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| Status |
Public on Jun 05, 2021 |
| Title |
Deep and accurate detection of m6A RNA modifications in human and mouse cells using miCLIP2 and m6Aboost machine learning [a] |
| Organisms |
Homo sapiens; Mus musculus |
| Experiment type |
Expression profiling by high throughput sequencing
Other
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| Summary |
N6-methyladenosine (m6A) is the most abundant internal RNA modification in eukaryotic mRNAs and influences many aspects of RNA processing, such as RNA stability and translation. miCLIP (m6A individual-nucleotide resolution UV crosslinking and immunoprecipitation) is an antibody-based approach to map m6A sites in the transcriptome with single-nucleotide resolution. However, due to broad antibody reactivity, reliable identification of m6A sites from miCLIP data remains challenging. Here, we present several experimental and computational innovations that significantly improve transcriptome-wide detection of m6A sites. Based on the recently developed iCLIP2 protocol, the optimised miCLIP2 results in high-complexity libraries using less input material, leading to a more comprehensive representation of m6A sites. Next, we established a robust computational pipeline to identify true m6A sites from our miCLIP2 data. The analyses are calibrated with data from Mettl3 knockout cells to learn the characteristics of m6A deposition, including a significant number of m6A sites outside of DRACH motifs. In order to make these results universally applicable, we trained a machine learning model, m6Aboost, based on the experimental and RNA sequence features. Importantly, m6Aboost allows prediction of genuine m6A sites in miCLIP data without filtering for DRACH motifs or the need for Mettl3 depletion. Using m6Aboost, we identify thousands of high-confidence m6A sites in different murine and human cell lines, which provide a rich resource for future analysis. Collectively, our combined experimental and computational methodology greatly improves m6A identification.
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| Overall design |
miCLIP2 experiments were performed on polyA+ RNA from human and mouse cells, with 3 biological replicates per condition. Mettl3 KO cells served to calibrate the m6A detection.
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| Contributor(s) |
Körtel N, Rücklé C, Zhou Y, Busch A, Sutandy FR, Dominissine D, Dieterich C, Zarnack K, König J |
| Citation(s) |
34157120 |
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| Submission date |
Dec 18, 2020 |
| Last update date |
Sep 08, 2022 |
| Contact name |
Kathi Zarnack |
| E-mail(s) |
kathi.zarnack@bmls.de
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| Organization name |
Goethe University Frankfurt
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| Department |
Buchmann Institute for Molecular Life Sciences (BMLS)
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| Street address |
Max-von-Laue-Str. 15
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| City |
Frankfurt |
| ZIP/Postal code |
60438 |
| Country |
Germany |
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| Platforms (2) |
| GPL18573 |
Illumina NextSeq 500 (Homo sapiens) |
| GPL19057 |
Illumina NextSeq 500 (Mus musculus) |
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| Samples (22)
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| This SubSeries is part of SuperSeries: |
| GSE163500 |
Deep and accurate detection of m6A RNA modifications in human and mouse cells using miCLIP2 and m6Aboost machine learning |
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| Relations |
| BioProject |
PRJNA686380 |
| SRA |
SRP298464 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE163491_HEK293T_m6Aboost_predicted_m6A_sites.bed.gz |
549.1 Kb |
(ftp)(http) |
BED |
| GSE163491_Mouse_heart_100ng_m6Aboost_predicted_m6A_sites.bed.gz |
13.1 Kb |
(ftp)(http) |
BED |
| GSE163491_Mouse_heart_1ug_m6Aboost_predicted_m6A_sites.bed.gz |
489.2 Kb |
(ftp)(http) |
BED |
| GSE163491_Mouse_heart_300ng_m6Aboost_predicted_m6A_sites.bed.gz |
97.0 Kb |
(ftp)(http) |
BED |
| GSE163491_Mouse_heart_50ng_m6Aboost_predicted_m6A_sites.bed.gz |
3.1 Kb |
(ftp)(http) |
BED |
| GSE163491_RAW.tar |
600.0 Mb |
(http)(custom) |
TAR (of BW, TXT) |
| GSE163491_mESC_m6Aboost_predicted_m6A_sites.bed.gz |
424.8 Kb |
(ftp)(http) |
BED |
| GSE163491_miCLIP_mESC_Mettl3_KO_1.merged.v2uniqMD.duprm.C2T.minus.bw |
4.0 Mb |
(ftp)(http) |
BW |
| GSE163491_miCLIP_mESC_Mettl3_KO_1.merged.v2uniqMD.duprm.C2T.plus.bw |
4.1 Mb |
(ftp)(http) |
BW |
| GSE163491_miCLIP_mESC_Mettl3_KO_1.merged.v2uniqMD.duprm.noC2T.minus.bw |
38.6 Mb |
(ftp)(http) |
BW |
| GSE163491_miCLIP_mESC_Mettl3_KO_1.merged.v2uniqMD.duprm.noC2T.plus.bw |
39.3 Mb |
(ftp)(http) |
BW |
| GSE163491_miCLIP_mESC_WT_1.merged.v2uniqMD.duprm.C2T.minus.bw |
4.7 Mb |
(ftp)(http) |
BW |
| GSE163491_miCLIP_mESC_WT_1.merged.v2uniqMD.duprm.C2T.plus.bw |
4.8 Mb |
(ftp)(http) |
BW |
| GSE163491_miCLIP_mESC_WT_1.merged.v2uniqMD.duprm.noC2T.minus.bw |
48.5 Mb |
(ftp)(http) |
BW |
| GSE163491_miCLIP_mESC_WT_1.merged.v2uniqMD.duprm.noC2T.plus.bw |
49.6 Mb |
(ftp)(http) |
BW |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
| Processed data are available on Series record |
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