TREX2 Exonuclease Causes Spontaneous Mutations and Stress-Induced Replication Fork Defects in Cells Expressing RAD51(K133A)
Authors
KO, Jun Ho, Mi Young SON, Qing ZHOU, Lucia MOLNÁROVÁ (703 Slovakia, belonging to the institution), Lambert SONG, Jarmila MLCOUSKOVA, Atis JEKABSONS (428 Latvia, belonging to the institution), Cristina MONTAGNA, Lumír KREJČÍ (203 Czech Republic, belonging to the institution) and Paul HASTY
DNA damage tolerance (DDT) and homologous recombination (HR) stabilize replication forks (RFs). RAD18/UBC13/three prime repair exonuclease 2 (TREX2)-mediated proliferating cell nuclear antigen (PCNA) ubiqui-tination is central to DDT, an error-prone lesion bypass pathway. RAD51 is the recombinase for HR. The RAD51(K133A) mutation increased spontaneous mutations and stress-induced RF stalls and nascent strand degradation. Here, we report in RAD51(K133A) cells that this phenotype is reduced by expressing a TREX2 H188A mutation that deletes its exonuclease activity. In RAD51(K133A) cells, knocking out RAD18 or overexpressing PCNA reduces spontaneous mutations, while expressing ubiquitination-incompetent PCNA(K164R )increases mutations, indicating DDT as causal. Deleting TREX2 in cells deficient for the RF maintenance proteins poly(ADP-ribose) polymerase 1 (PARP1) or FANCB increased nascent strand degradation that was rescued by TREX2(H188A), implying that TREX2 prohibits degradation independent of catalytic activity. A possible explanation for this occurrence is that TREX2(H188A) associates with UBC13 and ubiquitinates PCNA, suggesting a dual role for TREX2 in RF maintenance.
Links
EF16_027/0008360, research and development project
Name: Postdoc@MUNI
GA17-17720S, research and development project
Name: Vnitřní vlastnosti RAD51 vlákna a jeho biologické regulace
Investor: Czech Science Foundation
MUNI/G/1594/2019, interní kód MU
Name: RecQ4 – a protein hub required for proper replication and recombination and its implications in Rothmund-Thomson Syndrome
Investor: Masaryk University, INTERDISCIPLINARY - Interdisciplinary research projects
21-22593X, interní kód MU
Name: Identification and characterization of proteins involved in metabolism of G-quadruplexes and R-loops and molecular mechanisms of their relationship with replication