Clustered DNA Lesions Containing 5-Formyluracil and AP Site: Repair via the BER System

The regulation of repair processes including base excision repair (BER) in the presence of DNA damage is implemented by a cellular signal: poly(ADP-ribosyl)ation (PARylation), which is catalysed by PARP1 and PARP2. Despite ample studies, it is far from clear how BER is regulated by PARPs and how the roles are distributed between the PARPs. Here, we investigated the effects of PARP1, PARP2 and PARylation on activities of the main BER enzymes (APE1, DNA polymerase β [Polβ] and DNA ligase IIIα [LigIIIα]) in combination with BER scaffold protein XRCC1 in the nucleosomal context. We constructed nucleosome core particles with midward- or outward-oriented damage. It was concluded that in most cases, the presence of PARP1 leads to the suppression of the activities of APE1, Polβ and to a lesser extent LigIIIα. PARylation by PARP1 attenuated this effect to various degrees depending on the enzyme. PARP2 had an influence predominantly on the last stage of BER: DNA sealing. Nonetheless, PARylation by PARP2 led to Polβ inhibition and to significant stimulation of LigIIIα activities in a NAD+-dependent manner. On the basis of the obtained and literature data, we suggest a hypothetical model of the contribution of PARP1 and PARP2 to BER.

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“…Human LigIIIα and XRCC1 were purified according to Refs. 79 , 80 , whereas human APE1, rat Polβ and PARG according to Refs. 81 – 83 .…”

Section: Methodsmentioning

confidence: 99%

The contribution of PARP1, PARP2 and poly(ADP-ribosyl)ation to base excision repair in the nucleosomal context

Kutuzov

Belousova

Kurgina

et al. 2021

Sci Rep

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“…Recombinant human apurinic/apyrimidinic endonuclease 1 (APE1), rat DNA polymerase β (Polβ) and human tyrosyl-DNA phosphodiesterase 1 (TDP1) were produced by expression in Escherichia coli BL21(DE3)pLysS and purified as described previously ( 14 – 16 ). Human protein XRCC1 and human poly(ADP-ribose) polymerase 1 (PARP1) were produced by expression in E. coli Rosetta(DE3) and purified as described ( 17 , 18 ). A 24-kDa fragment of human PARP1 (p24) was produced by expression in E. coli BL21(DE3)pLysS and purified following the procedure described for PARP1 ( 18 ).…”

Section: Methodsmentioning

confidence: 99%

Quantitative characterization of protein–protein complexes involved in base excision DNA repair

et al. 2015

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Base Excision Repair (BER) efficiently corrects the most common types of DNA damage in mammalian cells. Step-by-step coordination of BER is facilitated by multiple interactions between enzymes and accessory proteins involved. Here we characterize quantitatively a number of complexes formed by DNA polymerase β (Polβ), apurinic/apyrimidinic endonuclease 1 (APE1), poly(ADP-ribose) polymerase 1 (PARP1), X-ray repair cross-complementing protein 1 (XRCC1) and tyrosyl-DNA phosphodiesterase 1 (TDP1), using fluorescence- and light scattering-based techniques. Direct physical interactions between the APE1-Polβ, APE1-TDP1, APE1-PARP1 and Polβ-TDP1 pairs have been detected and characterized for the first time. The combined results provide strong evidence that the most stable complex is formed between XRCC1 and Polβ. Model DNA intermediates of BER are shown to induce significant rearrangement of the Polβ complexes with XRCC1 and PARP1, while having no detectable influence on the protein–protein binding affinities. The strength of APE1 interaction with Polβ, XRCC1 and PARP1 is revealed to be modulated by BER intermediates to different extents, depending on the type of DNA damage. The affinity of APE1 for Polβ is higher in the complex with abasic site-containing DNA than after the APE1-catalyzed incision. Our findings advance understanding of the molecular mechanisms underlying coordination and regulation of the BER process.

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“…When an AP site is located directly opposite 5-fdU, it is repaired through the long-patch BER pathway; by contrast, when the AP site is shifted it is primarily repaired by the short-patch BER pathway. 238 …”

Section: Repair and Biological Consequences Of Oxidative Stress-inmentioning

confidence: 99%

Occurrence, Biological Consequences, and Human Health Relevance of Oxidative Stress-Induced DNA Damage

Cui

Niedernhofer

et al. 2016

Chem. Res. Toxicol.

146

135

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A variety of endogenous and exogenous agents can induce DNA damage and lead to genomic instability. Reactive oxygen species (ROS), an important class of DNA damaging agents, are constantly generated in cells as a consequence of endogenous metabolism, infection/inflammation, and/or exposure to environmental toxicants. A wide array of DNA lesions can be induced by ROS directly, including single-nucleobase lesions, tandem lesions, and hypochlorous acid (HOCl)/hypobromous acid (HOBr)-derived DNA adducts. ROS can also lead to lipid peroxidation, whose byproducts can also react with DNA to produce exocyclic DNA lesions. A combination of bioanalytical chemistry, synthetic organic chemistry, and molecular biology approaches have provided significant insights into the occurrence, repair, and biological consequences of oxidatively induced DNA lesions. The involvement of these lesions in the etiology of human diseases and aging was also investigated in the past several decades, suggesting that the oxidatively induced DNA adducts, especially bulky DNA lesions, may serve as biomarkers for exploring the role of oxidative stress in human diseases. The continuing development and improvement of LC-MS/MS coupled with the stable isotope-dilution method for DNA adduct quantification will further promote research about the clinical implications and diagnostic applications of oxidatively induced DNA adducts.

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Clustered DNA Lesions Containing 5-Formyluracil and AP Site: Repair via the BER System

Cited by 21 publications

References 34 publications

The contribution of PARP1, PARP2 and poly(ADP-ribosyl)ation to base excision repair in the nucleosomal context

The contribution of PARP1, PARP2 and poly(ADP-ribosyl)ation to base excision repair in the nucleosomal context

Quantitative characterization of protein–protein complexes involved in base excision DNA repair

Occurrence, Biological Consequences, and Human Health Relevance of Oxidative Stress-Induced DNA Damage

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