The DNA damage response (DDR) preserves genomic integrity. Small non-coding RNAs termed DDRNAs are generated at DNA double-strand breaks (DSBs) and are critical for DDR activation. Here we show that active DDRNAs specifically localize to their damaged homologous genomic sites in a transcription-dependent manner. Upon DNA damage, RNA polymerase II (RNAPII) binds to the MRE11/RAD50/NBS1 complex, is recruited to DSBs and synthesizes damage-induced long non-coding RNAs (dilncRNAs) from and towards DNA ends. DilncRNAs act both as DDRNA precursors and by recruiting DDRNAs through RNA:RNA pairing. Together dilncRNAs and DDRNAs fuel DDR focus formation and associate with 53BP1. Accordingly, inhibition of RNAPII prevents DDRNA recruitment, DDR activation and DNA repair. Antisense oligonucleotides matching dilncRNAs and DDRNAs impair site-specific DDR focus formation and DNA repair. We propose that DDR signalling sites, in addition to sharing a common pool of proteins, individually host a unique set of site-specific RNAs necessary for DDR activation.

Medulloblastoma is an aggressive brain malignancy with high incidence in childhood. Current treatment approaches have limited efficacy and severe side effects. Therefore, new risk-adapted therapeutic strategies based on molecular classification are required. MicroRNA expression analysis has emerged as a powerful tool to identify candidate molecules playing an important role in a large number of malignancies. However, no data are yet available on human primary medulloblastomas. A high throughput microRNA expression profiles was performed in human primary medulloblastoma specimens to investigate microRNA involvement in medulloblastoma carcinogenesis. We identified specific micro-RNA expression patterns which distinguish medulloblastoma differing in histotypes (anaplastic, classic and desmoplastic), in molecular features (ErbB2 or c-Myc overexpressing tumors) and in disease-risk stratification. MicroRNAs expression profile clearly differentiates medulloblastoma from either adult or fetal normal cerebellar tissues. Only a few microRNAs displayed upregulated expression, while most of them were downregulated in tumor samples, suggesting a tumor growth-inhibitory function. This property has been addressed for miR-9 and miR-125a, whose rescued expression promoted medulloblastoma cell growth arrest and apoptosis while targeting the proproliferative truncated TrkC isoform. In conclusion, misregulated microRNA expression profiles characterize human medulloblastomas, and may provide potential targets for novel therapeutic strategies.

Damage-induced long non-coding RNAs (dilncRNA) synthesized at DNA double-strand breaks (DSBs) by RNA polymerase II (RNAPII) are necessary for DNA damage response (DDR) foci formation. We demonstrate that induction of DSBs results in the assembly of functional promoters that include a complete RNAPII pre-initiation complex (PIC), MED1 and CDK9. Absence or inactivation of these factors causes DDR foci reduction both in vivo and in an in vitro system that reconstitutes DDR events on nucleosomes. We also show that dilncRNAs drive molecular crowding of DDR proteins such as 53BP1 into foci that exhibit liquid-liquid phase separation (LLPS) condensate properties. We propose that the assembly of DSB-induced transcriptional promoters drives RNA synthesis which stimulates phase separation of DDR factors in the shape of foci. Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use: