Detection of RNA modifications

Kellner, Stefanie; Burhenne, Jürgen; Helm, Mark

doi:10.4161/rna.7.2.11468

Cited by 115 publications

(96 citation statements)

References 102 publications

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“…tRNAs undergo extensive posttranscriptional base modifications in all organisms and genetic compartments that have been investigated (Sprinzl et al, 1998;Helm, 2006;Kellner et al, 2010;Phizicky and Alfonzo, 2010). These modifications are introduced at the posttranscriptional level by specific enzymes.…”

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confidence: 99%

Identification of Enzymes for Adenosine-to-Inosine Editing and Discovery of Cytidine-to-Uridine Editing in Nucleus-Encoded Transfer RNAs of Arabidopsis

2014

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In all organisms, transfer RNAs (tRNAs) contain numerous modified nucleotides. For many base modifications in tRNAs, the functional significance is not well understood, and the enzymes performing the modification reactions are unknown. Here, we have studied members of a family of putative nucleotide deaminases in the model plant Arabidopsis (Arabidopsis thaliana). We show that two Arabidopsis genes encoding homologs of yeast (Saccharomyces cerevisiae) tRNA adenosine deaminases catalyze adenosine-to-inosine editing in position 34 of several cytosolic tRNA species. The encoded proteins (AtTAD2 and AtTAD3, for tRNA-specific adenosine deaminase) localize to the nucleus and interact with each other in planta in bimolecular fluorescence complementation and coimmunoprecipitation assays. Both AtTAD2 and AtTAD3 are encoded by essential genes whose knockout is lethal and leads to arrested embryo development at the globular stage. Knockdown mutants for AtTAD2 and AtTAD3 display reduced growth and inefficient editing from adenosine to inosine in six nucleus-encoded tRNA species. Moreover, upon comparison of DNA and complementary DNA sequences, we discovered cytidine-to-uridine RNA editing in position 32 of two nucleus-encoded serine tRNAs, tRNA-serine(AGA) and tRNA-serine(GCT). This adds a unique type of RNA editing to the modifications occurring in nuclear genome-encoded RNAs in plants.

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Identification of Enzymes for Adenosine-to-Inosine Editing and Discovery of Cytidine-to-Uridine Editing in Nucleus-Encoded Transfer RNAs of Arabidopsis

2014

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“…Over 100 post-transcriptional modifications are present in tRNAs, many of which play crucial roles in tRNA folding and function such as codon recognition. [16][17][18] Because many such modifications inhibit Watson-Crick base paring and thus arrest reverse-transcription, 19 standard qRT-PCR would produce severely biased results with underrepresentation of heavily-modified tRNAs. This modification issue is inevitable with any PCR-based technology used for the detection and quantification of RNA because there is no experimental methodology that removes all tRNA modifications.…”

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confidence: 99%

Four-leaf clover qRT-PCR: A convenient method for selective quantification of mature tRNA

et al. 2015

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Transfer RNAs (tRNAs) play a central role in translation and also recently appear to have a variety of other functions in biological processes beyond translation. Here we report the development of Four-Leaf clover qRT-PCR (FL-PCR), a convenient PCR-based method, which can specifically quantify individual mature tRNA species. In FL-PCR, T4 RNA ligase 2 specifically ligates a stem-loop adapter to mature tRNAs but not to precursor tRNAs or tRNA fragments. Subsequent TaqMan qRT-PCR amplifies only unmodified regions of the tRNA-adapter ligation products; therefore, FL-PCR quantification is not influenced by tRNA post-transcriptional modifications. FL-PCR has broad applicability for the quantification of various tRNAs in different cell types, and thus provides a much-needed simple method for analyzing tRNA abundance and heterogeneity.

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“…tRNA HisGUG contains a guanine residue at np 37, which is often modified to 1-methyl-guanosine (m 1 G). The m 1 G modification inhibits reverse-transcription (22), which would result in the unsuccessful amplification of the 3′-tRNA HisGUG half by RACE. Interestingly, reduced levels of the 5′-tRNA AspGUC expression were observed in BmN4 cells whose cell cycle was arrested by double thymidine block (Fig.…”

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Sex hormone-dependent tRNA halves enhance cell proliferation in breast and prostate cancers

Honda

Loher

Shigematsu

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

295

464

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Sex hormones and their receptors play critical roles in the development and progression of the breast and prostate cancers. Here we report that a novel type of transfer RNA (tRNA)-derived small RNA, termed Sex HOrmone-dependent TRNA-derived RNAs (SHOTRNAs), are specifically and abundantly expressed in estrogen receptor (ER)-positive breast cancer and androgen receptor (AR)-positive prostate cancer cell lines. SHOT-RNAs are not abundantly present in ER − breast cancer, AR − prostate cancer, or other examined cancer cell lines from other tissues. ER-dependent accumulation of SHOT-RNAs is not limited to a cell culture system, but it also occurs in luminal-type breast cancer patient tissues. SHOT-RNAs are produced from aminoacylated mature tRNAs by angiogenin-mediated anticodon cleavage, which is promoted by sex hormones and their receptors. Resultant 5′-and 3′-SHOT-RNAs, corresponding to 5′-and 3′-tRNA halves, bear a cyclic phosphate (cP) and an amino acid at the 3′-end, respectively. By devising a "cP-RNA-seq" method that is able to exclusively amplify and sequence cP-containing RNAs, we identified the complete repertoire of 5′-SHOT-RNAs. Furthermore, 5′-SHOT-RNA, but not 3′-SHOT-RNA, has significant functional involvement in cell proliferation. These results have unveiled a novel tRNA-engaged pathway in tumorigenesis of hormone-dependent cancers and implicate SHOT-RNAs as potential candidates for biomarkers and therapeutic targets.tRNA | tRNA half | sex hormone | breast cancer | prostate cancer

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Detection of RNA modifications

Cited by 115 publications

References 102 publications

Identification of Enzymes for Adenosine-to-Inosine Editing and Discovery of Cytidine-to-Uridine Editing in Nucleus-Encoded Transfer RNAs of Arabidopsis

Identification of Enzymes for Adenosine-to-Inosine Editing and Discovery of Cytidine-to-Uridine Editing in Nucleus-Encoded Transfer RNAs of Arabidopsis

Four-leaf clover qRT-PCR: A convenient method for selective quantification of mature tRNA

Sex hormone-dependent tRNA halves enhance cell proliferation in breast and prostate cancers

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