.; Kim, H.S.; Jawahar, S.; Viard, M.; Talic, S.; Wustholz, K.L.; et al. Fluorescence blinking as an output signal for biosensing. ACS Sens. 2016, 1, 1295–1300. [Google Scholar] [CrossRef] [PubMed]
- Zakrevsky, P.; Parlea, L.; Viard, M.; Bindewald, E.; Afonin, K.A.; Shapiro, B.A. Preparation of a conditional RNA switch. Methods Mol. Biol. 2017, 1632, 303–324. [Google Scholar] [PubMed]
- Bindewald, E.; Afonin, K.A.; Viard, M.; Zakrevsky, P.; Kim, T.; Shapiro, B.A. Multistrand structure prediction of nucleic acid assemblies and design of RNA switches. Nano Lett. 2016, 16, 1726–1735. [Google Scholar] [CrossRef] [PubMed]
- Halman, J.R.; Satterwhite, E.; Roark, B.; Chandler, M.; Viard, M.; Ivanina, A.; Bindewald, E.; Kasprzak, W.K.; Panigaj, M.; Bui, M.N.; et al. Functionally-interdependent shape-switching nanoparticles with controllable properties. Nucleic Acids Res. 2017, 45, 2210–2220. [Google Scholar] [CrossRef]
- Chandler, M.; Ke, W.; Halman, J.; Panigaj, M.; Afonin, K.A. Reconfigurable nucleic acid materials for cancer therapy. In Nanooncology; Gonçalves, G., Tobias, G., Eds.; Springer: New York, NY, USA, 2018; pp. 365–385. [Google Scholar]
- Douglas, S.M.; Bachelet, I.; Church, G.M. A logic-gated nanorobot for targeted transport of molecular payloads. Science 2012, 335, 831–834. [Google Scholar] [CrossRef]
- Afonin, K.A.; Viard, M.; Kagiampakis, I.; Case, C.L.; Dobrovolskaia, M.A.; Hofmann, J.; Vrzak, A.; Kireeva, M.; Kasprzak, W.K.; KewalRamani, V.N.; et al. Triggering of RNA interference with RNA-RNA, RNA-DNA, and DNA-RNA nanoparticles. ACS Nano 2015, 9, 251–259. [Google Scholar] [CrossRef]
- Afonin, K.A.; Viard, M.; Martins, A.N.; Lockett, S.J.; Maciag, A.E.; Freed, E.O.; Heldman, E.; Jaeger, L.; Blumenthal, R.; Shapiro, B.A. Activation of different split functionalities upon re-association of RNA-DNA hybrids. Nat. Nanotechnol. 2013, 8, 296–304. [Google Scholar] [CrossRef]
- Sajja, S.; Chandler, M.; Fedorov, D.; Kasprzak, W.K.; Lushnikov, A.; Viard, M.; Shah, A.; Dang, D.; Dahl, J.; Worku, B.; et al. Dynamic behavior of RNA nanoparticles analyzed by AFM on a mica/air interface. Langmuir 2018. [Google Scholar] [CrossRef] [PubMed]
- Rackley, L.; Stewart, J.M.; Salotti, J.; Krokhotin, A.; Shah, A.; Halman, J.; Juneja, R.; Smollett, J.; Roark, B.; Viard, M.; et al. RNA fibers as optimized nanoscaffolds for siRNA coordination and reduced immunological recognition. Adv. Funct. Mater. 2018. [Google Scholar] [CrossRef]
- Ouellet, J. RNA fluorescence with light-up aptamers. Front. Chem. 2016, 4. [Google Scholar] [CrossRef] [PubMed]
- Marras, S.A.; Kramer, F.R.; Tyagi, S. Multiplex detection of single-nucleotide variations using molecular beacons. Genet. Anal. 1999, 14, 151–156. [Google Scholar] [CrossRef][Green Version]
- Feng, S.; Shang, Y.; Wu, F.; Ding, F.; Li, B.; Xu, J.; Xu, L.; Zhou, X. DNA nanomachines as evolved molecular beacons for in vitro and in vivo detection. Talanta 2014, 120, 141–147. [Google Scholar] [CrossRef] [PubMed]
- Bertrand, E.; Chartrand, P.; Schaefer, M.; Shenoy, S.M.; Singer, R.H.; Long, R.M. Localization of ASH1 mRNA particles in living yeast. Mol. Cell. 1998, 2, 437–445. [Google Scholar] [CrossRef]
- Valencia-Burton, M.; McCullough, R.M.; Cantor, C.R.; Broude, N.E. RNA visualization in live bacterial cells using fluorescent protein complementation. Nat. Methods 2007, 4, 421–427. [Google Scholar] [CrossRef]
