|
| Status |
Public on Dec 28, 2021 |
| Title |
The DGCR8 E518K mutation found in Wilms tumors leads to a partial miRNA processing defect that alters gene expression and biological processes (mRNA-seq) |
| Organism |
Mus musculus |
| Experiment type |
Expression profiling by high throughput sequencing
|
| Summary |
The microprocessor mutation DGCR8 E518K represents one of the most prevalent and stereotypical mutations in Wilms tumor (WT). It affects the dsRNA-binding domain (dsRBD) of DGCR8, which leads to a presumably disruptive charge reversal. The E518K mutation was expressed homozygously in all cases, indicative for the mutation to act in a recessive manner. Notably, this alteration has been almost exclusively reported in WT, suggesting a specific role in WT formation. To functionally characterize the DGCR8 E518K mutation in vitro, we employed DGCR8 knockout mouse embryonic stem cell (mESC) lines with inducible overexpression of wild-type (wt) or E518K-mutant DGCR8. Absence of endogenous DGCR8 mirrors the homozygous expression of mutant DGCR8 as observed in WT. Knockout of DGCR8 is known to result in a severe miRNA processing defect with a significant downregulation of all canonical miRNAs. Wild-type DGCR8 rescued this processing defect. DGCR8-E518K, in turn, was unable to fully restore miRNA expression and showed overall reduced miRNA expression levels, confirming our previous findings in bulk tumor tissue. Differentially expressed miRNAs comprised members of the embryonic stem cell specific cell cycle-regulating (ESCC) miRNAs and the let-7 miRNA family, whose antagonism is known to play a pivotal role in the regulation of critical gene expression programs in ES cells. Along with altered miRNA expression, DGCR8-E518K exhibited changes in target gene expression affecting various biological pathways. The known proliferation deficiency and cell cycle defect of DGCR8 KO mESCs could be rescued by DGCR8-E518K, albeit not to the full extent as by wild-type DGCR8. Likewise, DGCR8-E518K failed to fully block epithelial-to-mesenchymal transition (EMT). Upon embryoid body formation, however, both DGCR8-wt and -E518K were able to restore the differentiation capacity and to silence self-renewal in the knockout. Despite impaired miRNA processing and altered target gene regulation, DGCR8-E518K mESCs were still able to restore many biological processes in a DGCR8 knockout background. These findings suggest that partial reduction in activity or altered specificity might be critical in Wilms tumorigenesis.
|
| |
|
| Overall design |
We analyzed mRNA expression of DGCR8-KO, -E518K, and -wild-type mESCs after 48h induction with doxycycline (300 ng/mL). Two independent clones were used per condition.
|
| |
|
| Contributor(s) |
Vardapour R, Lenhof H, Kehl T, Kneitz S, Gessler M |
| Citation(s) |
34919667 |
| |
| Submission date |
Jan 21, 2021 |
| Last update date |
Dec 28, 2021 |
| Contact name |
Tim Kehl |
| E-mail(s) |
[email protected]
|
| Phone |
+49-681-302-68613
|
| Organization name |
Saarland University
|
| Department |
Center for Bioinformatics
|
| Street address |
Campus E2.1, P.O. box 151150
|
| City |
Saarbrücken |
| ZIP/Postal code |
66041 |
| Country |
Germany |
| |
|
| Platforms (1) |
|
| Samples (6)
|
|
| This SubSeries is part of SuperSeries: |
| GSE165270 |
The DGCR8 E518K mutation found in Wilms tumors leads to a partial miRNA processing defect that alters gene expression and biological processes |
|
| Relations |
| BioProject |
PRJNA693849 |
| SRA |
SRP302741 |
Less...
More...