Targeted mRNA demethylation in Arabidopsis using the human ALKBH5 catalytic domain

Abstract N6-methyladenosine (m6A) is an important epigenetic modification involved in RNA stability and translation regulation. Manipulating the expression of RNA m6A methyltransferases or demethylases makes it difficult to study the effect of specific RNA methylation. Recently, m6A editor was reported to manipulate RNA methylation in a locus-specific manner in human cells, but it remains unclear whether this tool can be used in plants. Here, we report the development of a plant m6A editors (PMEs) using dLwaCas13a (from L. wadei) and human m6A demethylase ALKBH5 catalytic domain. PMEs specifically demethylates m6A of targeted mRNAs (WUS, STM, FT, SPL3 and SPL9) to increase mRNAs stability. In addition, we discovered that a double ribozyme system can significantly improve the efficiency of RNA editing. Together, these findings demonstrated that PMEs for targeted transcript editing will be useful in plant functional genomics research.