Journal articles: 'Lagging-strand DNA synthesis'

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Giannattasio, Michele, and Dana Branzei. "DNA Replication Through Strand Displacement During Lagging Strand DNA Synthesis in Saccharomyces cerevisiae." Genes 10, no. 2 (February 21, 2019): 167. http://dx.doi.org/10.3390/genes10020167.

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This review discusses a set of experimental results that support the existence of extended strand displacement events during budding yeast lagging strand DNA synthesis. Starting from introducing the mechanisms and factors involved in leading and lagging strand DNA synthesis and some aspects of the architecture of the eukaryotic replisome, we discuss studies on bacterial, bacteriophage and viral DNA polymerases with potent strand displacement activities. We describe proposed pathways of Okazaki fragment processing via short and long flaps, with a focus on experimental results obtained in Saccharomyces cerevisiae that suggest the existence of frequent and extended strand displacement events during eukaryotic lagging strand DNA synthesis, and comment on their implications for genome integrity.

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Hernandez, Alfredo J., Seung-Joo Lee, and Charles C. Richardson. "Primer release is the rate-limiting event in lagging-strand synthesis mediated by the T7 replisome." Proceedings of the National Academy of Sciences 113, no. 21 (May 9, 2016): 5916–21. http://dx.doi.org/10.1073/pnas.1604894113.

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DNA replication occurs semidiscontinuously due to the antiparallel DNA strands and polarity of enzymatic DNA synthesis. Although the leading strand is synthesized continuously, the lagging strand is synthesized in small segments designated Okazaki fragments. Lagging-strand synthesis is a complex event requiring repeated cycles of RNA primer synthesis, transfer to the lagging-strand polymerase, and extension effected by cooperation between DNA primase and the lagging-strand polymerase. We examined events controlling Okazaki fragment initiation using the bacteriophage T7 replication system. Primer utilization by T7 DNA polymerase is slower than primer formation. Slow primer release from DNA primase allows the polymerase to engage the complex and is followed by a slow primer handoff step. The T7 single-stranded DNA binding protein increases primer formation and extension efficiency but promotes limited rounds of primer extension. We present a model describing Okazaki fragment initiation, the regulation of fragment length, and their implications for coordinated leading- and lagging-strand DNA synthesis.

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Koussa, Natasha C., and Duncan J. Smith. "Limiting DNA polymerase delta alters replication dynamics and leads to a dependence on checkpoint activation and recombination-mediated DNA repair." PLOS Genetics 17, no. 1 (January 25, 2021): e1009322. http://dx.doi.org/10.1371/journal.pgen.1009322.

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DNA polymerase delta (Pol δ) plays several essential roles in eukaryotic DNA replication and repair. At the replication fork, Pol δ is responsible for the synthesis and processing of the lagging-strand. At replication origins, Pol δ has been proposed to initiate leading-strand synthesis by extending the first Okazaki fragment. Destabilizing mutations in human Pol δ subunits cause replication stress and syndromic immunodeficiency. Analogously, reduced levels of Pol δ in Saccharomyces cerevisiae lead to pervasive genome instability. Here, we analyze how the depletion of Pol δ impacts replication origin firing and lagging-strand synthesis during replication elongation in vivo in S. cerevisiae. By analyzing nascent lagging-strand products, we observe a genome-wide change in both the establishment and progression of replication. S-phase progression is slowed in Pol δ depletion, with both globally reduced origin firing and slower replication progression. We find that no polymerase other than Pol δ is capable of synthesizing a substantial amount of lagging-strand DNA, even when Pol δ is severely limiting. We also characterize the impact of impaired lagging-strand synthesis on genome integrity and find increased ssDNA and DNA damage when Pol δ is limiting; these defects lead to a strict dependence on checkpoint signaling and resection-mediated repair pathways for cellular viability.

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Lukac, David, Zuzana Machacova, and Pavel Moudry. "Emetine blocks DNA replication via proteosynthesis inhibition not by targeting Okazaki fragments." Life Science Alliance 5, no. 12 (September 9, 2022): e202201560. http://dx.doi.org/10.26508/lsa.202201560.

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DNA synthesis of the leading and lagging strands works independently and cells tolerate single-stranded DNA generated during strand uncoupling if it is protected by RPA molecules. Natural alkaloid emetine is used as a specific inhibitor of lagging strand synthesis, uncoupling leading and lagging strand replication. Here, by analysis of lagging strand synthesis inhibitors, we show that despite emetine completely inhibiting DNA replication: it does not induce the generation of single-stranded DNA and chromatin-bound RPA32 (CB-RPA32). In line with this, emetine does not activate the replication checkpoint nor DNA damage response. Emetine is also an inhibitor of proteosynthesis and ongoing proteosynthesis is essential for the accurate replication of DNA. Mechanistically, we demonstrate that the acute block of proteosynthesis by emetine temporally precedes its effects on DNA replication. Thus, our results are consistent with the hypothesis that emetine affects DNA replication by proteosynthesis inhibition. Emetine and mild POLA1 inhibition prevent S-phase poly(ADP-ribosyl)ation. Collectively, our study reveals that emetine is not a specific lagging strand synthesis inhibitor with implications for its use in molecular biology.

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Kramer, M. Gabriela, Saleem A. Khan, and Manuel Espinosa. "Lagging-Strand Replication from the ssoA Origin of Plasmid pMV158 in Streptococcus pneumoniae: In Vivo and In Vitro Influences of Mutations in Two ConservedssoA Regions." Journal of Bacteriology 180, no. 1 (January 1, 1998): 83–89. http://dx.doi.org/10.1128/jb.180.1.83-89.1998.

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ABSTRACT The streptococcal plasmid pMV158 replicates by the rolling-circle mechanism. One feature of this replication mechanism is the generation of single-stranded DNA intermediates which are converted to double-stranded molecules. Lagging-strand synthesis initiates from the plasmid single-stranded origin, sso. We have used the pMV158-derivative plasmid pLS1 (containing the ssoA type of lagging-strand origin) and a set of pLS1 derivatives with mutations in two conserved regions of the ssoA (the recombination site B [RSB] and a conserved 6-nucleotide sequence [CS-6]) to identify sequences important for plasmid lagging-strand replication inStreptococcus pneumoniae. Cells containing plasmids with mutations in the RSB accumulated 30-fold more single-stranded DNA than cells containing plasmids with mutations in the CS-6 sequence. Specificity of lagging-strand synthesis was tested by the development of a new in vitro replication system with pneumococcal cell extracts. Four major initiation sites of lagging-strand DNA synthesis were observed. The specificity of initiation was maintained in plasmids with mutations in the CS-6 region. Mutations in the RSB region, on the other hand, resulted in the loss of specific initiation of lagging-strand synthesis and also severely reduced the efficiency of replication.

