{"DOI":"10.1093/nar/gkaa269","PMID":"32356894","abstract":"Abstract\n Studies on biological functions of N6-methyladenosine (m6A) modification in mRNA have drawn significant attention in recent years. Here we describe the construction and characterization of a CRISPR\u2013Cas13b-based tool for targeted demethylation of specific mRNA. A fusion protein, named dm6ACRISPR, was created by linking a catalytically inactive Type VI-B Cas13 enzyme from Prevotella sp. P5\u2013125 (dPspCas13b) to m6A demethylase AlkB homolog 5 (ALKBH5). dm6ACRISPR specifically demethylates m6A of targeted mRNA such as cytochrome b5 form A (CYB5A) to increase its mRNA stability. It can also demethylate \u03b2-catenin-encoding CTNNB1 mRNA that contains multiple m6A sites to trigger its translation. In addition, the dm6ACRISPR system incurs efficient demethylation of targeted epitranscriptome transcripts with limited off-target effects. Targeted demethylation of transcripts coding for oncoproteins such as epidermal growth factor receptor (EGFR) and MYC can suppress proliferation of cancer cells. Together, we provide a programmable and in vivo manipulation tool to study mRNA modification of specific genes and their related biological functions.","author":[{"family":"Li","given":"Jiexin"},{"family":"Chen","given":"Zhuojia"},{"family":"Chen","given":"Feng"},{"family":"Xie","given":"Guoyou"},{"family":"Ling","given":"Yuyi"},{"family":"Peng","given":"Yanxi"},{"family":"Lin","given":"Yu"},{"family":"Luo","given":"Nan"},{"family":"Chiang","given":"Cheng-Ming"},{"family":"Wang","given":"Hongsheng"}],"id":"unknown","issued":{"date-parts":[[2020,5,1]]},"language":"en","publisher":"Oxford University Press (OUP)","title":"Targeted mRNA demethylation using an engineered dCas13b-ALKBH5 fusion protein","type":"article-journal"}