RNA-protein interaction mapping via MS2- or Cas13-based APEX targeting.

Proc Natl Acad Sci U S A

Department of Genetics, Chan Zuckerberg Biohub, Stanford University, Stanford, CA 94305;

Published: September 2020

AI Article Synopsis

  • RNA-protein interactions play crucial roles in various cellular functions, necessitating better methods to study them in living cells.
  • Researchers utilized two systems, MS2-MCP and engineered CRISPR-Cas13, to direct an enzyme called APEX2 to specific RNAs, allowing for the identification of protein partners associated with human telomerase RNA (hTR).
  • This approach revealed new protein interactions and validated the role of mA demethylase ALKBH5 in modifying hTR and influencing telomerase activity.

Article Abstract

RNA-protein interactions underlie a wide range of cellular processes. Improved methods are needed to systematically map RNA-protein interactions in living cells in an unbiased manner. We used two approaches to target the engineered peroxidase APEX2 to specific cellular RNAs for RNA-centered proximity biotinylation of protein interaction partners. Both an MS2-MCP system and an engineered CRISPR-Cas13 system were used to deliver APEX2 to the human telomerase RNA hTR with high specificity. One-minute proximity biotinylation captured candidate binding partners for hTR, including more than a dozen proteins not previously linked to hTR. We validated the interaction between hTR and the -methyladenosine (mA) demethylase ALKBH5 and showed that ALKBH5 is able to erase the mA modification on endogenous hTR. ALKBH5 also modulates telomerase complex assembly and activity. MS2- and Cas13-targeted APEX2 may facilitate the discovery of novel RNA-protein interactions in living cells.

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Source
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7486720PMC
http://dx.doi.org/10.1073/pnas.2006617117DOI Listing

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