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Site-Specific m6A Erasing via Conditionally Stabilized CRISPR-Cas13b Editor
Authors:Ying Xu  Yufan Wang  Fu-Sen Liang
Institution:Department of Chemistry, Case Western Reserve University, 2080 Adelbert Rd, Cleveland, OH 44106 USA
Abstract:N6-methyladenosine (m6A) on RNAs plays an important role in regulating various biological processes and CRIPSR technology has been employed for programmable m6A editing. However, the bulky size of CRISPR protein and constitutively expressed CRISPR/RNA editing enzymes can interfere with the native function of target RNAs and cells. Herein, we reported a conditional m6A editing platform (FKBP*-dCas13b-ALK) based on a ligand stabilized dCas13 editor. The inducible expression of this m6A editing system was achieved by adding or removing the Shield-1 molecule. We further demonstrated that the targeted recruitment of dCas13b-m6A eraser fusion protein and site-specific m6A erasing were achieved under the control of Shield-1. Moreover, the release and degradation of dCas13b fusion protein occurred faster than the restoration of m6A on the target RNAs after Shield-1 removal, which provides an ideal opportunity to study the m6A function with minimal steric interference from bulky dCas13b fusion protein.
Keywords:Cas13b  Protein Degradation  RNA  Shield-1  m6A
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