SUMOylation at K707 of DGCR8 controls direct function of primary microRNA

DGCR8 (DiGeorge syndrome critical region gene 8) is essential for primary microRNA (pri-miRNA) processing in the cell nucleus. It specifically combines with Drosha, a nuclear RNase III enzyme, to form the Microprocessor complex (MC) that cleaves pri-miRNA to precursor miRNA (pre-miRNA), which is further processed to mature miRNA by Dicer, a cytoplasmic RNase III enzyme. Increasing evidences suggest that pri-/pre-miRNAs have direct functions in regulation of gene expression, however the underlying mechanism how it is fine-tuned remains unclear. Here we find that DGCR8 is modified by SUMO1 at the major site K⁷⁰⁷, which can be promoted by its ERK-activated phosphorylation. SUMOylation of DGCR8 enhances the protein stability by preventing the degradation via the ubiquitin proteasome pathway. More importantly, SUMOylation of DGCR8 does not alter its association with Drosha, the MC activity and miRNA biogenesis, but rather influences its affinity with pri-miRNAs. This altered affinity of DGCR8 with pri-miRNAs seems to control the direct functions of pri-miRNAs in recognition and repression of the target mRNAs, which is evidently linked to the DGCR8 function in regulation of tumorigenesis and cell migration. Collectively, our data suggest a novel mechanism that SUMOylation of DGCR8 controls direct functions of pri-miRNAs in gene silencing.     

Homodimerization of HYL1 ensures the correct selection of cleavage sites in primary miRNA

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.     

A Highlights from MBoC Selection: Drosha protein levels are translationally regulated during Xenopus oocyte maturation

MicroRNAs (miRNAs) are ∼21-nucleotide-long, single-stranded noncoding RNAs that regulate gene expression. Biogenesis of miRNAs is mediated by the two RNase III-like enzymes, Drosha and Dicer. Here we study miRNA biogenesis during maturation of Xenopus oocytes to eggs using microinjection of pri-miRNAs. We show that processing of exogenous and endogenous primary miRNAs (pri-miRNAs) is strongly enhanced upon maturation of oocytes to eggs. Overexpression of cloned Xenopus Drosha in oocytes, however, boosts pri-miRNA processing dramatically, indicating that Drosha is a rate-limiting factor in Xenopus oocytes. This developmental regulation of Drosha is controlled by poly(A) length addition to the Drosha mRNA, which boosts translation upon transition from oocytes to eggs. Processing of pri-miRNAs by Drosha and Dicer has been shown to be affected by adenosine-to-inosine deamination–type RNA editing. Using activated Xenopus eggs for microinjection experiments, we demonstrate that RNA editing can reduce pri-miRNA processing in vivo. This processing block is determined by the structural but not sequence changes introduced by RNA editing.