Transcript Slide 1
HR=homologous recombination NHEJ=Non-homologous end joining In HR, 100-1000 bases removed to make long 3’ overhang that invades copy on sister chromatid. Or other chromosome? (unlikely due to expected slow kinetics). This suggests greater dependence on RNR for HR than NHEJ (but what about salvage of what was just taken out? Or salvage imported in from the outside). If 25-mer defines position in genome uniquely, this implies that cells use much bigger sequences to make absolutely sure the invasion is in the right place, consistent with HR repair being a high fidelity route. dNTPs needed by HR NHEJ also important in S-phase We conclude that blocking the ability to generate dNTPs de novo does not significantly impact how well NHEJ proficient, G1/G0 enriched cells repair IRgenerated DSBs, consistent with our predictions. [HU] may be low enough that NHEJ is not impacted; higher [HU] may have killed both routes and this may explain why 200 uM was used in this study. NHEJ is important HR not that important And what is left, HR, depends on dNTPs Same data as previous Table rad51 Interacts with Lig4 = Pol alpha/delta inhibitor Rad51 focus positive cells were apparent in Xrs6 cells but not K1 cells, indicating HR is employed for DSB repair in G1 cells only when NHEJ is not available (Fig. 2C) Feedforward says don’t try HR without dNTPs Less so without DNA pol Note HR much slower than NHEJ for pure IR p53R2 Overexpression does not help if NHEJ is working fine in wild type cells. p53R2 is overexpressed p53R2 Overexpressed Helps cells with only HR active, i.e. NHEJ minus. Conclude: differences in RNR activity does not significantly determine if cells successfully repair double strand breaks, only which pathway (HR or NHEJ) they use to repair them Abstract, last line: “This may explain in part why G1/G0 cells, which have reduced ribonucleotide reductase activity, rely more on NHEJ for DSB repair.” More likely NHEJ is used in G1 due to slow kinetics of HR, supported by Fig. 2 In Intro: As a consequence, RNR activity rises in early S, and falls after G2 – a fluctuation that correlates well with the extent cells perform HR. Fails to say that S to G2 also lines up with sister chromatids being there, early S would discriminate, i.e. if HR is very active in early S then they are right. NHEJ’s ability to sustain efficient repair regardless of the activity of the de novo pathway is thus a strong rationale for using a repair pathway suggested to be intrinsically more error prone than the alternative (HR). I disagree here, I think it is for faster kinetics.