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Spiering, Michelle M., Philip Hanoian, Swathi Gannavaram, and Stephen J. Benkovic. "RNA primer–primase complexes serve as the signal for polymerase recycling and Okazaki fragment initiation in T4 phage DNA replication." Proceedings of the National Academy of Sciences 114, no. 22 (May 15, 2017): 5635–40. http://dx.doi.org/10.1073/pnas.1620459114.

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The opposite strand polarity of duplex DNA necessitates that the leading strand is replicated continuously whereas the lagging strand is replicated in discrete segments known as Okazaki fragments. The lagging-strand polymerase sometimes recycles to begin the synthesis of a new Okazaki fragment before finishing the previous fragment, creating a gap between the Okazaki fragments. The mechanism and signal that initiate this behavior—that is, the signaling mechanism—have not been definitively identified. We examined the role of RNA primer–primase complexes left on the lagging ssDNA from primer synthesis in initiating early lagging-strand polymerase recycling. We show for the T4 bacteriophage DNA replication system that primer–primase complexes have a residence time similar to the timescale of Okazaki fragment synthesis and the ability to block a holoenzyme synthesizing DNA and stimulate the dissociation of the holoenzyme to trigger polymerase recycling. The collision with primer–primase complexes triggering the early termination of Okazaki fragment synthesis has distinct advantages over those previously proposed because this signal requires no transmission to the lagging-strand polymerase through protein or DNA interactions, the mechanism for rapid dissociation of the holoenzyme is always collision, and no unique characteristics need to be assigned to either identical polymerase in the replisome. We have modeled repeated cycles of Okazaki fragment initiation using a collision with a completed Okazaki fragment or primer–primase complexes as the recycling mechanism. The results reproduce experimental data, providing insights into events related to Okazaki fragment initiation and the overall functioning of DNA replisomes.

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Parenteau, Julie, and Raymund J. Wellinger. "Accumulation of Single-Stranded DNA and Destabilization of Telomeric Repeats in Yeast Mutant Strains Carrying a Deletion of RAD27." Molecular and Cellular Biology 19, no. 6 (June 1, 1999): 4143–52. http://dx.doi.org/10.1128/mcb.19.6.4143.

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ABSTRACT The Saccharomyces cerevisiae RAD27 gene encodes the yeast homologue of the mammalian FEN-1 nuclease, a protein that is thought to be involved in the processing of Okazaki fragments during DNA lagging-strand synthesis. One of the predicted DNA lesions occurring in rad27 strains is the presence of single-stranded DNA of the template strand for lagging-strand synthesis. We examined this prediction by analyzing the terminal DNA structures generated during telomere replication in rad27strains. The lengths of the telomeric repeat tracts were found to be destabilized in rad27 strains, indicating that naturally occurring direct repeats are subject to tract expansions and contractions in such strains. Furthermore, abnormally high levels of single-stranded DNA of the templating strand for lagging-strand synthesis were observed in rad27 cells. Overexpression of Dna2p in wild-type cells also yielded single-stranded DNA regions on telomeric DNA and caused a cell growth arrest phenotype virtually identical to that seen for rad27 cells grown at the restrictive temperature. Furthermore, overexpression of the yeast exonuclease Exo1p alleviated the growth arrest induced by both conditions, overexpression of Dna2p and incubation of rad27cells at 37°C. However, the telomere heterogeneity and the appearance of single-stranded DNA are not prevented by the overexpression of Exo1p in these strains, suggesting that this nuclease is not simply redundant with Rad27p. Our data thus provide in vivo evidence for the types of DNA lesions predicted to occur when lagging-strand synthesis is deficient and suggest that Dna2p and Rad27p collaborate in the processing of Okazaki fragments.

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Serra-Cardona, Albert, Chuanhe Yu, Xinmin Zhang, Xu Hua, Yuan Yao, Jiaqi Zhou, Haiyun Gan, and Zhiguo Zhang. "A mechanism for Rad53 to couple leading- and lagging-strand DNA synthesis under replication stress in budding yeast." Proceedings of the National Academy of Sciences 118, no. 38 (September 16, 2021): e2109334118. http://dx.doi.org/10.1073/pnas.2109334118.

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In response to DNA replication stress, DNA replication checkpoint kinase Mec1 phosphorylates Mrc1, which in turn activates Rad53 to prevent the generation of deleterious single-stranded DNA, a process that remains poorly understood. We previously reported that lagging-strand DNA synthesis proceeds farther than leading strand in rad53-1 mutant cells defective in replication checkpoint under replication stress, resulting in the exposure of long stretches of the leading-strand templates. Here, we show that asymmetric DNA synthesis is also observed in mec1-100 and mrc1-AQ cells defective in replication checkpoint but, surprisingly, not in mrc1∆ cells in which both DNA replication and checkpoint functions of Mrc1 are missing. Furthermore, depletion of either Mrc1 or its partner, Tof1, suppresses the asymmetric DNA synthesis in rad53-1 mutant cells. Thus, the DNA replication checkpoint pathway couples leading- and lagging-strand DNA synthesis by attenuating the replication function of Mrc1-Tof1 under replication stress.

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Sparks, Melanie A., Peter M. Burgers, and Roberto Galletto. "Pif1, RPA, and FEN1 modulate the ability of DNA polymerase δ to overcome protein barriers during DNA synthesis." Journal of Biological Chemistry 295, no. 47 (September 10, 2020): 15883–91. http://dx.doi.org/10.1074/jbc.ra120.015699.

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Successful DNA replication requires carefully regulated mechanisms to overcome numerous obstacles that naturally occur throughout chromosomal DNA. Scattered across the genome are tightly bound proteins, such as transcription factors and nucleosomes, that are necessary for cell function, but that also have the potential to impede timely DNA replication. Using biochemically reconstituted systems, we show that two transcription factors, yeast Reb1 and Tbf1, and a tightly positioned nucleosome, are strong blocks to the strand displacement DNA synthesis activity of DNA polymerase δ. Although the block imparted by Tbf1 can be overcome by the DNA-binding activity of the single-stranded DNA-binding protein RPA, efficient DNA replication through either a Reb1 or a nucleosome block occurs only in the presence of the 5'-3' DNA helicase Pif1. The Pif1-dependent stimulation of DNA synthesis across strong protein barriers may be beneficial during break-induced replication where barriers are expected to pose a problem to efficient DNA bubble migration. However, in the context of lagging strand DNA synthesis, the efficient disruption of a nucleosome barrier by Pif1 could lead to the futile re-replication of newly synthetized DNA. In the presence of FEN1 endonuclease, the major driver of nick translation during lagging strand replication, Pif1-dependent stimulation of DNA synthesis through a nucleosome or Reb1 barrier is prevented. By cleaving the short 5' tails generated during strand displacement, FEN1 eliminates the entry point for Pif1. We propose that this activity would protect the cell from potential DNA re-replication caused by unwarranted Pif1 interference during lagging strand replication.

