PERSISTENT DNA DAMAGE AT TELOMERES, CAUSED BY TRF2-MEDIATED DNA REPAIR INHIBITION, TRIGGERS CELLULAR SENESCENCE AND IS ASSOCIATED WITH PRIMATES AGEING [Tesi di dottorato]
Università degli Studi di Milano, 2014-03-25

The DNA damage response (DDR) coordinates DNA repair events and transiently arrests cell-cycle progression until DNA damage has been removed. If the damage is not resolved, cells can enter an irreversible cell cycle arrest called cellular senescence. In irradiation-induced senescent cells a large fraction of persistent DDR markers are associated with telomeric DNA, both in cultured cells and in in vivo tissues. The aim of my PhD project was to investigate the mechanism underlying this phenomenon. I showed that persistent DDR activation has a causative role for the senescence-associated cell cycle exit and that a double-strand break (DSB) within telomeric repeats is inducing a more protracted DDR activation compared with a non-telomeric one in human cells. The DDR persistency at telomeres is neither dependent on their heterochromatic state nor on TRF2 loss from telomeres during senescence establishment. Rather, TRF2 recruitment next to a DSB, in the absence of telomeric DNA, is sufficient to induce a more protracted site-specific DDR focus and to impair DSB repair in mouse cells. Ageing is associated with accumulation of markers of DDR activation. In terminally differentiated brain neurons from old primates, I observed DDR activation at telomeres that were not critically short. Taken together, these results strongly suggest that TRF2 inhibits DNA repair at broken telomeres, contributing to the accumulation of unrepaired, DDR-positive telomeres during ageing. This can in turn trigger cellular senescence and impair tissue homeostasis providing a mechanism for ageing also in non-proliferating tissues. Finally, I focused my attention on DICER and DROSHA-dependent DNA damage response RNAs (DDRNAs), novel components of the DDR machinery, which have been described to be necessary for DDR activation at DSBs. I showed that RNase A treatment as well as DICER or DROSHA down-regulation impair DDR activation at uncapped telomeres and that DICER and DROSHA may have a role in chromosomal fusions. Furthermore, in cells with dysfunctional telomeres, the inhibition of telomeric DDRNAs using inhibitory oligonucleotide molecules with a complementary sequence can prevent DDR activation and senescent-associated cell cycle arrest. These data indicate that at uncapped telomeres, DDRNAs with telomeric sequences are generated and that they are necessary for DDR activation and chromosomal fusions.

diritti: info:eu-repo/semantics/openAccess
supervisor: F. d'Adda di Fagagna ; internal cosupervisor: M. Foiani ; external cosupervisor: M.A. Blasco
FOIANI, MARCO
Settore BIO/11 - - Biologia Molecolare


Tesi di dottorato. | Lingua: Inglese. | Paese: | BID: TD16001649