Analysis of queuosine and 2-thio tRNA modifications by high throughput sequencing
| Autor: | Christopher D Katanski, Christopher P Watkins, Wen Zhang, Matthew Reyer, Samuel Miller, Tao Pan |
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| Rok vydání: | 2022 |
| Předmět: | |
| Zdroj: | Nucleic Acids Research. 50:e99-e99 |
| ISSN: | 1362-4962 0305-1048 |
| DOI: | 10.1093/nar/gkac517 |
| Popis: | Queuosine (Q) is a conserved tRNA modification at the wobble anticodon position of tRNAs that read the codons of amino acids Tyr, His, Asn, and Asp. Q-modification in tRNA plays important roles in the regulation of translation efficiency and fidelity. Queuosine tRNA modification is synthesized de novo in bacteria, whereas in mammals the substrate for Q-modification in tRNA is queuine, the catabolic product of the Q-base of gut bacteria. This gut microbiome dependent tRNA modification may play pivotal roles in translational regulation in different cellular contexts, but extensive studies of Q-modification biology are hindered by the lack of high throughput sequencing methods for its detection and quantitation. Here, we describe a periodate-treatment method that enables single base resolution profiling of Q-modification in tRNAs by Nextgen sequencing from biological RNA samples. Periodate oxidizes the Q-base, which results in specific deletion signatures in the RNA-seq data. Unexpectedly, we found that periodate-treatment also enables the detection of several 2-thio-modifications including τm5s2U, mcm5s2U, cmnm5s2U, and s2C by sequencing in human and E. coli tRNA. We term this method periodate-dependent analysis of queuosine and sulfur modification sequencing (PAQS-seq). We assess Q- and 2-thio-modifications at the tRNA isodecoder level, and 2-thio modification changes in stress response. PAQS-seq should be widely applicable in the biological studies of Q- and 2-thio-modifications in mammalian and microbial tRNAs. |
| Databáze: | OpenAIRE |
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