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Nasheuer, Heinz Peter, and Nichodemus O. Onwubiko. "Lagging Strand Initiation Processes in DNA Replication of Eukaryotes—Strings of Highly Coordinated Reactions Governed by Multiprotein Complexes." Genes 14, no. 5 (April 29, 2023): 1012. http://dx.doi.org/10.3390/genes14051012.

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In their influential reviews, Hanahan and Weinberg coined the term ‘Hallmarks of Cancer’ and described genome instability as a property of cells enabling cancer development. Accurate DNA replication of genomes is central to diminishing genome instability. Here, the understanding of the initiation of DNA synthesis in origins of DNA replication to start leading strand synthesis and the initiation of Okazaki fragment on the lagging strand are crucial to control genome instability. Recent findings have provided new insights into the mechanism of the remodelling of the prime initiation enzyme, DNA polymerase α-primase (Pol-prim), during primer synthesis, how the enzyme complex achieves lagging strand synthesis, and how it is linked to replication forks to achieve optimal initiation of Okazaki fragments. Moreover, the central roles of RNA primer synthesis by Pol-prim in multiple genome stability pathways such as replication fork restart and protection of DNA against degradation by exonucleases during double-strand break repair are discussed.

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11

Liu, Guoqi, Xiaomi Chen, and Michael Leffak. "Oligodeoxynucleotide Binding to (CTG) · (CAG) Microsatellite Repeats Inhibits Replication Fork Stalling, Hairpin Formation, and Genome Instability." Molecular and Cellular Biology 33, no. 3 (November 19, 2012): 571–81. http://dx.doi.org/10.1128/mcb.01265-12.

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ABSTRACT(CTG)n· (CAG)ntrinucleotide repeat (TNR) expansion in the 3′ untranslated region of the dystrophia myotonica protein kinase (DMPK) gene causes myotonic dystrophy type 1. However, a direct link between TNR instability, the formation of noncanonical (CTG)n· (CAG)nstructures, and replication stress has not been demonstrated. In a human cell model, we found that (CTG)45· (CAG)45causes local replication fork stalling, DNA hairpin formation, and TNR instability. Oligodeoxynucleotides (ODNs) complementary to the (CTG)45· (CAG)45lagging-strand template eliminated DNA hairpin formation on leading- and lagging-strand templates and relieved fork stalling. Prolonged cell culture, emetine inhibition of lagging-strand synthesis, or slowing of DNA synthesis by low-dose aphidicolin induced (CTG)45· (CAG)45expansions and contractions. ODNs targeting the lagging-strand template blocked the time-dependent or emetine-induced instability but did not eliminate aphidicolin-induced instability. These results show directly that TNR replication stalling, replication stress, hairpin formation, and instability are mechanistically linkedin vivo.

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Kulczyk, Arkadiusz W., Arne Moeller, Peter Meyer, Piotr Sliz, and Charles C. Richardson. "Cryo-EM structure of the replisome reveals multiple interactions coordinating DNA synthesis." Proceedings of the National Academy of Sciences 114, no. 10 (February 21, 2017): E1848—E1856. http://dx.doi.org/10.1073/pnas.1701252114.

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We present a structure of the ∼650-kDa functional replisome of bacteriophage T7 assembled on DNA resembling a replication fork. A structure of the complex consisting of six domains of DNA helicase, five domains of RNA primase, two DNA polymerases, and two thioredoxin (processivity factor) molecules was determined by single-particle cryo-electron microscopy. The two molecules of DNA polymerase adopt a different spatial arrangement at the replication fork, reflecting their roles in leading- and lagging-strand synthesis. The structure, in combination with biochemical data, reveals molecular mechanisms for coordination of leading- and lagging-strand synthesis. Because mechanisms of DNA replication are highly conserved, the observations are relevant to other replication systems.

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Gao, Yang, Yanxiang Cui, Tara Fox, Shiqiang Lin, Huaibin Wang, Natalia de Val, Z. Hong Zhou, and Wei Yang. "Structures and operating principles of the replisome." Science 363, no. 6429 (January 24, 2019): eaav7003. http://dx.doi.org/10.1126/science.aav7003.

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Visualization in atomic detail of the replisome that performs concerted leading– and lagging–DNA strand synthesis at a replication fork has not been reported. Using bacteriophage T7 as a model system, we determined cryo–electron microscopy structures up to 3.2-angstroms resolution of helicase translocating along DNA and of helicase-polymerase-primase complexes engaging in synthesis of both DNA strands. Each domain of the spiral-shaped hexameric helicase translocates sequentially hand-over-hand along a single-stranded DNA coil, akin to the way AAA+ ATPases (adenosine triphosphatases) unfold peptides. Two lagging-strand polymerases are attached to the primase, ready for Okazaki fragment synthesis in tandem. A β hairpin from the leading-strand polymerase separates two parental DNA strands into a T-shaped fork, thus enabling the closely coupled helicase to advance perpendicular to the downstream DNA duplex. These structures reveal the molecular organization and operating principles of a replisome.

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Hiasa, H., and K. J. Marians. "Primase couples leading- and lagging-strand DNA synthesis from oriC." Journal of Biological Chemistry 269, no. 8 (February 1994): 6058–63. http://dx.doi.org/10.1016/s0021-9258(17)37569-5.

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Holt, Ian J., Heather E. Lorimer, and Howard T. Jacobs. "Coupled Leading- and Lagging-Strand Synthesis of Mammalian Mitochondrial DNA." Cell 100, no. 5 (March 2000): 515–24. http://dx.doi.org/10.1016/s0092-8674(00)80688-1.

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Kreisel, Katrin, Martin K. M. Engqvist, Josephine Kalm, Liam J. Thompson, Martin Boström, Clara Navarrete, John P. McDonald, Erik Larsson, Roger Woodgate, and Anders R. Clausen. "DNA polymerase η contributes to genome-wide lagging strand synthesis." Nucleic Acids Research 47, no. 5 (December 28, 2018): 2425–35. http://dx.doi.org/10.1093/nar/gky1291.

