Flexible pri-miRNA structures enable tunable production of 5' isomiRs

Drosha cleavage of a pri-miRNA defines mature microRNA sequence. Drosha cleavage at alternative positions generates 5 isoforms (isomiRs) which have distinctive functions. To understand how pri-miRNA structures influence Drosha cleavage, we performed a systematic analysis of the maturation of endogenous pri-miRNAs and their variants both in vitro and in vivo. We show that, in addition to previously known features, the overall structural flexibility of pri-miRNA impacts Drosha cleavage fidelity. Internal loops and nearby G{middle dot}U wobble pairs on the pri-miRNA stem induce the use of non-canonical cleavage sites by Drosha, resulting in 5 isomiR production. By analyzing patient data deposited in The Cancer Genome Atlas, we provide evidence that alternative Drosha cleavage of pri-miRNAs is a tunable process that responds to the level of pri-miRNA-associated RNA-binding proteins. Together, our findings reveal that Drosha cleavage fidelity can be modulated by altering pri-miRNA structure, a potential mechanism underlying 5 isomiR biogenesis in tumors. HIGHLIGHTSO_LIFlexible pri-miRNA structures lead to 5 isomiR production C_LIO_LIInternal loops and G{middle dot}U pairs of pri-miRNA contribute to alternative Drosha cleavages C_LIO_LIAlternative Drosha cleavage results in 5 isomiRs from both strands of pre-miRNAs C_LIO_LI5 isomiR production is upregulated by pri-miRNA-associated RBPs in cancers C_LI GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/456839v1_ufig1.gif" ALT="Figure 1"> View larger version (37K): org.highwire.dtl.DTLVardef@cab9aorg.highwire.dtl.DTLVardef@1d613b1org.highwire.dtl.DTLVardef@1a6e200org.highwire.dtl.DTLVardef@13f7607_HPS_FORMAT_FIGEXP M_FIG C_FIG