Journal
FEBS OPEN BIO
Volume 9, Issue 11, Pages 1860-1868Publisher
WILEY
DOI: 10.1002/2211-5463.12706
Keywords
APEX2; birA*; nascent peptide; proximity labeling; RNA binding protein; RNA-protein interactions
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RNA biology is orchestrated by the dynamic interactions of RNAs and RNA-binding proteins (RBPs). In the present study, we describe a new method of proximity-dependent protein labeling to detect RNA-protein interactions [RNA-bound protein proximity labeling (RBPL)]. We selected the well-studied RNA-binding protein PUF to examine the current proximity labeling enzymes birA* and APEX2. A new version of birA*, BASU, was used to validate that the PUF protein binds its RNA motif. We further optimized the RBPL labeling system using an inducible expression system. The RBPL (lambda N-BASU) labeling experiments exhibited high signal-to-noise ratios. We subsequently determined that RBPL (lambda N-BASU) is more suitable than RBPL (lambda N-APEX2) for the detection of RNA-protein interactions in live cells. Interestingly, our results also reveal that proximity labeling is probably capable of biotinylating proximate nascent peptide.
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