Characterisation of three zinc finger proteins ZC3H8, ZC3H11A and ZC3H14, identified as new partners of ATM, the central regulator of biological response to DNA double strand breaks
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The integrity of our genome is very important therefore, our cells have developed efficient DNA repair mechanisms to control this integrity and avoid mutations. Although the DNA double strand break response pathway is well known, there are still many aspects of this pathway that required further characterisation. The ATM protein is a major player in the DNA Damage Response (DDR). Through a proteomic screen of ATM three C3H1-type zinc finger proteins ZC3H8, ZC3H11A and ZC3H14 were identified as new ATM interacting partners. These proteins had been implicated in several aspects of RNA biogenesis but nothing has been reported about their implication in the DDR. We found these three zinc finger proteins being chromatin bound proteins and forming foci within the nucleus, with ZC3H14 co-localising with Nuclear Speckles (SC35). Depletion of ZC3H11A and ZC3H14 induces spontaneous damage, which results in block of the replication fork since accumulation of BrdU foci and pRPA2 (S4/S8) foci is visible upon ZC3H11A and ZC3H14 knock down. R-loop resolution is also affected when ZC3H11A and ZC3H14 are depleted suggesting that their role in RNA biogenesis might be important to avoid transcription/replication collision. Human cells are sensitive to IR, ICRF-193, but not to Olaparib, when ZC3H14 is depleted. Depletion of ZC3H14 also leads to persistent γH2AX foci and pATM S1981 foci confirming a role for this protein in DSBs repair. ZC3H14 is involved in the NHEJ, but not in the HR, repair pathway. The recruitment to site of DNA double strand breaks for 53BP1, BRCA1, FK2, RNF168 and RNF8, but not MDC1, is affected upon ZC3H14 depletion suggesting that ZC3H14 is required for DSBs repair pathway choice. ZC3H14 interacts with MDC1 and its depletion increases the total MDC1 protein levels, before and after damage, with the MDC1 lower band being more abundant than the upper band. Surprisingly, depletion of ZC3H14 does not affect the MDC1-RNF8 interaction.
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