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| Status |
Public on Jul 08, 2020 |
| Title |
The DGCR8-UGU interaction in microRNA biogenesis |
| Organism |
Homo sapiens |
| Experiment type |
Non-coding RNA profiling by high throughput sequencing
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| Summary |
Human Microprocessor is a trimeric complex composed of the RNase III enzyme, DROSHA, and a dimer of DGCR8 (its cofactor). Microprocessor initiates the biogenesis of microRNA by processing primary microRNAs (pri-miRNAs), during which, its cleavage efficiency and accuracy are enhanced because DGCR8 interacts with the apical UGU motif of pri-miRNAs. However, the mechanism of DGCR8-UGU interaction and influence of this interaction on the expression of cellular miRNA are still elusive. In this study, we demonstrated that the Rhed domain (i.e., the RNA-binding heme domain, amino acids 285-478) of DGCR8 is sufficient to recognize and interact with UGU. In addition, we identified three amino acid residues in Rhed (amino acids 361-363), which are critical for the UGU interaction, and we showed that these residues are essential for the Microprocessor complex to accurately and efficiently process pri-miRNAs in vitro. Furthermore, we found that within the DGCR8 dimer, the UGU-binding site from just one monomer is capable of discriminating between UGU- and non-UGU-containing pri-miRNAs. Finally, we showed that these amino acids are more important for the expression of UGU miRNAs than non-UGU miRNAs in human cells. This study improves our understanding of the substrate-recognizing mechanism of DGCR8, and implicates the roles of this recognition in differentiating miRNA expression in human cells.
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| Overall design |
DGCR8-KO cells were transfected with pCK empty, pCK-DGCR8 WT or pCK-DGCR8 mut1 plasmid. Small RNA sequencing libraries of the transfected DGCR8-KO cells were generated using the TruSeq Small RNA Library Prep kit. The experiment was repeated three times.
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| Contributor(s) |
Nguyen TA, Dang TL, Li S, Nguyen TL, Nguyen TD |
| Citation(s) |
32620823 |
| Submission date |
Nov 11, 2019 |
| Last update date |
Jul 08, 2020 |
| Contact name |
Duc Trung Nguyen |
| E-mail(s) |
tdnguyen@connect.ust.hk
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| Phone |
+582 93510497
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| Organization name |
The Hong Kong University of Science and Technology
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| Department |
Life Science
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| Lab |
Anh Lab
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| Street address |
Clear Water Bay
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| City |
Hong Kong |
| ZIP/Postal code |
00000 |
| Country |
Hong Kong |
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| Platforms (1) |
| GPL24676 |
Illumina NovaSeq 6000 (Homo sapiens) |
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| Samples (9)
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| GSM4568420 |
DGCR8 KO transfected with pCK empty plasmid replicate 1 |
| GSM4568421 |
DGCR8 KO transfected with pCK empty plasmid replicate 2 |
| GSM4568422 |
DGCR8 KO transfected with pCK empty plasmid replicate 3 |
| GSM4568423 |
DGCR8 KO transfected with DGCR8 WT plasmid replicate 1 |
| GSM4568424 |
DGCR8 KO transfected with DGCR8 WT plasmid replicate 2 |
| GSM4568425 |
DGCR8 KO transfected with DGCR8 WT plasmid replicate 3 |
| GSM4568426 |
DGCR8 KO transfected with DGCR8 mut1 plasmid replicate 1 |
| GSM4568427 |
DGCR8 KO transfected with DGCR8 mut1 plasmid replicate 2 |
| GSM4568428 |
DGCR8 KO transfected with DGCR8 mut1 plasmid replicate 3 |
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| Relations |
| BioProject |
PRJNA588809 |
| SRA |
SRP229498 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE140209_rawcount.csv.gz |
10.3 Kb |
(ftp)(http) |
CSV |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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