Rapid and systematic analysis of the RNA recognition specificities of RNA-binding proteins.
Metazoan genomes encode hundreds of RNA-binding proteins (RBPs) but RNA-binding preferences for relatively few RBPs have been well defined. Current techniques for determining RNA targets, including in vitro selection and RNA co-immunoprecipitation, require significant time and labor investment. Here we introduce RNAcompete, a method for the systematic analysis of RNA binding specificities that uses a single binding reaction to determine the relative preferences of RBPs for short RNAs that contain a complete range of k-mers in structured and unstructured RNA contexts. We tested RNAcompete by analyzing nine diverse RBPs (HuR, Vts1, FUSIP1, PTB, U1A, SF2/ASF, SLM2, RBM4 and YB1). RNAcompete identified expected and previously unknown RNA binding preferences. Using in vitro and in vivo binding data, we demonstrate that preferences for individual 7-mers identified by RNAcompete are a more accurate representation of binding activity than are conventional motif models. We anticipate that RNAcompete will be a valuable tool for the study of RNA-protein interactions.
Ray
Debashish
D
Banting and Best Department of Medical Research, University of Toronto, Toronto, Ontario, Canada.
Kazan
Hilal
H
Chan
Esther T
ET
Peña Castillo
Lourdes
L
Chaudhry
Sidharth
S
Talukder
Shaheynoor
S
Blencowe
Benjamin J
BJ
Morris
Quaid
Q
Hughes
Timothy R
TR
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D030561:Databases, Nucleic Acid; D016678:Genome; D008969:Molecular Sequence Data; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D012313:RNA / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012372:ROC Curve; D013379:Substrate Specificity
D016601:RNA-Binding Proteins; D012313:RNA