Human mirtrons can express functional microRNAs simultaneously from both arms in a flanking exon-independent manner.

Mirtrons are short intronic microRNA (miRNA) precursors representing an alternative, Drosha/DGCR8-independent miRNA biogenesis pathway. In this study we characterized three predicted human mirtrons. Their expression was proven to be context-independent, since functional mirtrons could be derived either from their endogenous or from a heterologous coding environment. Systematic testing revealed that both 5'- and 3'-arms of mir-877 are capable of producing functional miRNA simultaneously in the various cell types examined. On the other hand, experimental validations revealed that the predicted mir-1233 is not a bona fide mirtron. For functional mirtrons, we were able to detect mature mirtron-derived miRNAs for the first time by qRT-PCR or northern blot analysis, when silencing activity was proven by functional assays. Our results emphasize the need for functional testing of both arms of miRNAs and the importance of experimentally validating human mirtrons since, in spite of being localized in a short intron, predicted species could mature via other miRNA processing pathways.