Title | Characterizing Expression and Processing of Precursor and Mature Human tRNAs by Hydro-tRNAseq and PAR-CLIP. |
Publication Type | Journal Article |
Year of Publication | 2017 |
Authors | Gogakos T, Brown M, Garzia A, Meyer C, Hafner M, Tuschl T |
Journal | Cell Rep |
Volume | 20 |
Issue | 6 |
Pagination | 1463-1475 |
Date Published | 2017 Aug 08 |
ISSN | 2211-1247 |
Keywords | HEK293 Cells, Humans, RNA Processing, Post-Transcriptional, RNA, Transfer, Sequence Analysis, RNA |
Abstract | The participation of tRNAs in fundamental aspects of biology and disease necessitates an accurate, experimentally confirmed annotation of tRNA genes and curation of tRNA sequences. This has been challenging because RNA secondary structure, nucleotide modifications, and tRNA gene multiplicity complicate sequencing and mapping efforts. To address these issues, we developed hydro-tRNAseq, a method based on partial alkaline RNA hydrolysis that generates fragments amenable for sequencing. To identify transcribed tRNA genes, we further complemented this approach with photoactivatable crosslinking and immunoprecipitation (PAR-CLIP) of SSB/La, a conserved protein involved in pre-tRNA processing. Our results show that approximately half of all predicted tRNA genes are transcribed in human cells. We also report nucleotide modification sites and their order of introduction, and we identify tRNA leaders, trailers, and introns. By using complementary sequencing-based methodologies, we present a human tRNA atlas and determine expression levels of mature and processing intermediates of tRNAs in human cells. |
DOI | 10.1016/j.celrep.2017.07.029 |
Alternate Journal | Cell Rep |
PubMed ID | 28793268 |
PubMed Central ID | PMC5564215 |
Grant List | R01 CA159227 / CA / NCI NIH HHS / United States U19 CA179564 / CA / NCI NIH HHS / United States / / Howard Hughes Medical Institute / United States UH2 AR067689 / AR / NIAMS NIH HHS / United States UH3 TR000933 / TR / NCATS NIH HHS / United States UH2 TR000933 / TR / NCATS NIH HHS / United States |
Submitted by kej2006 on June 6, 2018 - 4:10pm