In vivo nuclear RNA structurome reveals the RNA-structure regulation of mRNA processing in plants

mRNA processing is critical for gene expression. A challenge in regulating mRNA processing is how to recognize the actual mRNA processing sites such as splice sites and polyadenylation sites when the sequence content is insufficient for this purpose. Previous studies suggested that RNA structure affects mRNA processing. However, the regulatory role of RNA structure in mRNA processing remains unclear. Here, we performed in vivo SHAPE chemical profiling on Arabidopsis nuclear RNAs and generated the in vivo nuclear RNA structure landscape. We found that nuclear mRNAs fold differently from cytosolic mRNAs. Notably, we discovered a two-nucleotide single-stranded RNA structure feature upstream of 5’ splice site that regulates splicing and is responsible for the selection of alternative 5’ splice sites. Moreover, we found branch point single-strandedness is associated with 3’ splice site recognition. We also identified an RNA structure feature comprising two close-by single-stranded regions that is specifically associated with both polyadenylation and alternative polyadenylation events. Our work demonstrates an RNA structure regulatory mechanism for mRNA processing. NAI treatment was applied to Arabidopsis seedlings under in vivo and deproteinized conditions. Control samples without NAI treatment were used for each conditions. RNA-sequencing was performed for NAI treated and control samples by using Illumina Hiseq 4000.