Suppressed catalytic activity of base excision repair enzymes on  rotationally positioned uracil in nucleosomes

Beard, Brian C.; Wilson, Samuel H.; Smerdon, Michael J.

doi:10.1073/pnas.1330328100

Cited by 143 publications

(161 citation statements)

References 61 publications

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“…It is possible that nick ligation does not require extensive contact or movement of the ligase along the DNA molecule. Nucleotide excision repair is another example for which the presence of nucleosomes is also inhibitory (16)(17)(18)(19)(20). Furthermore, chromatin remodeling factors, which serve to recover nucleotide excision repair on synthetic dinucleosomes, have been shown to be inhibited by the presence of histone H1 within the nucleosomal arrays (21,22,43).…”

Section: Discussionmentioning

confidence: 99%

“…Now that many repair processes can be reconstituted in vitro, studies are beginning to examine the impact of chromatin structure on repair. Nucleotide excision repair and base excision repair were shown to be impaired on nucleosome substrates relative to naked DNA (16)(17)(18)(19)(20) and repair efficiencies are recovered by the presence of ATP-dependent chromatin remodeling factors (21)(22)(23). In contrast, human DNA ligase I and Flap I endonuclease function efficiently on model nucleosome substrates (24,25).…”

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confidence: 99%

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Phosphorylation of linker histones by DNA-dependent protein kinase is required for DNA ligase IV-dependent ligation in the presence of histone H1

Kysela

Chovanec

Jeggo

2005

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

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DNA nonhomologous end-joining in vivo requires the DNA-dependent protein kinase (DNA-PK) and DNA ligase IV͞XRCC4 (LX) complexes. Here, we have examined the impact of histone octamers and linker histone H1 on DNA end-joining in vitro. Packing of the DNA substrate into dinucleosomes does not significantly inhibit ligation by LX. However, LX ligation activity is substantially reduced by the incorporation of linker histones. This inhibition is independent of the presence of core histone octamers and cannot be restored by addition of Ku alone but can be partially rescued by DNA-PK. The kinase activity of DNA-PK is essential for the recovery of end-joining. DNA-PK efficiently phosphorylates histone H1. Phosphorylated histone H1 has a reduced affinity for DNA and a decreased capacity to inhibit end-joining. Our findings raise the possibility that DNA-PK may act as a linker histone kinase by phosphorylating linker histones in the vicinity of a DNA break and coupling localized histone H1 release from DNA ends, with the recruitment of LX to carry out double-stranded ligation. Thus, by using histone H1-bound DNA as a template, we have reconstituted the end-joining step of DNA nonhomologous end-joining in vitro with a requirement for DNA-PK.chromatin ͉ DNA double-stranded break repair ͉ nonhomologous end-joining D NA nonhomologous end-joining (NHEJ) is the major mechanism for the repair of DNA double-stranded (ds) breaks (DSBs) in mammalian cells and functions to effect DNA rearrangement during V(D)J recombination. Five proteins forming two complexes are required for NHEJ in mammalian cells, namely DNA ligase IV͞XRCC4 (LX) complex and the DNA-dependent protein kinase (DNA-PK) complex, which encompasses the two subunits of the Ku heterodimer and a large catalytic subunit (DNA-PKcs) (1, 2). Recently, a sixth protein, Artemis, has been shown to cleave the hairpin intermediate generated during V(D)J recombination (3). Although DNA-PK phosphorylation is required for the hairpin cleavage activity of Artemis and the biochemical activities of the DNA-PK and LX complexes have been established in vitro, little is known about the role of DNA-PK in DSB rejoining or its in vivo phosphorylation targets. Current models suggest that DNA-PK may regulate NHEJ (4-8).Recently, we have shown that LX is most efficient on substrates Ϸ400 bp long and inefficient on short (Ϸ50-bp) oligomers (9). Ku can stimulate LX ligation in a manner that requires freedom for Ku to translocate inwardly on the DNA (9-11). However, the presence of the intact DNA-PK complex does not provide any further stimulation of end-joining, and, indeed, its presence can be inhibitory (see Results). Our finding that efficient LX ligation in the presence of Ku requires inward translocation of Ku raises the question of how the presence of nucleosomes and the complex structural packaging of DNA into chromatin might impact the efficiency of NHEJ. The fundamental structural unit of chromatin is the nucleosome, an octameric complex of two copies of each core histone (H2A, H2B, H3, an...

