RNAs are involved in key cellular functions and are the only known bio-organic molecules within living cells that store genetic information and act as catalysts. Recently, new strategies have demonstrated that the number of non-coding RNAs (ncRNAs) in genomes is much greater than previously believed. While these strategies can identify ncRNA genes, such approaches are unable to report information on RNAs at the posttranscriptional level. Mass spectrometry (MS) has become an enabling technology for the fields of molecular and cell biology, and these developments have significantly enhanced our ability to study RNA. The redundancy of the genetic code results in 61 different anticodons available for transfer RNA (tRNA) decoding of messenger RNA. This redundancy leads to isoaccepting tRNAs, which are two or more tRNAs that carry the same amino acid but which have different tRNA sequences. Moreover, within an organism's genome, one or more copies may exist for a particular tRNA gene. In most organisms only a subset of these possible tRNAs, including isoacceptors, will be expressed, and the abundance of specific tRNAs may vary. For example, E. coli has 81 tRNA genes to express 53 different tRNAs which can decode 41 different codons. Because posttranscriptionally modified nucleosides have a significant influence on the decoding process, tRNA characterization is incomplete without an accurate characterization of the identities, sequence locations, and amounts of modified nucleosides. Because tRNAs are posttranscriptionally modified and these modifications have structural and functional significance, methods that enable the sensitive and accurate determination of tRNA population and extent of tRNA modification are required. This talk describes several of the recent advances in RNA modification analysis made in our lab, with an emphasis on methods suitable for higher-throughput characterization of transfer and ribosomal RNAs.

New mass spectrometry assays for identifying and characterizing modified RNAs

247th National Meeting (2014)

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