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Spiering, Michelle M., Scott W. Nelson, and Stephen J. Benkovic. "Repetitive lagging strand DNA synthesis by the bacteriophage T4 replisome." Molecular BioSystems 4, no. 11 (2008): 1070. http://dx.doi.org/10.1039/b812163j.

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Onwubiko, Nichodemus O., Angela Borst, Suraya A. Diaz, Katharina Passkowski, Felicia Scheffel, Ingrid Tessmer, and Heinz P. Nasheuer. "SV40 T antigen interactions with ssDNA and replication protein A: a regulatory role of T antigen monomers in lagging strand DNA replication." Nucleic Acids Research 48, no. 7 (March 4, 2020): 3657–77. http://dx.doi.org/10.1093/nar/gkaa138.

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Abstract DNA replication is a central process in all living organisms. Polyomavirus DNA replication serves as a model system for eukaryotic DNA replication and has considerably contributed to our understanding of basic replication mechanisms. However, the details of the involved processes are still unclear, in particular regarding lagging strand synthesis. To delineate the complex mechanism of coordination of various cellular proteins binding simultaneously or consecutively to DNA to initiate replication, we investigated single-stranded DNA (ssDNA) interactions by the SV40 large T antigen (Tag). Using single molecule imaging by atomic force microscopy (AFM) combined with biochemical and spectroscopic analyses we reveal independent activity of monomeric and oligomeric Tag in high affinity binding to ssDNA. Depending on ssDNA length, we obtain dissociation constants for Tag-ssDNA interactions (KD values of 10–30 nM) that are in the same order of magnitude as ssDNA binding by human replication protein A (RPA). Furthermore, we observe the formation of RPA-Tag-ssDNA complexes containing hexameric as well as monomeric Tag forms. Importantly, our data clearly show stimulation of primase function in lagging strand Okazaki fragment synthesis by monomeric Tag whereas hexameric Tag inhibits the reaction, redefining DNA replication initiation on the lagging strand.

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Lee, Joonsoo, Paul D. Chastain, Jack D. Griffith, and Charles C. Richardson. "Lagging strand synthesis in coordinated DNA synthesis by bacteriophage T7 replication proteins." Journal of Molecular Biology 316, no. 1 (February 2002): 19–34. http://dx.doi.org/10.1006/jmbi.2001.5325.

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Yeeles, Joseph T. P. "Discontinuous leading-strand synthesis: a stop–start story." Biochemical Society Transactions 42, no. 1 (January 23, 2014): 25–34. http://dx.doi.org/10.1042/bst20130262.

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Reconstitution experiments using replication proteins from a number of different model organisms have firmly established that, in vitro, DNA replication is semi-discontinuous: continuous on the leading strand and discontinuous on the lagging strand. The mechanism by which DNA is replicated in vivo is less clear. In fact, there have been many observations of discontinuous replication in the absence of exogenous DNA-damaging agents. It has also been proposed that replication is discontinuous on the leading strand at least in part because of DNA lesion bypass. Several recent studies have revealed mechanistic details of pathways where replication of the leading strand introduces discontinuities. These mechanisms and their potential contributions to observations of discontinuous replication in vivo will be discussed.

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Ohki, Rieko, Toshiki Tsurimoto, and Fuyuki Ishikawa. "In Vitro Reconstitution of the End Replication Problem." Molecular and Cellular Biology 21, no. 17 (September 1, 2001): 5753–66. http://dx.doi.org/10.1128/mcb.21.17.5753-5766.2001.

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ABSTRACT The end replication problem hypothesis proposes that the ends of linear DNA cannot be replicated completely during lagging strand DNA synthesis. Although the idea has been widely accepted for explaining telomere attrition during cell proliferation, it has never been directly demonstrated. In order to take a biochemical approach to understand how linear DNA ends are replicated, we have established a novel in vitro linear simian virus 40 DNA replication system. In this system, terminally biotin-labeled linear DNAs are conjugated to avidin-coated beads and subjected to replication reactions. Linear DNA was efficiently replicated under optimized conditions, and replication products that had replicated using the original DNA templates were specifically analyzed by purifying bead-bound replication products. By exploiting this system, we showed that while the leading strand is completely synthesized to the end, lagging strand synthesis is gradually halted in the terminal ∼500-bp region, leaving 3′ overhangs. This result is consistent with observations in telomerase-negative mammalian cells and formally demonstrates the end replication problem. This study provides a basis for studying the details of telomere replication.

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Wanrooij, S., J. M. Fuste, G. Farge, Y. Shi, C. M. Gustafsson, and M. Falkenberg. "Human mitochondrial RNA polymerase primes lagging-strand DNA synthesis in vitro." Proceedings of the National Academy of Sciences 105, no. 32 (August 6, 2008): 11122–27. http://dx.doi.org/10.1073/pnas.0805399105.

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Lee, Joonsoo, Paul D. Chastain, Takahiro Kusakabe, Jack D. Griffith, and Charles C. Richardson. "Coordinated Leading and Lagging Strand DNA Synthesis on a Minicircular Template." Molecular Cell 1, no. 7 (June 1998): 1001–10. http://dx.doi.org/10.1016/s1097-2765(00)80100-8.

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Delagoutte, Emmanuelle, and Giuseppe Baldacci. "5′CAG and 5′CTG Repeats Create Differential Impediment to the Progression of a Minimal Reconstituted T4 Replisome Depending on the Concentration of dNTPs." Molecular Biology International 2011 (August 10, 2011): 1–14. http://dx.doi.org/10.4061/2011/213824.

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Instability of repetitive sequences originates from strand misalignment during repair or replicative DNA synthesis. To investigate the activity of reconstituted T4 replisomes across trinucleotide repeats (TNRs) during leading strand DNA synthesis, we developed a method to build replication miniforks containing a TNR unit of defined sequence and length. Each minifork consists of three strands, primer, leading strand template, and lagging strand template with a 5′ single-stranded (ss) tail. Each strand is prepared independently, and the minifork is assembled by hybridization of the three strands. Using these miniforks and a minimal reconstituted T4 replisome, we show that during leading strand DNA synthesis, the dNTP concentration dictates which strand of the structure-forming 5′CAG/5′CTG repeat creates the strongest impediment to the minimal replication complex. We discuss this result in the light of the known fluctuation of dNTP concentration during the cell cycle and cell growth and the known concentration balance among individual dNTPs.

