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Global chromatin changes induced by altered tonicity interferes with DNA damage response signaling and DNA double-strand break repair
The results of our experiments reveal that global changes in chromatin structure achieved by hypotonic or hypertonic treatment have severe consequences on DDR signaling and DSB repair and thereby endanger genomic stability. Chromatin relaxation by itself results in transient arrest of cells in G2-phase, which might be due to ATM activation. However, in response to IR DDR signaling is reduced in terms of pATM, γH2AX and 53BP1 foci formation. Surprisingly, we still see a strong G2-checkpoint response upon DSB induction, which is dependent on ATR as well as on ATM. Further investigation of upstream signaling of ATM will help to understand these diverse responses mediated by ATM. Since we observe functional DNA end resection in globally relaxed chromatin, an ATR mediated checkpoint response is promoted. However, DSB repair by the main repair pathways is disturbed, as we see less cNHEJ and especially HRR. Also altEJ fails to backup cNHEJ and HRR, but SSA is three-fold increased upon hypotonic treatment. As this is seen in experiments utilizing reporter cell lines, it would be interesting to investigate the response of SSA to chromatin relaxation on IR-induced DSBs. While impairments of faithful DSB repair are seen and in addition only mutagenic DSB repair is functional, the modest effects found in survival are unexpected. However, as continuous hypotonic treatment is toxic to the cells, the adaptation of the protocol to transient treatment explains the small radiosensitizing effect observed. DDR signaling experiments and also measurements of CCP show that effects of global chromatin relaxation mediated by hypotonic treatment are completely reversible. Global chromatin condensation achieved by hypertonic treatment arrests cells in G1-, G2- and M-phase, while it enhances DDR signaling in terms of ATM activation and H2AX phosphorylation in response to IR. However, DSB repair efficiency of all investigated repair pathways is compromised (cNHEJ) or completely abrogated (HRR, altEJ and SSA). This might be explained by ...
Global chromatin changes induced by altered tonicity interferes with DNA damage response signaling and DNA double-strand break repair
The results of our experiments reveal that global changes in chromatin structure achieved by hypotonic or hypertonic treatment have severe consequences on DDR signaling and DSB repair and thereby endanger genomic stability. Chromatin relaxation by itself results in transient arrest of cells in G2-phase, which might be due to ATM activation. However, in response to IR DDR signaling is reduced in terms of pATM, γH2AX and 53BP1 foci formation. Surprisingly, we still see a strong G2-checkpoint response upon DSB induction, which is dependent on ATR as well as on ATM. Further investigation of upstream signaling of ATM will help to understand these diverse responses mediated by ATM. Since we observe functional DNA end resection in globally relaxed chromatin, an ATR mediated checkpoint response is promoted. However, DSB repair by the main repair pathways is disturbed, as we see less cNHEJ and especially HRR. Also altEJ fails to backup cNHEJ and HRR, but SSA is three-fold increased upon hypotonic treatment. As this is seen in experiments utilizing reporter cell lines, it would be interesting to investigate the response of SSA to chromatin relaxation on IR-induced DSBs. While impairments of faithful DSB repair are seen and in addition only mutagenic DSB repair is functional, the modest effects found in survival are unexpected. However, as continuous hypotonic treatment is toxic to the cells, the adaptation of the protocol to transient treatment explains the small radiosensitizing effect observed. DDR signaling experiments and also measurements of CCP show that effects of global chromatin relaxation mediated by hypotonic treatment are completely reversible. Global chromatin condensation achieved by hypertonic treatment arrests cells in G1-, G2- and M-phase, while it enhances DDR signaling in terms of ATM activation and H2AX phosphorylation in response to IR. However, DSB repair efficiency of all investigated repair pathways is compromised (cNHEJ) or completely abrogated (HRR, altEJ and SSA). This might be explained by ...
Global chromatin changes induced by altered tonicity interferes with DNA damage response signaling and DNA double-strand break repair
Krieger, Lisa Marie (Autor:in) / Iliakis, George
19.12.2018
Hochschulschrift
Elektronische Ressource
Englisch
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