Poster abstracts

Poster number 39 submitted by Saurja DasGupta

RNA assembly in an RNA World

Saurja DasGupta (Dept. of Molecular Biology, Massachusetts General Hospital), Aleksander Radakovic (Biological and Biomedical Science Graduate Program, Harvard Medical School), Travis S. Walton (Dept. of Biological Chemistry and Molecular Pharmacology, Harvard Medical School), Stephanie J. Zhang (Dept. of Pathology, Boston Childrens Hospital), Jack W. Szostak (Dept. of Chemistry, The University of Chicago)

Abstract:
RNA is central to the emergence of early life due to its ability to function as genes as well as enzymes (ribozymes).[1] Therefore, RNA assembly processes were crucial for the replication of primordial genomes and for generating the enzymes required for primordial life. The first ribozymes must have been generated via the nonenzymatic assembly of activated RNA building blocks like phosphorimidazolides, and the emergence of ribozyme catalysis would have significantly accelerated RNA assembly. We have experimentally reconstructed prebiotically plausible pathways that would facilitate this crucial transition between nonenzymatic and enzymatic RNA assembly. First, we used in vitro selection to identify ribozymes that catalyze RNA ligation using the phosphorimidazolide substrates of nonenzymatic RNA ligation[2] and showed that substrate activation and RNA-catalyzed RNA ligation can occur in the same microenvironment. Next, we demonstrated that functional chimeric ribozymes, like the aforementioned phosphorimidazolide ligase, may be generated by the enzyme-free self-assembly of 3′ aminoacylated RNA oligomers.[3] These results provide an exciting scenario for robust prebiotic RNA assembly.
But how do we move to more biological RNA assembly? Using directed evolution, we optimized the phosphorimidazolide ligase to use more biologically relevant triphosphate substrates for RNA ligation. Interestingly, the new ribozyme was able to utilize both phosphorimidazolide and triphosphate substrates, making it an excellent evolutionary intermediate. We delineated a quasi-neutral mutational pathway that links the parent phosphorimidazolide ligase and the new triphosphate ligase sequences. This suggests that the transition from RNA assembly with prebiotic substrates to that with more biological substrates may be possible by simple neutral mutational drifts across the RNA sequence space, which underscores the ease of functional diversification in enzymes of the RNA World.

References:
[1] W. Gilbert, Origin of life: The RNA world. Nature, 319, 618 (1986).
[2] T. Walton, S. DasGupta, D. Duzdevich, S. S. Oh, J. W. Szostak, In vitro
selection of ribozyme ligases that use prebiotically plausible 2-
aminoimidazole-activated substrates. Proc. Natl. Acad. Sci. U.S.A. 117,
5741–5748 (2020).
[3] A. Radakovic, S. DasGupta, T. H. Wright, Harry Aitken, Jack W. Szostak,
Nonenzymatic assembly of active chimeric ribozymes from aminoacylated RNA
oligonucleotides. Proc. Natl. Acad. Sci. U.S.A. 119, e2116840119 (2022).

Keywords: RNA World, Ribozyme, Evolution