GSE112632
Specific Parallel Analysis of RNA Ends
Many evolutionarily conserved miRNAs in plants regulate transcription factors with key functions in plant development. Hence, mutations in core components of the miRNA biogenesis machinery causes strong growth defects. An essential aspect of miRNA biogenesis is the precise excision of the small RNA from an arm of the fold-back precursor. Defects in the selection of the correct miRNA sequence will ultimately affect miRNA target specificity, plant development and other processes controlled by these small RNAs. Intriguingly, plant miRNA precursors are largely variable in size and shape and can be processed by different modes. Here, we optimized genomic approaches to detect processing intermediates during miRNA biogenesis. We identified and characterized an endogenous miRNA whose processing is triggered by a terminal branched loop. Plant miRNA processing can be initiated by internal bubbles, small terminal loops or branched loops followed by dsRNA segments of 15-17 bp. Interestingly, precision and efficiency vary with the processing modes suggesting intrinsic differences between miRNA biogenesis pathways. Despite the various potential structural determinants present in a single a miRNA precursor, we found that DCL1 is mostly guided by a predominant structural region in each precursor in wild-type plants. However, genomic studies of miRNA processing intermediates in fiery1, hyl1 and se mutants revealed the existence of cleavage signatures consistent with the recognition of alternative or cryptic processing determinants in miRNA precursors. The results provide a general view of the mechanisms underlying the specificity of miRNA biogenesis in plants. Overall design: Processing intermediates are indentified on wt and mutant plants