Roncador, M. | Search bioRxiv

bioRxiv2018

Genomic landscape and chronological reconstruction of driver events in multiple myeloma

Multiple myeloma (MM) has a heterogeneous genome, evolving through both pre-clinical and post-diagnosis phases. Here, using sequences from 67 MM genomes serially collected from 30 patients together with public datasets, we establish a hierarchy of driver lesions. Point mutations, structural variants and copy number aberrations define at least 7 genomic subgroups of MM, each with distinct sets of co-operating driver mutations. Complex structural events are major drivers of MM, including chromothripsis, chromoplexy and a replication-based mechanism of templated insertions: these typically occur early. Hyperdiploidy also occurs early, with individual chromosomes often gained in more than one chronological epoch of MM evolution, showing a preferred order of acquisition. Positively selected point mutations frequently occur in later phases of disease development, as do structural variants involving MYC. Thus, initiating driver events of MM, drawn from a limited repertoire of structural and numerical chromosomal changes, shape preferred trajectories of subsequent evolution.

cancer biology↗

bioRxiv2018

DROSHA associates to DNA damage sites and is required for DNA repair

The DNA damage response (DDR) is the signaling cascade through which a cell recognizes DNA lesions, and promotes their resolution via the repair pathways of Non-Homologous End Joining (NHEJ), or Homologous Recombination (HR). We recently demonstrated that DROSHA boosts DDR signaling by processing damage-induced long non-coding RNAs into smaller DNA damage response RNAs (DDRNAs). However, the location at which DROSHA exerts its DDR functions, relative to sites of DNA damage, remains unknown.\n\nTo investigate DROSHAs localization during DDR activation, we used the DiVA cellular system, which allows the controlled induction of several DNA double strand breaks (DSBs) in the human genome. Indeed, by genome wide chromatin immunoprecipitation followed by next generation sequencing, we demonstrate that DROSHA associates with DSBs. In support of this, DSB-recruitment of DROSHA is detectable at the single-cell level by Proximity Ligation Assay between DROSHA and known DDR markers, and by DNA damage in situ ligation followed by Proximity Ligation Assay (DI-PLA), which demonstrates proximity of DROSHA to DNA ends. DROSHA recruitment occurs at both genic and inter-genic DSBs, suggesting that its recruitment is independent from ongoing transcription preceding damage generation. DROSHAs recruitment to DNA lesions occurs throughout the cell cycle, and with a preference for NHEJ-prone DSBs. Consistently, inhibition of the HR pathway increases DROSHA recruitment, and DROSHA knock down strongly impairs NHEJ efficiency in a GFP-reporter cellular system for monitoring NHEJ DNA repair. Overall, these results demonstrate that DROSHA acts locally at sites of DNA damage to promote NHEJ DNA repair.

cell biology↗