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Large-scale evaluation of the ability of RNA-binding proteins to activate exon inclusion.

Jonathan C Schmok, Manya Jain, Lena A Street, Alex T Tankka, Danielle Schafer, Hsuan-Lin Her, Sara Elmsaouri, Maya L Gosztyla, Evan A Boyle, Pratibha Jagannatha, En-Ching Luo, Ester J Kwon, Marko Jovanovic, Gene W Yeo

Nature biotechnology · 2024 · Vol. 42 (9) · pp. 1429-1441

PubMed ↗ DOI ↗ PMC full text ↗

Abstract

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 developed tethered function luciferase-based splicing reporters that provide rapid, scalable and robust readouts of exon inclusion changes and used these to evaluate 718 human RBPs. We performed enhanced cross-linking immunoprecipitation, RNA sequencing and affinity purification-mass spectrometry to investigate a subset of candidates with no prior association with splicing. Integrative analysis of these assays indicates surprising roles for TRNAU1AP, SCAF8 and RTCA in the modulation of 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 these identified domains, we engineered programmable fusion proteins that outperform current artificial splicing factors at manipulating inclusion of reporter and endogenous exons. This tethering approach characterizes the ability of RBPs to induce exon inclusion and yields new molecular parts for programmable splicing control.

Linked datasets 3

GSE232599 GSE · via ncbi_elink
GEO ↗

Systematic identification of RNA-binding proteins and tethered domains that activate exon splicing inclusion

Homo sapiens

GSE232598 GSE · via ncbi_elink
GEO ↗

Systematic identification of RNA-binding proteins and tethered domains that activate exon splicing inclusion [RNA-seq]

Homo sapiens

30 data files
FileTypeSize
shNT_1_R1.fastq.gz RNA-Seq 5.0 GB
shNT_2_R1.fastq.gz RNA-Seq 5.2 GB
shNT_3_R1.fastq.gz RNA-Seq 6.3 GB
shRTCA_1_R1.fastq.gz RNA-Seq 5.3 GB
shRTCA_2_R1.fastq.gz RNA-Seq 4.7 GB
shRTCA_3_R1.fastq.gz RNA-Seq 5.0 GB
shSCAF8_1_R1.fastq.gz RNA-Seq 4.9 GB
shSCAF8_2_R1.fastq.gz RNA-Seq 5.3 GB
shSCAF8_3_R1.fastq.gz RNA-Seq 4.8 GB
shSTAU2_1_R1.fastq.gz RNA-Seq 4.8 GB
shSTAU2_2_R1.fastq.gz RNA-Seq 4.6 GB
shSTAU2_3_R1.fastq.gz RNA-Seq 4.7 GB
shTRNAU1AP_1_R1.fastq.gz RNA-Seq 5.6 GB
shTRNAU1AP_2_R1.fastq.gz RNA-Seq 4.8 GB
shTRNAU1AP_3_R1.fastq.gz RNA-Seq 5.6 GB
SRR24579591_GSM7359814 RNA-Seq 6.3 GB
SRR24579592_GSM7359813 RNA-Seq 5.2 GB
SRR24579593_GSM7359812 RNA-Seq 5.0 GB
SRR24579597_GSM7359808 RNA-Seq 4.7 GB
SRR24579598_GSM7359807 RNA-Seq 4.6 GB
SRR24579599_GSM7359806 RNA-Seq 4.8 GB
SRR24579600_GSM7359805 RNA-Seq 4.8 GB
SRR24579601_GSM7359804 RNA-Seq 5.3 GB
SRR24579602_GSM7359803 RNA-Seq 4.9 GB
SRR24579603_GSM7359802 RNA-Seq 5.0 GB
SRR24579604_GSM7359801 RNA-Seq 4.7 GB
SRR24579605_GSM7359800 RNA-Seq 5.3 GB
SRR24579606_GSM7359799 RNA-Seq 5.6 GB
SRR24579607_GSM7359798 RNA-Seq 4.8 GB
SRR24579608_GSM7359797 RNA-Seq 5.6 GB
GSE232597 GSE · via ncbi_elink
GEO ↗

Systematic identification of RNA-binding proteins and tethered domains that activate exon splicing inclusion [eCLIP-seq]

