Isomer-specific effect of microRNA miR-29b on nuclear morphology

Targeting mRNAs via seed region pairing is the canonical mechanism by which microRNAs (miRNAs) regulate cellular functions and disease processes. Emerging evidence suggests miRNAs might also act through other mechanisms. miRNA isomers that contain identical seed region sequences, such as miR-29a and miR-29b, provide naturally occurring, informative models for identifying those miRNA effects that are independent of seed region pairing. miR-29a and miR-29b are both expressed in HeLa cells, and miR-29b has been reported to localize to the nucleus in early mitosis because of unique nucleotide sequences on its 3' end. Here, we sought to better understand the mechanism of miR-29b nuclear localization and its function in cell division. We hypothesized that its nuclear localization may be facilitated by protein-miRNA interactions unique to miR-29b. Specific blockade of miR-29b resulted in striking nuclear irregularities not observed following miR-29a blockade. We also observed that miR-29b, but not miR-29a, is enriched in the nucleus and perinuclear clusters during mitosis. Targeted proteomic analysis of affinity-purified samples identified several proteins interacting with synthetic oligonucleotides mimicking miR-29b, but these proteins did not interact with miR-29a. One of these proteins, ADP/ATP translocase 2 (ANT2), known to be involved in mitotic spindle formation, colocalized with miR-29b in perinuclear clusters independently of Argonaute 2. Of note, ANT2 knockdown resulted in nuclear irregularities similar to those observed following miR-29b blockade and prevented nuclear uptake of endogenous miR-29b. Our findings reveal that miR-29 regulates nuclear morphology during mitosis and that this critical function is unique to the miR-29b isoform.