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Kadyrov, Farid A., and John W. Drake. "Conditional Coupling of Leading-strand and Lagging-strand DNA Synthesis at Bacteriophage T4 Replication Forks." Journal of Biological Chemistry 276, no. 31 (June 4, 2001): 29559–66. http://dx.doi.org/10.1074/jbc.m101310200.

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Weston-Hafer, K., and D. E. Berg. "Deletions in plasmid pBR322: replication slippage involving leading and lagging strands." Genetics 127, no. 4 (April 1, 1991): 649–55. http://dx.doi.org/10.1093/genetics/127.4.649.

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Abstract We test here whether a class of deletions likely to result from errors during DNA replication arise preferentially during synthesis of either the leading or the lagging DNA strand. Deletions were obtained by reversion of particular insertion mutant alleles of the pBR322 amp gene. The alleles contain insertions of palindromic DNAs bracketed by 9-bp direct repeats of amp sequence; in addition, bp 2 to 5 in one arm of the palindrome form a direct repeat with 4 bp of adjoining amp sequence. Prior work had shown that reversion to Ampr results from deletions with endpoints in the 8- or 4-bp repeat, and that the 4-bp repeats are used preferentially because one of them is in the palindrome. To test the role of leading and lagging strand synthesis in deletion formation, we reversed the direction of replication of the amp gene by inverting the pBR322 replication origin, and also constructed new mutant alleles with a 4-bp repeat starting counterclockwise rather than clockwise of the insertion. In both cases the 4-bp repeats were used preferentially as deletion endpoints. A model is presented in which deletions arise during elongation of the strand that copies the palindrome before the adjoining 4-bp repeat, and in which preferential use of the 4-bp repeats independent of the overall direction of replication implies that deletions arise during syntheses of both leading and lagging strands.

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Khristich, Alexandra N., Jillian F. Armenia, Robert M. Matera, Anna A. Kolchinski, and Sergei M. Mirkin. "Large-scale contractions of Friedreich’s ataxia GAA repeats in yeast occur during DNA replication due to their triplex-forming ability." Proceedings of the National Academy of Sciences 117, no. 3 (January 7, 2020): 1628–37. http://dx.doi.org/10.1073/pnas.1913416117.

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Friedreich’s ataxia (FRDA) is a human hereditary disease caused by the presence of expanded (GAA)n repeats in the first intron of the FXN gene [V. Campuzano et al., Science 271, 1423–1427 (1996)]. In somatic tissues of FRDA patients, (GAA)n repeat tracts are highly unstable, with contractions more common than expansions [R. Sharma et al., Hum. Mol. Genet. 11, 2175–2187 (2002)]. Here we describe an experimental system to characterize GAA repeat contractions in yeast and to conduct a genetic analysis of this process. We found that large-scale contraction is a one-step process, resulting in a median loss of ∼60 triplet repeats. Our genetic analysis revealed that contractions occur during DNA replication, rather than by various DNA repair pathways. Repeats contract in the course of lagging-strand synthesis: The processivity subunit of DNA polymerase δ, Pol32, and the catalytic domain of Rev1, a translesion polymerase, act together in the same pathway to counteract contractions. Accumulation of single-stranded DNA (ssDNA) in the lagging-strand template greatly increases the probability that (GAA)n repeats contract, which in turn promotes repeat instability in rfa1, rad27, and dna2 mutants. Finally, by comparing contraction rates for homopurine-homopyrimidine repeats differing in their mirror symmetry, we found that contractions depend on a repeat’s triplex-forming ability. We propose that accumulation of ssDNA in the lagging-strand template fosters the formation of a triplex between the nascent and fold-back template strands of the repeat. Occasional jumps of DNA polymerase through this triplex hurdle, result in repeat contractions in the nascent lagging strand.

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Hamdan, Samir M., Joseph J. Loparo, Masateru Takahashi, Charles C. Richardson, and Antoine M. van Oijen. "Dynamics of DNA replication loops reveal temporal control of lagging-strand synthesis." Nature 457, no. 7227 (November 23, 2008): 336–39. http://dx.doi.org/10.1038/nature07512.

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Hamdan, Samir M., Joseph J. Loparo, Masateru Takahashi, Charles C. Richardson, and Antoine M. Vanoijen. "Dynamics Of DNA Replication Loops Reveal Temporal Control Of Lagging-strand Synthesis." Biophysical Journal 96, no. 3 (February 2009): 568a. http://dx.doi.org/10.1016/j.bpj.2008.12.3720.

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Cerron, Fernando, Grzegorz L. Ciesielski, Laurie S. Kaguni, Francisco J. Cao, and Borja Ibarra. "Mechanism of SSB Displacement by Replicative DNA Polymerases During Lagging Strand Synthesis." Biophysical Journal 116, no. 3 (February 2019): 74a. http://dx.doi.org/10.1016/j.bpj.2018.11.443.

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Hedglin, Mark, Binod Pandey, and Stephen J. Benkovic. "Stability of the human polymerase δ holoenzyme and its implications in lagging strand DNA synthesis." Proceedings of the National Academy of Sciences 113, no. 13 (March 14, 2016): E1777—E1786. http://dx.doi.org/10.1073/pnas.1523653113.

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In eukaryotes, DNA polymerase δ (pol δ) is responsible for replicating the lagging strand template and anchors to the proliferating cell nuclear antigen (PCNA) sliding clamp to form a holoenzyme. The stability of this complex is integral to every aspect of lagging strand replication. Most of our understanding comes fromSaccharomyces cerevisaewhere the extreme stability of the pol δ holoenzyme ensures that every nucleobase within an Okazaki fragment is faithfully duplicated before dissociation but also necessitates an active displacement mechanism for polymerase recycling and exchange. However, the stability of the human pol δ holoenzyme is unknown. We designed unique kinetic assays to analyze the processivity and stability of the pol δ holoenzyme. Surprisingly, the results indicate that human pol δ maintains a loose association with PCNA while replicating DNA. Such behavior has profound implications on Okazaki fragment synthesis in humans as it limits the processivity of pol δ on undamaged DNA and promotes the rapid dissociation of pol δ from PCNA on stalling at a DNA lesion.

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32

Yonesaki, T. "Involvement of a replicative DNA helicase of bacteriophage T4 in DNA recombination." Genetics 138, no. 2 (October 1, 1994): 247–52. http://dx.doi.org/10.1093/genetics/138.2.247.

