Mechanisms of molecular self-replication have the potential to shed light on the origins of life. In particular, self-replication through RNA-catalysed templated RNA synthesis is thought to have supported a primordial ‘RNA world’. However, existing polymerase ribozymes lack the capacity to synthesize RNAs approaching their own size. Here, we report the in vitro evolution of such catalysts directly in the RNA-stabilizing medium of water ice, which yielded RNA polymerase ribozymes specifically adapted to sub-zero temperatures and able to synthesize RNA in ices at temperatures as low as −19 °C. The combination of cold-adaptive mutations with a previously described 5′ extension operating at ambient temperatures enabled the design of a first polymerase ribozyme capable of catalysing the accurate synthesis of an RNA sequence longer than itself (adding up to 206 nucleotides), an important stepping stone towards RNA self-replication.
This is a preview of subscription content, access via your institution
Open Access articles citing this article.
Nature Communications Open Access 03 June 2022
BMC Evolutionary Biology Open Access 03 April 2019
Nature Communications Open Access 13 June 2018
Subscribe to Journal
Get full journal access for 1 year
only $9.92 per issue
All prices are NET prices.
VAT will be added later in the checkout.
Tax calculation will be finalised during checkout.
Get time limited or full article access on ReadCube.
All prices are NET prices.
Atkins, J. F., Gesteland, R. F. & Cech, T. R. (eds) RNA Worlds (Cold Spring Harbor Laboratory, 2011).
Gilbert, W. Origin of life: the RNA world. Nature 319, 618 (1986).
Powner, M. W., Gerland, B. & Sutherland, J. D. Synthesis of activated pyrimidine ribonucleotides in prebiotically plausible conditions. Nature 459, 239–242 (2009).
Bowler, F. R. et al. Prebiotically plausible oligoribonucleotide ligation facilitated by chemoselective acetylation. Nature Chem. 5, 383–389 (2013).
Engelhart, A. E., Powner, M. W. & Szostak, J. W. Functional RNAs exhibit tolerance for non-heritable 2′–5′ versus 3′–5′ backbone heterogeneity. Nature Chem. 5, 390–394 (2013).
Szostak, J. W., Bartel, D. P. & Luisi, P. L. Synthesizing life. Nature 409, 387–390 (2001).
Robertson, M. P. & Joyce, G. F. The origins of the RNA world. Cold Spring Harb. Perspect. Biol. http://dx.doi.org/10.1101/cshperspect.a003608 (2010).
Ellington, A. D., Chen, X., Robertson, M. & Syrett, A. Evolutionary origins and directed evolution of RNA. Int. J. Biochem. Cell Biol. 41, 254–265 (2009).
Johnston, W. K., Unrau, P. J., Lawrence, M. S., Glasner, M. E. & Bartel, D. P. RNA-catalysed RNA polymerization: accurate and general RNA-templated primer extension. Science 292, 1319–1325 (2001).
Zaher, H. S. & Unrau, P. J. Selection of an improved RNA polymerase ribozyme with superior extension and fidelity. RNA 13, 1017–1026 (2007).
Wochner, A., Attwater, J., Coulson, A. & Holliger, P. Ribozyme-catalysed transcription of an active ribozyme. Science 332, 209–212 (2011).
Bartel, D. P. & Szostak, J. W. Isolation of new ribozymes from a large pool of random sequences. Science 261, 1411–1418 (1993).
Ekland, E. H., Szostak, J. W. & Bartel, D. P. Structurally complex and highly active RNA ligases derived from random RNA sequences. Science 269, 364–370 (1995).
Lawrence, M. S. & Bartel, D. P. Processivity of ribozyme-catalysed RNA polymerization. Biochemistry 42, 8748–8755 (2003).
Attwater, J., Wochner, A., Pinheiro, V. B., Coulson, A. & Holliger, P. Ice as a protocellular medium for RNA replication. Nature Commun. 1, 76 (2010).
