RNase-mediated protein footprint sequencing reveals protein-binding sites throughout the human transcriptome

Genome Biol. 2014 Jan 7;15(1):R3. doi: 10.1186/gb-2014-15-1-r3.

Abstract

Although numerous approaches have been developed to map RNA-binding sites of individual RNA-binding proteins (RBPs), few methods exist that allow assessment of global RBP-RNA interactions. Here, we describe PIP-seq, a universal, high-throughput, ribonuclease-mediated protein footprint sequencing approach that reveals RNA-protein interaction sites throughout a transcriptome of interest. We apply PIP-seq to the HeLa transcriptome and compare binding sites found using different cross-linkers and ribonucleases. From this analysis, we identify numerous putative RBP-binding motifs, reveal novel insights into co-binding by RBPs, and uncover a significant enrichment for disease-associated polymorphisms within RBP interaction sites.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Binding Sites
  • Genetic Association Studies
  • HeLa Cells
  • Humans
  • Polymorphism, Single Nucleotide
  • Protein Footprinting / methods*
  • RNA-Binding Proteins / genetics
  • RNA-Binding Proteins / metabolism
  • Ribonucleases / genetics
  • Ribonucleases / metabolism
  • Sequence Analysis, RNA
  • Transcriptome / genetics*

Substances

  • RNA-Binding Proteins
  • Ribonucleases