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| Status |
Public on Sep 25, 2023 |
| Title |
Systematic identification of RNA-binding proteins and tethered domains that activate exon splicing inclusion [eCLIP-seq] |
| Organism |
Homo sapiens |
| Experiment type |
Other
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| Summary |
RNA-binding proteins (RBPs) modulate alternative splicing outcomes to determine isoform expression and cellular survival. To identify RBPs that directly drive alternative exon inclusion, we evaluated 718 human RBPs with tethered function luciferase-based splicing reporter assays to identify 58 candidates, including known splicing factors such as RBFOX and serine-arginine proteins. We performed enhanced CLIP, RNA-seq, and affinity purification-mass spectrometry to investigate a subset of the 11 candidates with no prior association with splicing. Integrative analysis of these assays indicated the surprising roles of TRNAU1AP, SCAF8, and RTCA in modulating hundreds of endogenous splicing events. We also leveraged our tethering assays and top candidates to identify potent and compact exon inclusion activation domains for splicing modulation applications. Using identified domains, we engineered programmable fusion proteins which outperformed current artificial splicing factors at manipulating inclusion of reporter and endogenous exons. Altogether, our tethering approach characterized the ability of RBPs to induce exon inclusion and yielded new molecular parts for programmable splicing control.
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| Overall design |
eCLIP-seq with TRNAU1AP-specific antibody in HEK cells, or V5-specific antibody in HEK cells overexpressing V5-tagged targets
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| Contributor(s) |
Schmok JC, Jain M, Street LA, Gosztyla ML, Boyle EA, Jagannatha P, Luo E, Jovanovic M, Yeo GW |
| Citation missing |
Has this study been published? Please login to update or notify GEO. |
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| Submission date |
May 16, 2023 |
| Last update date |
Sep 25, 2023 |
| Contact name |
Brian Yee |
| E-mail(s) |
brian.alan.yee@gmail.com
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| Organization name |
UCSD
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| Department |
Health
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| Lab |
Yeo
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| Street address |
9500 Gilman Dr
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| City |
La Jolla |
| State/province |
CA |
| ZIP/Postal code |
92093 |
| Country |
USA |
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| Platforms (1) |
| GPL24676 |
Illumina NovaSeq 6000 (Homo sapiens) |
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| Samples (20)
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| GSM7359776 |
HEK293T cells, TRNAU1AP_IP_2 |
| GSM7359777 |
HEK293T cells, RTCA_IN_1 |
| GSM7359778 |
HEK293T cells, RTCA_IN_2 |
| GSM7359779 |
HEK293T cells, RTCA_IP_1 |
| GSM7359780 |
HEK293T cells, RTCA_IP_2 |
| GSM7359781 |
HEK293T cells, SCAF8_IN_1 |
| GSM7359782 |
HEK293T cells, SCAF8_IN_2 |
| GSM7359783 |
HEK293T cells, SCAF8_IP_1 |
| GSM7359784 |
HEK293T cells, SCAF8_IP_2 |
| GSM7359785 |
HEK293T cells, STAU2_IN_1 |
| GSM7359786 |
HEK293T cells, STAU2_IN_2 |
| GSM7359787 |
HEK293T cells, STAU2_IP_1 |
| GSM7359788 |
HEK293T cells, STAU2_IP_2 |
| GSM7359793 |
HEK293T cells, FLAG_IN_1 |
| GSM7359794 |
HEK293T cells, FLAG_IN_2 |
| GSM7359795 |
HEK293T cells, FLAG_IP_1 |
| GSM7359796 |
HEK293T cells, FLAG_IP_2 |
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| This SubSeries is part of SuperSeries: |
| GSE232599 |
Systematic identification of RNA-binding proteins and tethered domains that activate exon splicing inclusion |
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| Relations |
| BioProject |
PRJNA972979 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE232597_FLAG_IP1_enriched_windows.tsv.gz |
191.0 Kb |
(ftp)(http) |
TSV |
| GSE232597_FLAG_IP2_enriched_windows.tsv.gz |
529.6 Kb |
(ftp)(http) |
TSV |
| GSE232597_FLAG_reproducible_enriched_windows.tsv.gz |
115.8 Kb |
(ftp)(http) |
TSV |
| GSE232597_RTCA_IP1_enriched_windows.tsv.gz |
9.2 Mb |
(ftp)(http) |
TSV |
| GSE232597_RTCA_IP2_enriched_windows.tsv.gz |
9.5 Mb |
(ftp)(http) |
TSV |
| GSE232597_RTCA_reproducible_enriched_windows.tsv.gz |
8.4 Mb |
(ftp)(http) |
TSV |
| GSE232597_SCAF8_IP1_enriched_windows.tsv.gz |
1.9 Mb |
(ftp)(http) |
TSV |
| GSE232597_SCAF8_IP2_enriched_windows.tsv.gz |
3.8 Mb |
(ftp)(http) |
TSV |
| GSE232597_SCAF8_reproducible_enriched_windows.tsv.gz |
1.5 Mb |
(ftp)(http) |
TSV |
| GSE232597_STAU2_IP1_enriched_windows.tsv.gz |
7.9 Mb |
(ftp)(http) |
TSV |
| GSE232597_STAU2_IP2_enriched_windows.tsv.gz |
7.4 Mb |
(ftp)(http) |
TSV |
| GSE232597_STAU2_reproducible_enriched_windows.tsv.gz |
6.1 Mb |
(ftp)(http) |
TSV |
| GSE232597_TRNAU1AP_IP1_enriched_windows.tsv.gz |
1.0 Mb |
(ftp)(http) |
TSV |
| GSE232597_TRNAU1AP_IP2_enriched_windows.tsv.gz |
820.4 Kb |
(ftp)(http) |
TSV |
| GSE232597_TRNAU1AP_reproducible_enriched_windows.tsv.gz |
573.9 Kb |
(ftp)(http) |
TSV |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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