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

Phosphorylation of linker histones by DNA-dependent protein kinase is required for DNA ligase IV-dependent ligation in the presence of histone H1

Kysela

Chovanec

Jeggo

2005

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

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“…These include the FEN1-mediated DNA cleavage and DNA ligase I-mediated ligation [58,59]. However, replacement of the correct nucleotide by DNA polymerase beta is blocked by nucleosomes in vitro [60]. While it is possible that chromatin remodeling rather than gross chromatin alterations such as chromatin disassembly and reassembly may be sufficient for short patch BER, long patch BER is likely to require chromatin disassembly and reassembly.…”

Section: Chromatin Assembly During Excision Repairmentioning

confidence: 99%

Chromatin disassembly and reassembly during DNA repair

Linger

Tyler

2007

Mutation Research/Fundamental and Molecular Mechanisms of Mutag

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Current research is demonstrating that the packaging of the eukaryotic genome together with histone proteins into chromatin is playing a fundamental role in DNA repair and the maintenance of genomic integrity. As is well established to be the case for transcription, the chromatin structure dynamically changes during DNA repair. Recent studies indicate that the complete removal of histones from DNA and their subsequent reassembly onto DNA accompanies DNA repair. This review will present evidence indicating that chromatin disassembly and reassembly occur during DNA repair and that these are critical processes for cell survival after DNA repair. Concomitantly, candidate proteins utilized for these processes will be highlighted.

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“…Furthermore, the amount of AID protein used in in vitro reactions to detect DNA deamination is far in excess of that of in vivo reaction. It is also important to note that chromatin DNA may be a much less efficient substrate for deamination compared with naked DNA, as shown in uracil DNA glycosylase activity of UNG (34). In addition, U removal activity of UNG is shown to be dispensable for CSR (35).…”

Section: Requirement Of De Novo Protein Synthesis For ␥-H2axmentioning

confidence: 99%

De novo protein synthesis is required for activation-induced cytidine deaminase-dependent DNA cleavage in immunoglobulin class switch recombination

Begum

Kinoshita

Muramatsu

et al. 2004

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

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Activation-induced cytidine deaminase is required for the DNA cleavage step of Ig class switch recombination (CSR). However, its molecular mechanism is controversial. RNA-editing hypothesis postulates that activation-induced cytidine deaminase deaminates cytosine in an unknown mRNA to generate a new mRNA encoding an endonuclease for CSR and thus predicts that DNA cleavage depends on de novo protein synthesis. On the other hand, DNA deamination hypothesis proposes that DNA cleavage is initiated by cytosine deamination in DNA, followed by uracil removal by uracil DNA glycosylase. By using the chromatin immunoprecipitation assay to detect ␥-H2AX focus formation as a marker for DNA cleavage, we found that cycloheximide inhibited DNA cleavage in the Ig heavy-chain locus during CSR. Requirement of protein synthesis in the DNA cleavage step of CSR strengthens the RNAediting hypothesis.

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Suppressed catalytic activity of base excision repair enzymes on rotationally positioned uracil in nucleosomes

Cited by 143 publications

References 61 publications

Phosphorylation of linker histones by DNA-dependent protein kinase is required for DNA ligase IV-dependent ligation in the presence of histone H1

Phosphorylation of linker histones by DNA-dependent protein kinase is required for DNA ligase IV-dependent ligation in the presence of histone H1

Chromatin disassembly and reassembly during DNA repair

De novo protein synthesis is required for activation-induced cytidine deaminase-dependent DNA cleavage in immunoglobulin class switch recombination

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