Homo sapiens

40 data files
FileTypeSize
FLAG_IN1.fastq.gz RIP-Seq 928.9 MB
FLAG_IN2.fastq.gz RIP-Seq 1.0 GB
FLAG_IP1.fastq.gz RIP-Seq 864.1 MB
FLAG_IP2.fastq.gz RIP-Seq 802.7 MB
RTCA_IN1.fastq.gz RIP-Seq 824.8 MB
RTCA_IN2.fastq.gz RIP-Seq 808.7 MB
RTCA_IP1.fastq.gz RIP-Seq 816.4 MB
RTCA_IP2.fastq.gz RIP-Seq 858.7 MB
SCAF8_IN1.fastq.gz RIP-Seq 746.9 MB
SCAF8_IN2.fastq.gz RIP-Seq 882.8 MB
SCAF8_IP1.fastq.gz RIP-Seq 782.9 MB
SCAF8_IP2.fastq.gz RIP-Seq 852.6 MB
SRR24579324_GSM7359796 RIP-Seq 802.0 MB
SRR24579325_GSM7359795 RIP-Seq 864.0 MB
SRR24579326_GSM7359794 RIP-Seq 1.0 GB
SRR24579327_GSM7359793 RIP-Seq 928.0 MB
SRR24579332_GSM7359788 RIP-Seq 876.0 MB
SRR24579333_GSM7359787 RIP-Seq 925.0 MB
SRR24579334_GSM7359786 RIP-Seq 902.0 MB
SRR24579335_GSM7359785 RIP-Seq 805.0 MB
SRR24579336_GSM7359784 RIP-Seq 852.0 MB
SRR24579337_GSM7359783 RIP-Seq 782.0 MB
SRR24579338_GSM7359782 RIP-Seq 882.0 MB
SRR24579339_GSM7359781 RIP-Seq 746.0 MB
SRR24579340_GSM7359780 RIP-Seq 858.0 MB
SRR24579341_GSM7359779 RIP-Seq 816.0 MB
SRR24579342_GSM7359778 RIP-Seq 808.0 MB
SRR24579343_GSM7359777 RIP-Seq 824.0 MB
SRR24579344_GSM7359776 RIP-Seq 819.0 MB
SRR24579345_GSM7359775 RIP-Seq 514.0 MB
SRR24579346_GSM7359774 RIP-Seq 493.0 MB
SRR24579347_GSM7359773 RIP-Seq 652.0 MB
STAU2_IN1.fastq.gz RIP-Seq 805.7 MB
STAU2_IN2.fastq.gz RIP-Seq 902.6 MB
STAU2_IP1.fastq.gz RIP-Seq 925.0 MB
STAU2_IP2.fastq.gz RIP-Seq 876.9 MB
TRNAU1AP_IN1.fastq.gz RIP-Seq 652.9 MB
TRNAU1AP_IN2.fastq.gz RIP-Seq 493.3 MB
TRNAU1AP_IP1.fastq.gz RIP-Seq 514.1 MB
TRNAU1AP_IP2.fastq.gz RIP-Seq 819.3 MB

Potentially related datasets 1

These accessions were text-mined from the PMC full text. They may be referenced for comparison, cited from other studies, or otherwise mentioned without being primary data for this paper.

Accession Source Title
GSE232599 GSE · GEO GEO ↗ Systematic identification of RNA-binding proteins and tethered domains that activate exon splicing inclusion

Analysis pipelines 3

Pipeline Assay Steps Dataset
GSE232597 eCLIP Data Processing geo_data_processing eCLIP 5 GSE232597
GSE232598 RNA-seq Data Processing geo_data_processing RNA-seq 6 GSE232598
GSE232599 eCLIP Data Processing geo_data_processing eCLIP 5 GSE232599
Funding 16 grants
Grant Funder
U24 HG009889 NHGRI NIH HHS (United States)
S10 OD026929 NIH HHS (United States)
R01 HG004659 NHGRI NIH HHS (United States)
U54 CA209891 NCI NIH HHS (United States)
T32 CA067754 NCI NIH HHS (United States)
DGE-2038238 National Science Foundation (NSF)
R01 HG012216 NHGRI NIH HHS (United States)
R35 GM128802 NIGMS NIH HHS (United States)
R01 AG071869 NIA NIH HHS (United States)
P41 GM103504 NIGMS NIH HHS (United States)
R01HG012216 U.S. Department of Health & Human Services | National Institutes of Health (NIH)
R01AG071869 U.S. Department of Health & Human Services | National Institutes of Health (NIH)
R35GM128802 U.S. Department of Health & Human Services | National Institutes of Health (NIH)
2224211 National Science Foundation (NSF)
PGS D-532649-2019 Gouvernement du Canada | Natural Sciences and Engineering Research Council of Canada (Conseil de Recherches en Sciences Naturelles et en Génie du Canada)
T32CA067754 U.S. Department of Health & Human Services | National Institutes of Health (NIH)