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Abstract Bacteriophage T4 gene 41 encodes a replicative DNA helicase that is a subunit of the primosome which is essential for lagging-strand DNA synthesis. A mutation, rrh, was generated and selected in the helicase gene on the basis of limited DNA replication that ceases early. The survival of ultraviolet-irradiated phage and the frequency of genetic recombination are reduced by rrh. In addition, rrh diminishes the production of concatemeric DNA. These results strongly suggest that the gene 41 replicative helicase participates in DNA recombination.

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33

Nakamura, Mirai, Akira Nabetani, Takeshi Mizuno, Fumio Hanaoka, and Fuyuki Ishikawa. "Alterations of DNA and Chromatin Structures at Telomeres and Genetic Instability in Mouse Cells Defective in DNA Polymerase α." Molecular and Cellular Biology 25, no. 24 (December 15, 2005): 11073–88. http://dx.doi.org/10.1128/mcb.25.24.11073-11088.2005.

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ABSTRACT Telomere length is controlled by a homeostatic mechanism that involves telomerase, telomere-associated proteins, and conventional replication machinery. Specifically, the coordinated actions of the lagging strand synthesis and telomerase have been argued. Although DNA polymerase α, an enzyme important for the lagging strand synthesis, has been indicated to function in telomere metabolism in yeasts and ciliates, it has not been characterized in higher eukaryotes. Here, we investigated the impact of compromised polymerase α activity on telomeres, using tsFT20 mouse mutant cells harboring a temperature-sensitive polymerase α mutant allele. When polymerase α was temperature-inducibly inactivated, we observed sequential events that included an initial extension of the G-tail followed by a marked increase in the overall telomere length occurring in telomerase-independent and -dependent manners, respectively. These alterations of telomeric DNA were accompanied by alterations of telomeric chromatin structures as revealed by quantitative chromatin immunoprecipitation and immunofluorescence analyses of TRF1 and POT1. Unexpectedly, polymerase α inhibition resulted in a significantly high incidence of Robertsonian chromosome fusions without noticeable increases in other types of chromosomal aberrations. These results indicate that although DNA polymerase α is essential for genome-wide DNA replication, hypomorphic activity leads to a rather specific spectrum of chromosomal abnormality.

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34

Bullock, P. A., S. Tevosian, C. Jones, and D. Denis. "Mapping initiation sites for simian virus 40 DNA synthesis events in vitro." Molecular and Cellular Biology 14, no. 8 (August 1994): 5043–55. http://dx.doi.org/10.1128/mcb.14.8.5043-5055.1994.

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Primer RNA-DNA, a small (approximately 30-nucleotide) RNA-DNA hybrid molecule, was identified in recent studies of simian virus 40 DNA synthesis in vitro. The available evidence indicates that primer RNA-DNA is the product of the polymerase alpha-primase complex. Primer RNA-DNA is formed exclusively on lagging-strand DNA templates; it is synthesized initially in the vicinity of the simian virus 40 origin and at later times at sites progressively distal to the origin. To further characterize initiation events, template sequences encoding the 5' ends of both primer RNA and primer DNA, formed during a 5-s pulse, have been determined. Analyses of these sequences demonstrate the existence of an initiation signal for lagging-strand synthesis. At any given position, the initiation signal is located within those template sequences encoding primer RNA, situated proximal to the nucleotide encoding the 5' end of the RNA primer. In most instances, the sequence 5'-TTN-3' (where N encodes the nucleotide at the 5' end of the primer) is a feature of the initiation signal. Initiation signals are present, on average, once every 19 nucleotides. These results are discussed in terms of the mechanism of Okazaki fragment formation and possible links between prokaryotic and eukaryotic initiation events.

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35

Bullock, P. A., S. Tevosian, C. Jones, and D. Denis. "Mapping initiation sites for simian virus 40 DNA synthesis events in vitro." Molecular and Cellular Biology 14, no. 8 (August 1994): 5043–55. http://dx.doi.org/10.1128/mcb.14.8.5043.

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Primer RNA-DNA, a small (approximately 30-nucleotide) RNA-DNA hybrid molecule, was identified in recent studies of simian virus 40 DNA synthesis in vitro. The available evidence indicates that primer RNA-DNA is the product of the polymerase alpha-primase complex. Primer RNA-DNA is formed exclusively on lagging-strand DNA templates; it is synthesized initially in the vicinity of the simian virus 40 origin and at later times at sites progressively distal to the origin. To further characterize initiation events, template sequences encoding the 5' ends of both primer RNA and primer DNA, formed during a 5-s pulse, have been determined. Analyses of these sequences demonstrate the existence of an initiation signal for lagging-strand synthesis. At any given position, the initiation signal is located within those template sequences encoding primer RNA, situated proximal to the nucleotide encoding the 5' end of the RNA primer. In most instances, the sequence 5'-TTN-3' (where N encodes the nucleotide at the 5' end of the primer) is a feature of the initiation signal. Initiation signals are present, on average, once every 19 nucleotides. These results are discussed in terms of the mechanism of Okazaki fragment formation and possible links between prokaryotic and eukaryotic initiation events.

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36

Gawel, Damian, Magdalena Maliszewska-Tkaczyk, Piotr Jonczyk, Roel M. Schaaper, and Iwona J. Fijalkowska. "Lack of Strand Bias in UV-Induced Mutagenesis in Escherichia coli." Journal of Bacteriology 184, no. 16 (August 15, 2002): 4449–54. http://dx.doi.org/10.1128/jb.184.16.4449-4454.2002.

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ABSTRACT We have investigated whether UV-induced mutations are created with equal efficiency on the leading and lagging strands of DNA replication. We employed an assay system that permits measurement of mutagenesis in the lacZ gene in pairs of near-identical strains. Within each pair, the strains differ only in the orientation of the lacZ gene with respect to the origin of DNA replication. Depending on this orientation, any lacZ target sequence will be replicated in one orientation as a leading strand and as a lagging strand in the other orientation. In contrast to previous results obtained for mutations resulting from spontaneous replication errors or mutations resulting from the spontaneous SOS mutator effect, measurements of UV-induced mutagenesis in uvrA strains fail to show significant differences between the two target orientations. These data suggest that SOS-mediated mutagenic translesion synthesis on the Escherichia coli chromosome may occur with equal or similar probability on leading and lagging strands.

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37

Zhu, Yali, Zetang Wu, M. Cristina Cardoso, and Deborah S. Parris. "Processing of Lagging-Strand Intermediates In Vitro by Herpes Simplex Virus Type 1 DNA Polymerase." Journal of Virology 84, no. 15 (May 5, 2010): 7459–72. http://dx.doi.org/10.1128/jvi.01875-09.

