Polymerase theta-mediated end joining of replication-associated DNA breaks in <i>C. elegans</i>

Sf, Roerink; R, van Schendel; Tijsterman, Marcel

doi:10.1101/gr.170431.113

Cited by 152 publications

(169 citation statements)

References 37 publications

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“…Strikingly, one other eukaryotic polymerase has a related activity-Pol θ also directs synthesis across double-strand break ends, although it needs a short patch of terminal complementary sequence, or microhomology, to initiate synthesis-and it is essential for an alternate NHEJ pathway (29)(30)(31). More complex and longer-lived organisms are expected to need more flexible and efficient versions of end joining; it is clear that the use of specialized polymerases is an integral part of how this has been achieved.…”

Section: Discussionmentioning

confidence: 99%

Essential role for polymerase specialization in cellular nonhomologous end joining

Pryor

Waters

Aza

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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Nonhomologous end joining (NHEJ) repairs chromosome breaks and must remain effective in the face of extensive diversity in broken end structures. We show here that this flexibility is often reliant on the ability to direct DNA synthesis across strand breaks, and that polymerase (Pol) μ and Pol λ are the only mammalian DNA polymerases that have this activity. By systematically varying substrate in cells, we show each polymerase is uniquely proficient in different contexts. The templating nucleotide is also selected differently, with Pol μ using the unpaired base adjacent to the downstream 5′ phosphate even when there are available template sites further upstream of this position; this makes Pol μ more flexible but also less accurate than Pol λ. Loss of either polymerase alone consequently has clear and distinguishable effects on the fidelity of repair, but end remodeling by cellular nucleases and the remaining polymerase helps mitigate the effects on overall repair efficiency. Accordingly, when cells are deficient in both polymerases there is synergistic impact on NHEJ efficiency, both in terms of repair of defined substrates and cellular resistance to ionizing radiation. Pol μ and Pol λ thus provide distinct solutions to a problem for DNA synthesis that is unique to this pathway and play a key role in conferring on NHEJ the flexibility required for accurate and efficient repair.nonhomologous end joining | double-strand break repair | Pol X | polymerase mu | polymerase lambda

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Section: Discussionmentioning

confidence: 99%

Essential role for polymerase specialization in cellular nonhomologous end joining

Pryor

Waters

Aza

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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“…The inserted bases for another three variants appeared to have been templated from the neighboring sequence. Such palindromic and templated complex indels have been reported in model organisms around double-stranded break repairs through synthesisdependent microhomology-mediated end joining (SD-MMEJ) (Yu and McVey 2010) and theta-mediated end joining (TMEJ) (Roerink et al 2014). The formation of these indels likely follows a multistep process involving resection of break ends, hairpin …”

Section: Indel Formationmentioning

confidence: 90%

Characteristics of de novo structural changes in the human genome

et al. 2015

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Small insertions and deletions (indels) and large structural variations (SVs) are major contributors to human genetic diversity and disease. However, mutation rates and characteristics of de novo indels and SVs in the general population have remained largely unexplored. We report 332 validated de novo structural changes identified in whole genomes of 250 families, including complex indels, retrotransposon insertions, and interchromosomal events. These data indicate a mutation rate of 2.94 indels (1-20 bp) and 0.16 SVs (>20 bp) per generation. De novo structural changes affect on average 4.1 kbp of genomic sequence and 29 coding bases per generation, which is 91 and 52 times more nucleotides than de novo substitutions, respectively. This contrasts with the equal genomic footprint of inherited SVs and substitutions. An excess of structural changes originated on paternal haplotypes. Additionally, we observed a nonuniform distribution of de novo SVs across offspring. These results reveal the importance of different mutational mechanisms to changes in human genome structure across generations.

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“…The relevance of these activities in vivo was highlighted in D. melanogaster , where Polθ was shown to stimulate nucleotide insertions during DSB repair by alt-NHEJ 14 . More recently, Polθ was shown to promote end-joining of replication-associated DSBs in C. elegans 15 , preventing large deletions around G-rich DNA 16 . The exact function of Polθ during DSB repair in mammalian cells remains elusive.…”

mentioning

confidence: 99%

Mammalian polymerase θ promotes alternative NHEJ and suppresses recombination

Mateos-Gómez

Gong

Nair

et al. 2015

Nature

616

672

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The alternative nonhomologous end-joining (alt-NHEJ) machinery facilitates a number of genomic rearrangements, some of which can lead to cellular transformation. This error-prone repair pathway is triggered upon telomere de-protection to promote the formation of deleterious chromosome end-to-end fusions1,2,3. Using next-generation sequencing technology, we found that repair by alt-NHEJ yields non-TTAGGG nucleotide insertions at fusion breakpoints of dysfunctional telomeres. Investigating the enzymatic activity responsible for the random insertions enabled us to identify Polymerase theta (Polθ; encoded by PolQ) as a critical alt-NHEJ factor in mammalian cells. PolQ inhibition suppresses alt-NHEJ at dysfunctional telomeres, and hinders chromosomal translocations at non-telomeric loci. In addition, we found that PolQ loss results in increased rates of homology directed repair (HDR), evident by recombination of dysfunctional telomeres and accumulation of Rad51 at double stranded breaks. Lastly, we show that depletion of PolQ has a synergistic impact on cell survival in the absence of BRCA genes, suggesting that the inhibition of this mutagenic polymerase represents a valid therapeutic avenue for tumors carrying mutations in HDR genes.

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Polymerase theta-mediated end joining of replication-associated DNA breaks in C. elegans

Cited by 152 publications

References 37 publications

Essential role for polymerase specialization in cellular nonhomologous end joining

Essential role for polymerase specialization in cellular nonhomologous end joining

Characteristics of de novo structural changes in the human genome

Mammalian polymerase θ promotes alternative NHEJ and suppresses recombination

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