Direct observation of cytoskeleton-dependent trafficking of miRNA visualized by the introduction of pre-miRNA.

MicroRNA (miRNA) plays physiologically and pathologically important roles in post-transcriptional regulation. Although miRNA has been suggested to dynamically interact with cellular organelles, the dynamicity of intracellular miRNA behavior has remained unclear. Here, by introducing fluorescently labeled pre-miRNA into living cells, we improved the miRNA visualization method using exogenous miRNA precursors. Through the combination of our miRNA visualization method and single-molecule sensitive fluorescence microscopy, we quantitatively analyzed the process of miRNA maturation. Furthermore, single-particle tracking of fluorescent miRNA in cells revealed the directed movements of miRNA on cytoskeletal components (i.e., microtubules and actin filaments). Our results also suggest that cytoskeleton-dependent miRNA trafficking is associated with the interaction of miRNAs with the nucleus and the endoplasmic reticulum/Golgi apparatus. Our method should facilitate the elucidation of the mechanism and physiological significance of the subcellular localization and organelle interaction of miRNA.