MicroRNA Clustering Assists Processing of Suboptimal MicroRNA Hairpins Through the Action of the ERH Protein. MicroRNA Clustering Assists Processing of Suboptimal MicroRNA Hairpins Through the Action of the ERH Protein

Microprocessor initiates processing of microRNAs (miRNAs) from hairpin regions of primary transcripts (pri-miRNAs). Pri-miRNAs often contain multiple miRNA hairpins, and this clustered arrangement can assist processing of otherwise defective hairpins. We find that miR-451, which derives from a hairpin with a suboptimal terminal loop and a suboptimal stem length, accumulates to 40-fold higher levels when clustered with a helper hairpin. This phenomenon tolerates changes in hairpin order, linker lengths, and the identities of the helper hairpin, the recipient hairpin, the linker-sequence, and the RNA polymerase that transcribes the hairpins. It can act reciprocally and need not occur co-transcriptionally. It requires Microprocessor recognition of the helper hairpin and linkage of the two hairpins, yet predominantly manifests after helper-hairpin processing. It also requires Enhancer of Rudimentary Homology (ERH), which copurifies with Microprocessor and can dimerize and interact with other proteins that can dimerize, suggesting a model in which Microprocessor recruits another Microprocessor. Overall design: In this study, ERH was knocked down using inducible CRISPR interference (CRISPRi) in K562-dCas9-KRAB cells; small-RNA sequencing was performed on both ERH knockdown and contorl (no doxycycline induction) cells.