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ABSTRACT The processing of lagging-strand intermediates has not been demonstrated in vitro for herpes simplex virus type 1 (HSV-1). Human flap endonuclease-1 (Fen-1) was examined for its ability to produce ligatable products with model lagging-strand intermediates in the presence of the wild-type or exonuclease-deficient (exo−) HSV-1 DNA polymerase (pol). Primer/templates were composed of a minicircle single-stranded DNA template annealed to primers that contained 5′ DNA flaps or 5′ annealed DNA or RNA sequences. Gapped DNA primer/templates were extended but not significantly strand displaced by the wild-type HSV-1 pol, although significant strand displacement was observed with exo− HSV-1 pol. Nevertheless, the incubation of primer/templates containing 5′ flaps with either wild-type or exo− HSV-1 pol and Fen-1 led to the efficient production of nicks that could be sealed with DNA ligase I. Both polymerases stimulated the nick translation activity of Fen-1 on DNA- or RNA-containing primer/templates, indicating that the activities were coordinated. Further evidence for Fen-1 involvement in HSV-1 DNA synthesis is suggested by the ability of a transiently expressed green fluorescent protein fusion with Fen-1 to accumulate in viral DNA replication compartments in infected cells and by the ability of endogenous Fen-1 to coimmunoprecipitate with an essential viral DNA replication protein in HSV-1-infected cells.

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38

Khan, S. A. "Rolling-circle replication of bacterial plasmids." Microbiology and Molecular Biology Reviews 61, no. 4 (December 1997): 442–55. http://dx.doi.org/10.1128/mmbr.61.4.442-455.1997.

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Many bacterial plasmids replicate by a rolling-circle (RC) mechanism. Their replication properties have many similarities to as well as significant differences from those of single-stranded DNA (ssDNA) coliphages, which also replicate by an RC mechanism. Studies on a large number of RC plasmids have revealed that they fall into several families based on homology in their initiator proteins and leading-strand origins. The leading-strand origins contain distinct sequences that are required for binding and nicking by the Rep proteins. Leading-strand origins also contain domains that are required for the initiation and termination of replication. RC plasmids generate ssDNA intermediates during replication, since their lagging-strand synthesis does not usually initiate until the leading strand has been almost fully synthesized. The leading- and lagging-strand origins are distinct, and the displaced leading-strand DNA is converted to the double-stranded form by using solely the host proteins. The Rep proteins encoded by RC plasmids contain specific domains that are involved in their origin binding and nicking activities. The replication and copy number of RC plasmids, in general, are regulated at the level of synthesis of their Rep proteins, which are usually rate limiting for replication. Some RC Rep proteins are known to be inactivated after supporting one round of replication. A number of in vitro replication systems have been developed for RC plasmids and have provided insight into the mechanism of plasmid RC replication.

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39

Kuban, Wojciech, Magdalena Banach-Orlowska, Malgorzata Bialoskorska, Aleksandra Lipowska, Roel M. Schaaper, Piotr Jonczyk, and Iwona J. Fijalkowska. "Mutator Phenotype Resulting from DNA Polymerase IV Overproduction in Escherichia coli: Preferential Mutagenesis on the Lagging Strand." Journal of Bacteriology 187, no. 19 (October 1, 2005): 6862–66. http://dx.doi.org/10.1128/jb.187.19.6862-6866.2005.

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ABSTRACT We investigated the mutator effect resulting from overproduction of Escherichia coli DNA polymerase IV. Using lac mutational targets in the two possible orientations on the chromosome, we observed preferential mutagenesis during lagging strand synthesis. The mutator activity likely results from extension of mismatches produced by polymerase III holoenzyme.

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40

Gan, Haiyun, Chuanhe Yu, Sujan Devbhandari, Sushma Sharma, Junhong Han, Andrei Chabes, Dirk Remus, and Zhiguo Zhang. "Checkpoint Kinase Rad53 Couples Leading- and Lagging-Strand DNA Synthesis under Replication Stress." Molecular Cell 68, no. 2 (October 2017): 446–55. http://dx.doi.org/10.1016/j.molcel.2017.09.018.

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41

Kurat, Christoph F., Joseph T. P. Yeeles, Harshil Patel, Anne Early, and John F. X. Diffley. "Chromatin Controls DNA Replication Origin Selection, Lagging-Strand Synthesis, and Replication Fork Rates." Molecular Cell 65, no. 1 (January 2017): 117–30. http://dx.doi.org/10.1016/j.molcel.2016.11.016.

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42

Kresge, Nicole, Robert D. Simoni, and Robert L. Hill. "DNA Polymerase and Leading and Lagging Strand Synthesis: the Work of Bruce Alberts." Journal of Biological Chemistry 282, no. 4 (January 2007): e3-e5. http://dx.doi.org/10.1016/s0021-9258(20)72146-0.

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43

Cerrón, Fernando, Sara de Lorenzo, Kateryna M. Lemishko, Grzegorz L. Ciesielski, Laurie S. Kaguni, Francisco J. Cao, and Borja Ibarra. "Replicative DNA polymerases promote active displacement of SSB proteins during lagging strand synthesis." Nucleic Acids Research 47, no. 11 (April 10, 2019): 5723–34. http://dx.doi.org/10.1093/nar/gkz249.

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44

Barry, Jack, Mei Lie Wong,, and Bruce Alberts. "In vitro reconstitution of DNA replication initiated by genetic recombination: a T4 bacteriophage model for a type of DNA synthesis important for all cells." Molecular Biology of the Cell 30, no. 1 (January 2019): 146–59. http://dx.doi.org/10.1091/mbc.e18-06-0386.

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Using a mixture of 10 purified DNA replication and DNA recombination proteins encoded by the bacteriophage T4 genome, plus two homologous DNA molecules, we have reconstituted the genetic recombination–initiated pathway that initiates DNA replication forks at late times of T4 bacteriophage infection. Inside the cell, this recombination-dependent replication (RDR) is needed to produce the long concatemeric T4 DNA molecules that serve as substrates for packaging the shorter, genome-sized viral DNA into phage heads. The five T4 proteins that catalyze DNA synthesis on the leading strand, plus the proteins required for lagging-strand DNA synthesis, are essential for the reaction, as are a special mediator protein (gp59) and a Rad51/RecA analogue (the T4 UvsX strand-exchange protein). Related forms of RDR are widespread in living organisms—for example, they play critical roles in the homologous recombination events that can restore broken ends of the DNA double helix, restart broken DNA replication forks, and cross over chromatids during meiosis in eukaryotes. Those processes are considerably more complex, and the results presented here should be informative for dissecting their detailed mechanisms.

