Abstract
MicroRNA (miRNA) plays an important role in the control of gene expression. HYPONASTIC LEAVES1 (HYL1) is a double-stranded RNA-binding protein that forms a complex with DICER-LIKE1 (DCL1) and SERRATE (SE) to process primary miRNA (pri-miRNA) into mature miRNA. Although HYL1 has been shown to partner with DCL1 to enhance miRNA accuracy, the mechanism by which HYL1 selects the DCL1-targeted cleavage sites in pri-miRNA has remained unknown. By mutagenesis of HYL1 and analysis of in vivo pri-miRNA processing, we investigated the role of HYL1 in pri-miRNA cleavage. HYL1 forms homodimers in which the residues Gly147 and Leu165 in the dsRBD2 domain are shown to be critical. Disruption of HYL1 homodimerization causes incorrect cleavage at sites in pri-miRNA without interrupting the interaction of HYL1 with DCL1 and accumulation of pri-miRNAs in HYL1/pri-miRNA complexes, leading to a reduction in the efficiency and accuracy of miRNAs that results in strong mutant phenotypes of the plants. HYL1 homodimers may function as a molecular anchor for DCL1 to cleave at a distance from the ssRNA–dsRNA junction in pri-miRNA. These results suggest that HYL1 ensures the correct selection of pri-miRNA cleavage sites through homodimerization and thus contributes to gene silencing and plant development.
Highlights
MicroRNAs are ∼21 nucleotide small RNAs produced from partially paired stem-loop regions of primary miRNA transcripts [1]. miRNAs regulate gene expression at the post-transcriptional level by inhibiting the expression of mRNAs bearing fully or partly homologous target sequences [1,2,3]
DICER-LIKE1 (DCL1; the Arabidopsis homolog of Dicer) cleaves both pri-miRNA and pre-miRNA and generates miRNA in the nucleus [12,13], and the miRNAs are exported to the cytoplasm and incorporated in the RNA-induced silencing complex (RISC) for negatively regulation to gene expression through mRNA degradation or translation inhibition [13,14,15,16,17,18]
These findings indicate that HYPONASTIC LEAVES1 (HYL1) forms a homodimer through the dsRBD2 domains
Summary
MicroRNAs (miRNAs) are ∼21 nucleotide (nt) small RNAs produced from partially paired stem-loop regions of primary miRNA (pri-miRNA) transcripts [1]. miRNAs regulate gene expression at the post-transcriptional level by inhibiting the expression of mRNAs bearing fully or partly homologous target sequences [1,2,3]. MiRNAs regulate gene expression at the post-transcriptional level by inhibiting the expression of mRNAs bearing fully or partly homologous target sequences [1,2,3]. Pri-miRNAs are processed into miRNA precursors (pre-miRNAs) in nucleus by an RNaseIII-like enzyme named Drosha [7] that selectively cleaves RNA hairpins bearing a large terminal loop [8]. DICER-LIKE1 (DCL1; the Arabidopsis homolog of Dicer) cleaves both pri-miRNA and pre-miRNA and generates miRNA in the nucleus [12,13], and the miRNAs are exported to the cytoplasm and incorporated in the RNA-induced silencing complex (RISC) for negatively regulation to gene expression through mRNA degradation or translation inhibition [13,14,15,16,17,18]
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