- Demidov, V.V.; Dokholyan, N.V.; Witte-Hoffmann, C.; Chalasani, P.; Yiu, H.-W.; Ding, F.; Yu, Y.; Cantor, C.R.; Broude, N.E. Fast complementation of split fluorescent protein triggered by DNA hybridization. Proc. Nat. Acad. Sci. USA 2006, 103, 2052–2056. [Google Scholar] [CrossRef]
- Schwarz-Schilling, M.; Dupin, A.; Chizzolini, F.; Krishnan, S.; Mansy, S.S.; Simmel, F.C. Optimized assembly of a multifunctional RNA-protein nanostructure in a cell-free gene expression system. Nano Lett. 2018, 18, 2650–2657. [Google Scholar] [CrossRef]
- Grate, D.; Wilson, C. Laser-mediated, site-specific inactivation of RNA transcripts. Proc. Nat. Acad. Sci. USA 1999, 96, 6131–6136. [Google Scholar] [CrossRef] [PubMed][Green Version]
- Stojanovic, M.N.; Kolpashchikov, D.M. Modular aptameric sensors. J. Am. Chem. Soc. 2004, 126, 9266–9270. [Google Scholar] [CrossRef] [PubMed]
- Kolpashchikov, D.M. Binary malachite green aptamer for fluorescent detection of nucleic acids. J. Am. Chem. Soc. 2005, 127, 12442–12443. [Google Scholar] [CrossRef] [PubMed]
- Afonin, K.A.; Bindewald, E.; Yaghoubian, A.J.; Voss, N.; Jacovetty, E.; Shapiro, B.A.; Jaeger, L. In vitro assembly of cubic RNA-based scaffolds designed in silico. Nat. Nanotechnol. 2010, 5, 676–682. [Google Scholar] [CrossRef]
- Afonin, K.A.; Desai, R.; Viard, M.; Kireeva, M.L.; Bindewald, E.; Case, C.L.; Maciag, A.E.; Kasprzak, W.K.; Kim, T.; Sappe, A.; et al. Co-transcriptional production of RNA-DNA hybrids for simultaneous release of multiple split functionalities. Nucleic Acids Res. 2014, 42, 2085–2097. [Google Scholar] [CrossRef] [PubMed]
- Paige, J.S.; Wu, K.Y.; Jaffrey, S.R. RNA mimics of green fluorescent protein. Science 2012, 333, 642–646. [Google Scholar] [CrossRef] [PubMed]
- Paige, J.S.; Nguyen-Duc, T.; Song, W.; Jaffrey, S.R. Fluorescence imaging of cellular metabolites with RNA. Science 2012, 335, 1194. [Google Scholar] [CrossRef]
- Strack, R.L.; Disney, M.D.; Jaffrey, S.R. A superfolding Spinach2 reveals the dynamic nature of trinucleotide repeat–containing RNA. Nat. Methods 2013, 10, 1219. [Google Scholar] [CrossRef]
- Filonov, G.S.; Kam, C.W.; Song, W.; Jaffrey, S.R. In-gel imaging of RNA processing using broccoli reveals optimal aptamer expression strategies. Chem. Biol. 2015, 22, 649–660. [Google Scholar] [CrossRef]
- Filonov Grigory, S.; Jaffrey, S.R. RNA imaging with dimeric broccoli in live bacterial and mammalian cells. Curr. Protoc. Chem. Biol. 2016, 8, 1–28. [Google Scholar]
- Filonov, G.S.; Moon, J.D.; Svensen, N.; Jaffrey, S.R. Broccoli: Rapid selection of an RNA mimic of green fluorescent protein by fluorescence-based selection and directed evolution. J. Am. Chem. Soc. 2014, 136, 16299–16308. [Google Scholar] [CrossRef] [PubMed]
- Svensen, N.; Jaffrey, S.R. Fluorescent RNA aptamers as a tool to study RNA-modifying enzymes. Cell Chem. Biol. 2016, 23, 415–425. [Google Scholar] [PubMed]
- Autour, A.C.Y.; Jeng, S.D.; Cawte, A.; Abdolahzadeh, A.; Galli, A.; Panchapakesan, S.S.S.; Rueda, D.; Ryckelynck, M.; Unrau, P.J. Fluorogenic RNA mango aptamers for imaging small non-coding RNAs in mammalian cells. Nat. Commun. 2018, 9, 656. [Google Scholar] [CrossRef]
- Jepsen, M.D.E.; Sparvath, S.M.; Nielsen, T.B.; Langvad, A.H.; Grossi, G.; Gothelf, K.V.; Andersen, E.S. Development of a genetically encodable FRET system using fluorescent RNA aptamers. Nat. Commun. 2018, 9, 18. [Google Scholar] [CrossRef] [PubMed]
