SUMOylation at K707 of DGCR8 controls direct function of primary microRNA.

Changhong Zhu,Jinzhuo Dou,Cheng Chen,Hailong Zhang,Rong Deng,Jian Huang,Yanli Wang,Ming Xu,Hui Jin,Jianxiu Yu,Qin Chen,Xian Zhao,Ran Chen

doi:10.1093/nar/gkv741

Abstract

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.

Highlights

IntroductionA long primary transcript known as a pri-miRNA in the cell nucleus is cleaved by a Microprocessor complex (MC), which is mainly composed of Drosha, an RNase III enzyme and DGCR8, a double-stranded RNAbinding protein [1,2,3,4], to generate a characteristic stem-loop structure of about 70 bp long, known as a pre-miRNA
The microRNA biogenesis pathway has been thoroughly uncovered
The relative molecular weight (MW) of DGCR8 shifted from 115 to 135 kDa was observed in the presence of His-SUMO1DGCR8 and (SUMO1) alone, and the 135- and 155-kDa bands were greatly strengthened with additional plasmid Flag-Ubc9 (Figure 1B), indicating that DGCR8 can be modified by SUMO1 at multiple sites

Summary

Introduction

A long primary transcript known as a pri-miRNA in the cell nucleus is cleaved by a Microprocessor complex (MC), which is mainly composed of Drosha, an RNase III enzyme and DGCR8, a double-stranded RNAbinding protein [1,2,3,4], to generate a characteristic stem-loop structure of about 70 bp long, known as a pre-miRNA The latter molecule is subsequently exported by exportin to the cytoplasm and further cleaved into an ∼20–25-bp double-stranded RNA fragment by another RNAIII enzyme Dicer. Chen’s group has first reported that the different activities of miR-181a-1 and miR181c, which are members of the same miRNA gene family, are dependent on their pre-miRNA loop nucleotides other than nucleotide difference in their mature miRNA sequences [5] Later they found that pri-let-7 can directly interact with target mRNAs to show a direct function in target repression, whose activity is determined on loop nucleotides by modulating interactions between pri-let-7 and target mRNAs [6,7]. It has become increasingly clear that pri-/pre-miRNAs can serve as posttranscriptional regulators of miRNA activity besides as biogenesis intermediates

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Conclusion