Rewiring E2F1 with classical NHEJ via APLF suppression promotes bladder cancer invasiveness

Autor: Berdien A. H. Edelhäuser, George Iliakis, Fanghua Li, Christin Richter, Brigitte M. Pützer, Nico Murr, Stephan Marquardt, Alf Spitschak, Stella Logotheti
Rok vydání: 2019
Předmět:
0301 basic medicine
Genome instability
Transcriptional Activation
Cancer Research
endocrine system
Ku80
DNA End-Joining Repair
APLF
Non-homologous end-joining
Medizin
lcsh:RC254-282
03 medical and health sciences
0302 clinical medicine
Cell Line
Tumor
DNA-(Apurinic or Apyrimidinic Site) Lyase
E2F1
Humans
DNA Breaks
Double-Stranded
Neoplasm Invasiveness
Homologous Recombination
Poly-ADP-Ribose Binding Proteins
Promoter Regions
Genetic
Gene knockdown
Ku70
Chemistry
Research
Bladder cancer
miR-888
DNA repair protein XRCC4
DNA Methylation
lcsh:Neoplasms. Tumors. Oncology. Including cancer and carcinogens
Endonucleases
Cell biology
Non-homologous end joining
DNA-Binding Proteins
MicroRNAs
030104 developmental biology
DNA Repair Enzymes
Oncology
Urinary Bladder Neoplasms
030220 oncology & carcinogenesis
Gene Knockdown Techniques
biological phenomena
cell phenomena
and immunity
Homologous recombination
E2F1 Transcription Factor
Zdroj: Journal of Experimental & Clinical Cancer Research : CR
Journal of Experimental & Clinical Cancer Research, Vol 38, Iss 1, Pp 1-16 (2019)
ISSN: 1756-9966
Popis: 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. Electronic supplementary material The online version of this article (10.1186/s13046-019-1286-9) contains supplementary material, which is available to authorized users.
Databáze: OpenAIRE
Externí odkaz:
Nepřihlášeným uživatelům se plný text nezobrazuje