Nucleic Acids Res. 2017 Feb 28;45(4):2124-2136. doi: 10.1093/nar/gkw1120.

Identification of 2-methylthio cyclic N6-threonylcarbamoyladenosine (ms2ct6A) as a novel RNA modification at position 37 of tRNAs.

Kang BI1, Miyauchi K1, Matuszewski M2, D'Almeida GS3, Rubio MAT3, Alfonzo JD3, Inoue K1, Sakaguchi Y1, Suzuki T1, Sochacka E2, Suzuki T1.

Author information

1

Department of Chemistry and Biotechnology, Graduate School of Engineering, University of Tokyo, Tokyo 113-8656, Japan.

2

Institute of Organic Chemistry, Faculty of Chemistry, Lodz University of Technology, Lodz 90-924, Poland.

3

Department of Microbiology and The Center for RNA Biology, The Ohio State University, Columbus, OH 43210, USA.

Abstract

Transfer RNA modifications play pivotal roles in protein synthesis. N6-threonylcarbamoyladenosine (t6A) and its derivatives are modifications found at position 37, 3΄-adjacent to the anticodon, in tRNAs responsible for ANN codons. These modifications are universally conserved in all domains of life. t6A and its derivatives have pleiotropic functions in protein synthesis including aminoacylation, decoding and translocation. We previously discovered a cyclic form of t6A (ct6A) as a chemically labile derivative of t6A in tRNAs from bacteria, fungi, plants and protists. Here, we report 2-methylthio cyclic t6A (ms2ct6A), a novel derivative of ct6A found in tRNAs from Bacillus subtilis, plants and Trypanosoma brucei. In B. subtilis and T. brucei, ms2ct6A disappeared and remained to be ms2t6A and ct6A by depletion of tcdA and mtaB homologs, respectively, demonstrating that TcdA and MtaB are responsible for biogenesis of ms2ct6A.

© The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

PMID:

 

27913733

 

PMCID:

 

PMC5389704

 

DOI:

 

10.1093/nar/gkw1120

Nature. 2017 Feb 22;542(7642):494-497. doi: 10.1038/nature21396.

Editing and methylation at a single site by functionally interdependent activities.

Rubio MA1, Gaston KW1,2, McKenney KM1, Fleming IM1, Paris Z1,3, Limbach PA2, Alfonzo JD1.

Author information

1

Department of Microbiology, Ohio State Biochemistry Program and The Center for RNA Biology, The Ohio State University, Columbus, Ohio 43210, USA.

2

Rieveschl Laboratories for Mass Spectrometry, Department of Chemistry, University of Cincinnati, Cincinnati, Ohio 45221, USA.

3

Institute of Parasitology, Biology Centre and Faculty of Sciences, University of South Bohemia, 37005 České Budějovice (Budweis), Czech Republic.

Abstract

Nucleic acids undergo naturally occurring chemical modifications. Over 100 different modifications have been described and every position in the purine and pyrimidine bases can be modified; often the sugar is also modified. Despite recent progress, the mechanism for the biosynthesis of most modifications is not fully understood, owing, in part, to the difficulty associated with reconstituting enzyme activity in vitro. Whereas some modifications can be efficiently formed with purified components, others may require more intricate pathways. A model for modification interdependence, in which one modification is a prerequisite for another, potentially explains a major hindrance in reconstituting enzymatic activity in vitro. This model was prompted by the earlier discovery of tRNA cytosine-to-uridine editing in eukaryotes, a reaction that has not been recapitulated in vitro and the mechanism of which remains unknown. Here we show that cytosine 32 in the anticodon loop of Trypanosoma brucei tRNAThr is methylated to 3-methylcytosine (m3C) as a pre-requisite for C-to-U deamination. Formation of m3C in vitro requires the presence of both the T. brucei m3C methyltransferase TRM140 and the deaminase ADAT2/3. Once formed, m3C is deaminated to 3-methyluridine (m3U) by the same set of enzymes. ADAT2/3 is a highly mutagenic enzyme, but we also show that when co-expressed with the methyltransferase its mutagenicity is kept in check. This helps to explain how T. brucei escapes 'wholesale deamination' of its genome while harbouring both enzymes in the nucleus. This observation has implications for the control of another mutagenic deaminase, human AID, and provides a rationale for its regulation.

Comment in

• Molecular biology: RNA editing packs a one-two punch. [Nature. 2017]

PMID:

 

28230119

 

DOI:

 

10.1038/nature21396