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Rewiring E2F1 with classical NHEJ via APLF suppression promotes bladder cancer invasiveness.
- Source :
- Journal of Experimental & Clinical Cancer Research (17569966); 7/8/2019, Vol. 38 Issue 1, pN.PAG-N.PAG, 1p
- Publication Year :
- 2019
-
Abstract
- Background: Bladder cancer progression has been associated with dysfunctional repair of double-strand breaks (DSB), a deleterious type of DNA lesions that fuel genomic instability. Accurate DSB repair relies on two distinct pathways, homologous recombination (HR) and classical non-homologous end-joining (c-NHEJ). The transcription factor E2F1 supports HR-mediated DSB repair and protects genomic stability. However, invasive bladder cancers (BC) display, in contrast to non-invasive stages, genomic instability despite their high E2F1 levels. Hence, E2F1 is either inefficient in controlling DSB repair in this setting, or rewires the repair apparatus towards alternative, error-prone DSB processing pathways. Methods: RT-PCR and immunoblotting, in combination with bioinformatics tools were applied to monitor c-NHEJ factors status in high-E2F1-expressing, invasive BC versus low-E2F1-expressing, non-invasive BC. In vivo binding of E2F1 on target gene promoters was demonstrated by ChIP assays and E2F1 CRISPR-Cas9 knockdown. MIR888-dependent inhibition of APLF by E2F1 was demonstrated using overexpression and knockdown experiments, in combination with luciferase assays. Methylation status of MIR888 promoter was monitored by methylation-specific PCR. The changes in invasion potential and the DSB repair efficiency were estimated by Boyden chamber assays and pulse field electrophoresis, correspondingly. Results: Herein, we show that E2F1 directly transactivates the c-NHEJ core factors Artemis, DNA-PKcs, ligase IV, NHEJ1, Ku70/Ku80 and XRCC4, but indirectly inhibits APLF, a chromatin modifier regulating c-NHEJ. Inhibition is achieved by miR-888-5p, a testis-specific, X-linked miRNA which, in normal tissues, is often silenced via promoter methylation. Upon hypomethylation in invasive BC cells, MIR888 is transactivated by E2F1 and represses APLF. Consequently, E2F1/miR-888/APLF rewiring is established, generating conditions of APLF scarcity that compromise proper c-NHEJ function. Perturbation of the E2F1/miR-888/APLF axis restores c-NHEJ and ameliorates cell invasiveness. Depletion of miR-888 can establish a 'high E2F1/APLF/DCLRE1C' signature, which was found to be particularly favorable for BC patient survival. Conclusion: Suppression of the 'out-of-context' activity of miR-888 improves DSB repair and impedes invasiveness by restoring APLF. [ABSTRACT FROM AUTHOR]
- Subjects :
- CANCER invasiveness
BLADDER cancer
DNA damage
EIGENFUNCTIONS
TRANSCRIPTION factors
Subjects
Details
- Language :
- English
- ISSN :
- 17569966
- Volume :
- 38
- Issue :
- 1
- Database :
- Complementary Index
- Journal :
- Journal of Experimental & Clinical Cancer Research (17569966)
- Publication Type :
- Academic Journal
- Accession number :
- 137397734
- Full Text :
- https://doi.org/10.1186/s13046-019-1286-9