Attwater, J. et al. Chemical fidelity of an RNA polymerase ribozyme. Chem. Sci. 4, 2804–2814 (2013).
Dobson, C. M., Ellison, G. B., Tuck, A. F. & Vaida, V. Atmospheric aerosols as prebiotic chemical reactors. Proc. Natl Acad. Sci. USA 97, 11864–11868 (2000).
Chen, I. A., Salehi-Ashtiani, K. & Szostak, J. W. RNA catalysis in model protocell vesicles. J. Am. Chem. Soc. 127, 13213–13219 (2005).
Budin, I. & Szostak, J. W. Expanding roles for diverse physical phenomena during the origin of life. Annu. Rev. Biophys. 39, 245–263 (2010).
Monnard, P. A. Catalysis in abiotic structured media: an approach to selective synthesis of biopolymers. Cell. Mol. Life Sci. 62, 520–534 (2005).
Vlassov, A. V., Kazakov, S. A., Johnston, B. H. & Landweber, L. F. The RNA world on ice: a new scenario for the emergence of RNA information. J. Mol. Evol. 61, 264–273 (2005).
Muller, U. F. & Bartel, D. P. Improved polymerase ribozyme efficiency on hydrophobic assemblies. RNA 14, 552–562 (2008).
Kun, A., Santos, M. & Szathmary, E. Real ribozymes suggest a relaxed error threshold. Nature Genet. 37, 1008–1011 (2005).
Rajamani, S. et al. Effect of stalling after mismatches on the error catastrophe in nonenzymatic nucleic acid replication. J. Am. Chem. Soc. 132, 5880–5885 (2010).
Lawrence, M. S. & Bartel, D. P. New ligase-derived RNA polymerase ribozymes. RNA 11, 1173–1180 (2005).
Zuker, M. Mfold web server for nucleic acid folding and hybridization prediction. Nucleic Acids Res. 31, 3406–3415 (2003).
Strulson, C. A., Molden, R. C., Keating, C. D. & Bevilacqua, P. C. RNA catalysis through compartmentalization. Nature Chem. 4, 941–946 (2012).
Vlassov, A. V., Johnston, B. H., Landweber, L. F. & Kazakov, S. A. Ligation activity of fragmented ribozymes in frozen solution: implications for the RNA world. Nucleic Acids Res. 32, 2966–2974 (2004).
Shechner, D. M. et al. Crystal structure of the catalytic core of an RNA-polymerase ribozyme. Science 326, 1271–1275 (2009).
Shechner, D. M. & Bartel, D. P. The structural basis of RNA-catalysed RNA polymerization. Nature Struct. Mol. Biol. 18, 1036–1042 (2011).
Muller, U. F. & Bartel, D. P. Substrate 2′-hydroxyl groups required for ribozyme-catalysed polymerization. Chem. Biol. 10, 799–806 (2003).
Joyce, G. F. & Orgel, L. E. Non-enzymatic template-directed synthesis on RNA random copolymers. Poly(C,A) templates. J. Mol. Biol. 202, 677–681 (1988).
The authors thank J.N. Skepper (University of Cambridge) for help with SEM imaging, S. James and S. Brunner for help with MiSeq sequencing and analysis and M. Daly (MRC LMB) for help with FACS. This work was supported by a Homerton College, Cambridge Junior Research Fellowship (J.A.) and by the Medical Research Council (programme number U105178804).
The authors declare no competing financial interests.
About this article
Cite this article
Attwater, J., Wochner, A. & Holliger, P. In-ice evolution of RNA polymerase ribozyme activity. Nature Chem 5, 1011–1018 (2013). https://doi.org/10.1038/nchem.1781
This article is cited by
Nature Communications (2022)
The identification and characterization of a selected SAM-dependent methyltransferase ribozyme that is present in natural sequences
Nature Catalysis (2021)
Nature Reviews Earth & Environment (2021)
BMC Evolutionary Biology (2019)
Nature Communications (2018)