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Chemical Method to Sequence 5-Formylcytosine on RNA.

ABSTRACT: Epitranscriptomic RNA modifications can regulate biological processes, but there remains a major gap in our ability to identify and measure individual modifications at nucleotide resolution. Here we present Mal-Seq, a chemical method for sequencing 5-formylcytosine (f⁵C) modifications on RNA based on the selective and efficient malononitrile-mediated labeling of f⁵C residues to generate adducts that are read as C-to-T mutations upon reverse transcription and polymerase chain reaction amplification. We apply Mal-Seq to characterize the prevalence of f⁵C at the wobble position of mt-tRNA(Met) in different organisms and tissue types and find that high-level f⁵C modification is present in mammals but lacking in lower eukaryotes. Our work sheds light on mitochondrial tRNA modifications throughout eukaryotic evolution and provides a general platform for characterizing the f⁵C epitranscriptome.

SUBMITTER: Li A

PROVIDER: S-EPMC9357364 | biostudies-literature | 2022 Mar

REPOSITORIES: biostudies-literature

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Chemical Method to Sequence 5-Formylcytosine on RNA.

Li Ang A Sun Xuemeng X Arguello A Emilia AE Kleiner Ralph E RE

ACS chemical biology 20220225 3

Epitranscriptomic RNA modifications can regulate biological processes, but there remains a major gap in our ability to identify and measure individual modifications at nucleotide resolution. Here we present Mal-Seq, a chemical method for sequencing 5-formylcytosine (f<sup>5</sup>C) modifications on RNA based on the selective and efficient malononitrile-mediated labeling of f<sup>5</sup>C residues to generate adducts that are read as C-to-T mutations upon reverse transcription and polymerase chai ...[more]

PMID: 35212224

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Project description:MotivationsBiclustering is a clustering method that simultaneously clusters both the domain and range of a relation. A challenge in multiple sequence alignment (MSA) is that the alignment of sequences is often intended to reveal groups of conserved functional subsequences. Simultaneously, the grouping of the sequences can impact the alignment; precisely the kind of dual situation biclustering is intended to address.ResultsWe define a representation of the MSA problem enabling the application of biclustering algorithms. We develop a computer program for local MSA, BlockMSA, that combines biclustering with divide-and-conquer. BlockMSA simultaneously finds groups of similar sequences and locally aligns subsequences within them. Further alignment is accomplished by dividing both the set of sequences and their contents. The net result is both a multiple sequence alignment and a hierarchical clustering of the sequences. BlockMSA was tested on the subsets of the BRAliBase 2.1 benchmark suite that display high variability and on an extension to that suite to larger problem sizes. Also, alignments were evaluated of two large datasets of current biological interest, T box sequences and Group IC1 Introns. The results were compared with alignments computed by ClustalW, MAFFT, MUCLE and PROBCONS alignment programs using Sum of Pairs (SPS) and Consensus Count. Results for the benchmark suite are sensitive to problem size. On problems of 15 or greater sequences, BlockMSA is consistently the best. On none of the problems in the test suite are there appreciable differences in scores among BlockMSA, MAFFT and PROBCONS. On the T box sequences, BlockMSA does the most faithful job of reproducing known annotations. MAFFT and PROBCONS do not. On the Intron sequences, BlockMSA, MAFFT and MUSCLE are comparable at identifying conserved regions.AvailabilityBlockMSA is implemented in Java. Source code and supplementary datasets are available at http://aug.csres.utexas.edu/msa/

Dataset Information

Chemical Method to Sequence 5-Formylcytosine on RNA.

Publications

Chemical Method to Sequence 5-Formylcytosine on RNA.

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