Exportin 5 is a RanGTP-dependent dsRNA-binding protein that mediates nuclear export of pre-miRNAs

TLDR

MicroRNAs are ubiquitous in eukaryotes and are processed from primary transcripts into ~70‑nt stem–loop pre‑miRNAs that are exported to the cytoplasm for further maturation into ~22‑nt dsRNAs incorporated into ribonucleoprotein complexes. The study investigates the carrier‑mediated, saturable nuclear export of pre‑miRNAs. Using affinity chromatography with immobilized pre‑miRNAs, exportin 5 was identified as the specific carrier for pre‑miRNA export. Export of pre‑miRNAs is RanGTP‑dependent, mediated by exportin 5, which binds double‑stranded RNA in a sequence‑independent manner and can be inhibited by specific antibodies.

Abstract

microRNAs (miRNAs) are widespread among eukaryotes, and studies in several systems have revealed that miRNAs can regulate expression of specific genes. Primary miRNA transcripts are initially processed to ≈70-nucleotide (nt) stem–loop structures (pre-miRNAs), exported to the cytoplasm, further processed to yield ≈22-nt dsRNAs, and finally incorporated into ribonucleoprotein particles, which are thought to be the active species. Here we study nuclear export of pre-miRNAs and show that the process is saturable and thus carrier-mediated. Export is sensitive to depletion of nuclear RanGTP and, according to this criterion, mediated by a RanGTP-dependent exportin. An unbiased affinity chromatography approach with immobilized pre-miRNAs identified exportin 5 as the pre-miRNA-specific export carrier. We have cloned exportin 5 from Xenopus and demonstrate that antibodies raised against the Xenopus receptor specifically block pre-miRNA export from nuclei of Xenopus oocytes. We further show that exportin 5 interacts with double-stranded RNA in a sequence-independent manner.