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DNA Polymerases Are Error-Prone at Reca-Mediated Recombination

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DNA Polymerases Are Error-Prone at Reca-Mediated Recombination PAPER TYPE Cell Cycle 12:16, 2558–2563; August 15, 2013; © 2013 Landes Bioscience DNA polymerases are error-prone at RecA-mediated recombination intermediates Richard T Pomerantz1,*, Myron F Goodman2, and Michael E O’Donnell3 1Fels Institute for Cancer Research; Department of Biochemistry; Temple University School of Medicine; Philadelphia, PA USA; 2Department of Biological Sciences; University of Southern California; Los Angeles, CA USA; 3The Rockefeller University; Howard Hughes Medical Institute; New York, NY USA enetic studies have suggested that occur on both episomal and chromosomal GY-family translesion DNA poly- DNA and promote the survival of labora- merase IV (DinB) performs error-prone tory strains and natural isolates of bacteria distribute. recombination-directed replication under conditions of stress.6,7 Importantly, (RDR) under conditions of stress due stress-induced mutagenesis has many par- to its ability to promote mutations dur- allels to the adaptation of cancer cells to not ing double-strand break (DSB) repair growth-limiting conditions and stress and in growth-limited E. coli cells. In recent therefore may have implications for cancer Do studies we have demonstrated that pol progression and the ability of tumor cells IV is preferentially recruited to D-loop to develop resistance to therapy.3 recombination intermediates at stress- The central mechanism of adaptive induced concentrations and is highly mutagenesis in E. coli has been revealed mutagenic during RDR in vitro. These by several years of elegant genetic studies findings verify longstanding genetic data which have identified Y-family translesion that have implicated pol IV in promoting DNA polymerase IV (pol IV), also known stress-induced mutagenesis at D-loops. as DinB, as the major driver of stress- 1,2,8 In this Extra View, we demonstrate the induced mutations. Pol IV is highly Bioscience. surprising finding that A-family pol I, upregulated to ~2500 molecules/cell which normally exhibits high-fidelity during the SOS response and is further DNA synthesis, is highly error-prone at induced by an additional ~2-fold to ~5000 Keywords: DNA replication, D-loops like pol IV. These findings indi- molecules/cell during stationary phase by homologous recombination, double- cate that DNA polymerases are intrin- the RpoS general stress response.8,9 This strand break repair, stress-induced sically error-prone at RecA-mediated makes pol IV by far the most abundant Landes mutagenesis, adaptive mutagen- D-loops and suggest that auxiliary DNA polymerase in growth-limited cells. esis, break-induced replication factors are necessary for suppressing Considering that pol IV is an error-prone mutations during RDR in non-stressed Y-family DNA polymerase, it is there- Abbreviations: HR, homologous recom- proliferating cells. fore not surprising that it promotes most bination; RDR, recombination directed (~85%) adaptive mutations under growth- replication; DSB, double-strand break; Introduction limiting conditions8. ©2013 pol, DNA polymerase; D-loop, displace- Where pol IV elicits its mutagenic ment loop; BIR, break-induced replica- Adaptive mutagenesis, also known as activity during stress has also been revealed tion; ALT, alternative lengthening of stress-induced mutagenesis, is a major by genetic studies. For example, pol IV- telomeres evolutionary process that allows bacteria induced adaptive mutations have been Submitted: 06/24/2013 to overcome adverse environmental condi- shown to require DSB repair proteins such tions, such as nutrient starvation and expo- as RecA, RecBCD and RuvABC, and such Accepted: 07/09/2013 sure to antibiotics.1-3 For example, adaptive mutations are mostly targeted to regions http://dx.doi.org/10.4161/cc.25691 mutagenesis has been shown to promote near DSBs.10 Hence, previous genetics antibiotic resistance, which is becoming a indicate that pol IV promotes adaptive *Correspondence to: Richard T Pomerantz; major threat to modern medicine.4,5 It is mutations at or near displacement loop Email: [email protected] now evident that stress-induced mutations (D-loop) recombination intermediates, 2558 Cell Cycle Volume 12 Issue 16 PAPER TYPE EXTRA VIEW which are necessary for DSB repair. A requires upregulation of the polymerase exposure to antibiotics.18 This activity of general model of pol IV involvement by the SOS response and induction of the pol IV appears to potentiate cell death in stress-induced mutagenesis during RpoS stress response, which occurs in sta- during antibiotic treatment, presumably DSB repair is illustrated in Figure 1. tionary-phase cells (Fig. 1).9,12 Altogether, due to a high frequency of DSBs caused DSBs are first