Robust single-cell discovery of RNA targets of RNA binding proteins and ribosomes

doi:10.21203/rs.3.rs-87224/v1

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Robust single-cell discovery of RNA targets of RNA binding proteins and ribosomes

https://doi.org/10.21203/rs.3.rs-87224/v1

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published 01 May, 2021

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RNA binding proteins (RBPs) are critical regulators of gene expression and RNA processing that are required for gene function. Yet, the dynamics of RBP regulation in single cells is unknown. To address this gap in understanding, we developed STAMP (Surveying Targets by APOBEC Mediated Profiling), which efficiently detects RBP-RNA interactions. STAMP does not rely on UV-crosslinking or immunoprecipitation and, when coupled with single-cell capture, can identify RBP- and cell type-specific RNA-protein interactions for multiple RBPs and cell types in single-pooled experiments. Pairing STAMP with long-read sequencing also yields RBP target sites for full-length isoforms. Finally, conducting STAMP using small ribosomal subunits (Ribo-STAMP) allows analysis of transcriptome-wide ribosome association in single cells. STAMP enables the study of RBP-RNA interactomes and translational landscapes with unprecedented cellular resolution.

Biotechnology and Bioengineering

RNA binding proteins (RBPs)

RBP regulation

STAMP (Surveying Targets by APOBEC Mediated Profiling)

RNA binding proteins and ribosomes

GWY is co-founder, member of the Board of Directors, on the SAB, equity holder, and paid consultant for Locana and Eclipse BioInnovations. GWY is a visiting professor at the National University of Singapore. GWY’s interests have been reviewed and approved by the University of California, San Diego in accordance with its conflict of interest policies. The authors declare no other competing financial interests.

TableS1.xlsx
Table S1. All control-STAMP edit site coordinates and levels; related to Figures 1-4 and Figures S1-S4
TableS2.xlsx
Table S2. All RBP-STAMP (RBFOX2, SLBP, TIA1) edit site coordinates and levels; related to Figures 1, 2, and Figures S1, S2
TableS3.xlsx
Table S3. Differential gene expression from RBFOX2-STAMP as evaluated by DEseq2
TableS4.xlsx
Tabls S4. All Ribo-STAMP (RPS2, RPS3, GSTP1) edit site coordinates and levels; related to Figure 3 and Figure S3
TableS5.xlsx
Tabls S5. Differential gene editing between background-cluster and RBFOX2- and TIA1-clusters, related to Figure 6D
TableS6.xlsx
Table S6. Differential gene expression between HEK293T cells and NPCs, related to Figure 6F and S6E
TableS7.xlsx
Table S7. Differential gene editing between HEK293T cells and NPCs for RBFOX2-STAMP, related to Figure 6G-J
TableS8.xlsx
Table S8. Differential gene editing between background-cluster and RPS2-,RBFOX2- and TIA1-clusters, related to Figure 7D
TableS9.xlsx
Table S9. Differential gene editing between RPS2-cluster subsets, related to Figure 7G
TableS10.xlsx
Table S10. Differential gene editing between cell cycle phase subsets, related to Figure 7I and Figure S7I
TableS11.xlsx
Table S11. Oligonucleotides used in this study

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Robust single-cell discovery of RNA targets of RNA binding proteins and ribosomes

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