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

Nat Methods. 2021 May;18(5):507-519. doi: 10.1038/s41592-021-01128-0. Epub 2021 May 7.

Abstract

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 ultraviolet cross-linking or immunoprecipitation and, when coupled with single-cell capture, can identify RBP-specific and cell-type-specific RNA-protein interactions for multiple RBPs and cell types in single, pooled experiments. Pairing STAMP with long-read sequencing yields RBP target sites in an isoform-specific manner. Finally, Ribo-STAMP leverages small ribosomal subunits to measure transcriptome-wide ribosome association in single cells. STAMP enables the study of RBP-RNA interactomes and translational landscapes with unprecedented cellular resolution.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Animals
  • Binding Sites
  • Gene Expression Profiling
  • HEK293 Cells
  • Humans
  • Nanopore Sequencing
  • RNA / chemistry
  • RNA / metabolism*
  • RNA-Binding Proteins / chemistry
  • RNA-Binding Proteins / metabolism*
  • Sequence Analysis, RNA
  • Single-Cell Analysis / methods*
  • Transcriptome

Substances

  • RNA-Binding Proteins
  • RNA