Discovery and biological characterization of geranylated RNA in bacteria

Abstract

A general MS-based screen for unusually hydrophobic cellular small molecule–RNA conjugates revealed geranylated RNA in Escherichia coli, Enterobacter aerogenes, Pseudomonas aeruginosa and Salmonella enterica var. Typhimurium. The geranyl group is conjugated to the sulfur atom in two 5-methylaminomethyl-2-thiouridine nucleotides. These geranylated nucleotides occur in the first anticodon position of tRNAGluUUC, tRNALysUUU and tRNAGlnUUG at a frequency of up to 6.7% (400 geranylated nucleotides per cell). RNA geranylation can be increased or abolished by mutation or deletion of the selU (ybbB ) gene in E. coli, and purified SelU protein in the presence of geranyl pyrophosphate and tRNA can produce geranylated tRNA. The presence or absence of the geranyl group in tRNAGluUUC, tRNALysUUU and tRNAGlnUUG affects codon bias and frameshifting during translation. These RNAs represent the first reported examples of oligoisoprenylated cellular nucleic acids.

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Figure 1: Discovery of two hydrophobic small molecule–RNA conjugates with [M-H] m/z = 824.200 and 868.189.
Figure 2: MS characterization of two new hydrophobic small molecule–RNA conjugates.
Figure 3: Structural elucidation of two geranylated nucleosides.
Figure 4: Characterization of geranylated cellular RNAs.
Figure 5: Biological abundance and properties of geranylated RNA.

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Acknowledgements

This work was supported by the Howard Hughes Medical Institute and the US National Institutes of Health (NIH)-National Institute of General Medical Sciences (NIGMS) (R01GM065865). C.E.D. acknowledges fellowship from the Novartis Foundation. A.M.L. was supported by a NIH National Research Service Award Postdoctoral Fellowship (F32GM095028). We thank S.-L. Zheng for his help with X-ray diffraction and structural determination of synthetic geranyl-2-thiouracil. We thank A. Saghatelian (Harvard University) for providing P. aeruginosa and I. Chen (Harvard University) for providing S. Typhimurium and BL2 facilities. We are also grateful to J. Carlson for his assistance and M. Ibba for helpful discussions.

Author information

C.E.D., Y.C., A.M.L. and Y.G.C. designed and performed the experiments. All authors analyzed the results and wrote the manuscript.

Correspondence to David R Liu.

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Supplementary information

Supplementary Text and Figures

Supplementary Methods and Supplementary Results (PDF 3375 kb)

Supplementary Data Set 1

geranyl-2-thiouracil (CIF 14 kb)

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Dumelin, C., Chen, Y., Leconte, A. et al. Discovery and biological characterization of geranylated RNA in bacteria. Nat Chem Biol 8, 913–919 (2012). https://doi.org/10.1038/nchembio.1070

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