- Song, W.; Filonov, G.S.; Kim, H.; Hirsch, M.; Li, X.; Moon, J.D.; Jaffrey, S.R. Imaging RNA polymerase III transcription using a photostable RNA–fluorophore complex. Nat. Chem. Biol. 2017, 13, 1187. [Google Scholar] [CrossRef]
- Rogers, T.A.; Andrews, G.E.; Jaeger, L.; Grabow, W.W. Fluorescent monitoring of RNA assembly and processing using the split-spinach aptamer. ACS Synth. Biol. 2015, 4, 162–166. [Google Scholar] [CrossRef]
- Warner, K.D.; Chen, M.C.; Song, W.; Strack, R.L.; Thorn, A.; Jaffrey, S.R.; Ferre-D‘Amare, A.R. Structural basis for activity of highly efficient RNA mimics of green fluorescent protein. Nat. Struct. Mol. Biol. 2014, 21, 658–663. [Google Scholar] [CrossRef]
- Zadeh, J.N.; Steenberg, C.D.; Bois, J.S.; Wolfe, B.R.; Pierce, M.B.; Khan, A.R.; Dirks, R.M.; Pierce, N.A. NUPACK: Analysis and design of nucleic acid systems. J. Comput. Chem. 2011, 32, 170–173. [Google Scholar] [CrossRef]
- Afonin, K.A.; Kireeva, M.; Grabow, W.W.; Kashlev, M.; Jaeger, L.; Shapiro, B.A. Co-transcriptional assembly of chemically modified RNA nanoparticles functionalized with siRNAs. Nano Lett. 2012, 12, 5192–5195. [Google Scholar] [CrossRef]
- Britton, P.; Green, P.; Kottier, S.; Mawditt, K.L.; Penzes, Z.; Cavanagh, D.; Skinner, M.A. Expression of bacteriophage T7 RNA polymerase in avian and mammalian cells by a recombinant fowlpox virus. J. Gen. Virol. 1996, 77, 963–967. [Google Scholar] [CrossRef][Green Version]
- Elroy-Stein, O.; Moss, B. Cytoplasmic expression system based on constitutive synthesis of bacteriophage T7 RNA polymerase in mammalian cells. Proc. Nat. Acad. Sci. USA 1990, 87, 6743–6747. [Google Scholar] [CrossRef] [PubMed]
- Goldsworthy, V.; LaForce, G.; Abels, S.; Khisamutdinov, E. Fluorogenic RNA aptamers: A nano-platform for fabrication of simple and combinatorial logic gates. Nanomaterials 2018, 8, 984. [Google Scholar] [CrossRef] [PubMed]
- Lloyd, J.; Tran, C.H.; Wadhwani, K.; Cuba Samaniego, C.; Subramanian, H.K.K.; Franco, E. Dynamic control of aptamer-ligand activity using strand displacement reactions. ACS Synth. Biol. 2018, 7, 30–37. [Google Scholar] [CrossRef] [PubMed]
- Ke, W.; Hong, E.; Saito, R.F.; Rangel, M.C.; Wang, J.; Viard, M.; Richardson, M.; Khisamutdinov, E.F.; Panigaj, M.; Dokholyan, N.V.; et al. RNA-DNA fibers and polygons with controlled immunorecognition activate RNAi, FRET, and transcriptional regulation of NF-κB in human cells. Nucleic Acids Res. 2018. [Google Scholar] [CrossRef]
Sample Availability: Samples of the compounds are not available from the authors. |
© 2018 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
Share and Cite
Chandler, M.; Lyalina, T.; Halman, J.; Rackley, L.; Lee, L.; Dang, D.; Ke, W.; Sajja, S.; Woods, S.; Acharya, S.; et al. Broccoli Fluorets: Split Aptamers as a User-Friendly Fluorescent Toolkit for Dynamic RNA Nanotechnology. Molecules 2018, 23, 3178. https://doi.org/10.3390/molecules23123178
Chandler M, Lyalina T, Halman J, Rackley L, Lee L, Dang D, Ke W, Sajja S, Woods S, Acharya S, et al. Broccoli Fluorets: Split Aptamers as a User-Friendly Fluorescent Toolkit for Dynamic RNA Nanotechnology. Molecules. 2018; 23(12):3178. https://doi.org/10.3390/molecules23123178
Chicago/Turabian StyleChandler, Morgan, Tatiana Lyalina, Justin Halman, Lauren Rackley, Lauren Lee, Dylan Dang, Weina Ke, Sameer Sajja, Steven Woods, Shrija Acharya, and et al. 2018. "Broccoli Fluorets: Split Aptamers as a User-Friendly Fluorescent Toolkit for Dynamic RNA Nanotechnology" Molecules 23, no. 12: 3178. https://doi.org/10.3390/molecules23123178