processed by the RecBCD genetic studies have provided compelling by incomplete repair of closely spaced nuclease/helicase complex that resects the evidence that pol IV mutagenic RDR 8-oxo-dGMP lesions.18 Considering that DNA and promotes RecA filament forma- activity in growth-limited cells is the the RpoS response upregulates pol IV by tion along the resulting 3′ single-strand central mechanism of adaptive mutagen- ~100% and promotes resistance to oxi- DNA tails. RecA filaments then promote esis. Considering bacteria spend extended dation, the ability of pol IV to incorpo- strand invasion within a homologous periods in growth-limiting conditions in rate 8-oxo-dGTP may also be beneficial donor duplex, resulting in a D-loop. Pol IV nature, pol IV involvement in RDR is for survival.19,20 Although the mode of is then recruited to the D-loop, presum- likely a major activity for this enzyme. replication in which pol IV incorporates ably along with replication co-factors, and Although pol IV is strongly implicated 8-oxo-dGTP has yet to be elucidated, the copies the sequence information from the in RDR, it is important to point out that suggested high frequency of 8-oxo-dGTP undamaged DNA donor molecule, result- Y-family DNA polymerases are typically incorporation during antibiotic treatment ing in mutations in close proximity to known for their ability to promote replica- indicates that pol IV is actively engaged DSBs. This recombination-directed repli- tion past DNA lesions in a pathway called in replication during stress, which may cation (RDR) process is often referred to translesion synthesis (TLS).13,14 Since be accounted for by pol IV RDR activ- as D-loop extension or break-induced rep- high-fidelity replicative DNA polymerases ity.18 Taken together, pol IV was probably distribute. lication (BIR), which is also error-prone stall upon encountering lesions in DNA selected to perform multiple functions in in yeast.11 Pol IV involvement in error- due to their stringent active site struc- DNA damage tolerance in stressed cells, prone RDR (stress-induced mutagenesis) tures, replication forks become arrested including RDR, TLS, and 8-oxo-dGMP not or even collapse upon collision with DNA incorporation. lesions in the leading strand; lesions in the Do lagging strand do not arrest replication Pol IV Preferentially Extends forks. In contrast to high-fidelity DNA D-Loops during Stress polymerases, low-fidelity Y-family DNA polymerases possess relatively open active Although previous genetics data have sites, which allows them to accommodate strongly implicated pol IV in RDR in aberrant DNA structures due to damage stressed cells, direct biochemical evidence such as cyclobutane pyrimidine dimmers, for this activity has been lacking. In recent which are frequently caused by exposure studies, we directly examined the ability 13-16 to UV light. Other common DNA of pol IV to promote RDR by reconsti- Bioscience. lesions bypassed by translesion DNA poly- tuting D-loop extension in vitro.21 These merases include abasic sites and 8-oxo- studies show that pol IV is proficient in 7,8-dihydroguanine (8-oxoG), which RDR, whereas the related Y-family pol V occur regularly in normal unstressed cel- is unable to perform this activity under lular environments as result of hydrolysis identical conditions. Pol IV is there- and oxidation, respectively. fore readily recruited to RecA mediated Landes Interestingly, mounting evidence indi- D-loops and exhibits robust strand dis- cates that although Y-family DNA polymer- placement activity on supercoiled DNA. ases are mostly error-prone and inefficient Since pol IV is highly upregulated during on undamaged DNA, they are relatively the SOS response, we examined whether accurate and proficient when bypassing the β clamp, which confers processivity their respective lesions.14 For example, onto DNA polymerases, is needed for pol ©2013 pol IV is more proficient and accurate in IV RDR activity. Using concentrations of replication opposite deoxyguanosine-con- pol IV relative to those in SOS-induced Figure 1. Model of pol IV activity during error- taining adducts at the N2-position (i.e., cells, we found that the polymerase does prone recombination. DSBs are processed N2-furfuryl-dG) compared with undam- not require β for RDR.21 Hence, the by the RecBCD helicase-nuclease complex, 17 which resects the DNA and facilitates loading aged nucleotides. Pol κ, the mammalian abundant levels of pol IV in stressed cells of RecA onto ssDNA. The RecA filament pro- ortholog of pol IV, is also more efficient in may preclude the need for β. These recent motes strand invasion within a homologous replication of such adducts.17 findings verify the ability of pol IV to DNA donor resulting in D-loop formation. Intriguingly, pol IV has also been promote RDR, as suggested by previous Pol IV is preferentially
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