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45

Bainbridge, Lewis J., Rebecca Teague, and Aidan J. Doherty. "Repriming DNA synthesis: an intrinsic restart pathway that maintains efficient genome replication." Nucleic Acids Research 49, no. 9 (March 21, 2021): 4831–47. http://dx.doi.org/10.1093/nar/gkab176.

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Abstract To bypass a diverse range of fork stalling impediments encountered during genome replication, cells possess a variety of DNA damage tolerance (DDT) mechanisms including translesion synthesis, template switching, and fork reversal. These pathways function to bypass obstacles and allow efficient DNA synthesis to be maintained. In addition, lagging strand obstacles can also be circumvented by downstream priming during Okazaki fragment generation, leaving gaps to be filled post-replication. Whether repriming occurs on the leading strand has been intensely debated over the past half-century. Early studies indicated that both DNA strands were synthesised discontinuously. Although later studies suggested that leading strand synthesis was continuous, leading to the preferred semi-discontinuous replication model. However, more recently it has been established that replicative primases can perform leading strand repriming in prokaryotes. An analogous fork restart mechanism has also been identified in most eukaryotes, which possess a specialist primase called PrimPol that conducts repriming downstream of stalling lesions and structures. PrimPol also plays a more general role in maintaining efficient fork progression. Here, we review and discuss the historical evidence and recent discoveries that substantiate repriming as an intrinsic replication restart pathway for maintaining efficient genome duplication across all domains of life.

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46

Martin, Aegina Adams, Isabelle Dionne, Raymund J. Wellinger, and Connie Holm. "The Function of DNA Polymerase α at Telomeric G Tails Is Important for Telomere Homeostasis." Molecular and Cellular Biology 20, no. 3 (February 1, 2000): 786–96. http://dx.doi.org/10.1128/mcb.20.3.786-796.2000.

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ABSTRACT Telomere length control is influenced by several factors, including telomerase, the components of telomeric chromatin structure, and the conventional replication machinery. Although known components of the replication machinery can influence telomere length equilibrium, little is known about why mutations in certain replication proteins cause dramatic telomere lengthening. To investigate the cause of telomere elongation in cdc17/pol1 (DNA polymerase α) mutants, we examined telomeric chromatin, as measured by its ability to repress transcription on telomere-proximal genes, and telomeric DNA end structures in pol1-17 mutants. pol1-17 mutants with elongated telomeres show a dramatic loss of the repression of telomere-proximal genes, or telomeric silencing. In addition,cdc17/pol1 mutants grown under telomere-elongating conditions exhibit significant increases in single-stranded character in telomeric DNA but not at internal sequences. The single strandedness is manifested as a terminal extension of the G-rich strand (G tails) that can occur independently of telomerase, suggesting thatcdc17/pol1 mutants exhibit defects in telomeric lagging-strand synthesis. Interestingly, the loss of telomeric silencing and the increase in the sizes of the G tails at the telomeres temporally coincide and occur before any detectable telomere lengthening is observed. Moreover, the G tails observed incdc17/pol1 mutants incubated at the semipermissive temperature appear only when the cells pass through S phase and are processed by the time cells reach G1. These results suggest that lagging-strand synthesis is coordinated with telomerase-mediated telomere maintenance to ensure proper telomere length control.

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47

Prelich, Gregory, and Bruce Stillman. "Coordinated leading and lagging strand synthesis during SV40 DNA replication in vitro requires PCNA." Cell 53, no. 1 (April 1988): 117–26. http://dx.doi.org/10.1016/0092-8674(88)90493-x.

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48

Muzi-Falconi, Marco, Michele Giannattasio, Marco Foiani, and Paolo Plevani. "The DNA Polymerase _-Primase Complex: Multiple Functions and Interactions." Scientific World JOURNAL 3 (2003): 21–33. http://dx.doi.org/10.1100/tsw.2003.05.

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DNA polymerase _ (pol _) holds a special position among the growing family of eukaryotic DNA polymerases. In fact, pol _ is associated with DNA primase to form a four subunit complex and, as a consequence, is the only enzyme able to start DNA synthesis de novo. Because of this peculiarity the major role of the DNA polymerase _-primase complex (pol-prim) is in the initiation of DNA replication at chromosomal origins and in the discontinuous synthesis of Okazaki fragments on the lagging strand of the replication fork. However, pol-prim seems to play additional roles in other complex cellular processes, such as the response to DNA damage, telomere maintenance, and the epigenetic control of higher order chromatin assembly.

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Gangavarapu, Venkateswarlu, Satya Prakash, and Louise Prakash. "Requirement of RAD52 Group Genes for Postreplication Repair of UV-Damaged DNA in Saccharomyces cerevisiae." Molecular and Cellular Biology 27, no. 21 (September 4, 2007): 7758–64. http://dx.doi.org/10.1128/mcb.01331-07.

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ABSTRACT In Saccharomyces cerevisiae, replication through DNA lesions is promoted by Rad6-Rad18-dependent processes that include translesion synthesis by DNA polymerases η and ζ and a Rad5-Mms2-Ubc13-controlled postreplicational repair (PRR) pathway which repairs the discontinuities in the newly synthesized DNA that form opposite from DNA lesions on the template strand. Here, we examine the contributions of the RAD51, RAD52, and RAD54 genes and of the RAD50 and XRS2 genes to the PRR of UV-damaged DNA. We find that deletions of the RAD51, RAD52, and RAD54 genes impair the efficiency of PRR and that almost all of the PRR is inhibited in the absence of both Rad5 and Rad52. We suggest a role for the Rad5 pathway when the lesion is located on the leading strand template and for the Rad52 pathway when the lesion is located on the lagging strand template. We surmise that both of these pathways operate in a nonrecombinational manner, Rad5 by mediating replication fork regression and template switching via its DNA helicase activity and Rad52 via a synthesis-dependent strand annealing mode. In addition, our results suggest a role for the Rad50 and Xrs2 proteins and thereby for the MRX complex in promoting PRR via both the Rad5 and Rad52 pathways.

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Nethanel, T., and G. Kaufmann. "Two DNA polymerases may be required for synthesis of the lagging DNA strand of simian virus 40." Journal of Virology 64, no. 12 (1990): 5912–18. http://dx.doi.org/10.1128/jvi.64.12.5912-5918.1990.

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Journal articles on the topic 'Lagging-strand DNA synthesis'

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