Translesion Synthesis Past 5-Formylcytosine-Mediated DNA–Peptide Cross-Links by hPolη Is Dependent on the Local DNA Sequence

Thomforde, Jenna; Fu, Iwen; Rodriguez, Freddys; Pujari, Suresh S.; Broyde, Suse; Tretyakova, Natalia

doi:10.1021/acs.biochem.1c00130

Cited by 9 publications

(11 citation statements)

References 49 publications

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“…This approach allows for efficient, site specific bioconjugation of protein to DNA without the need for harsh chemicals such as NaCNBH 3 . Molecular dynamics simulations reveal that the presence of an oxygen in place of CH 2 group at the ϵ position of lysine has minimal effect on protein and DPC structure, [8a] allowing for future use of these model lesions in biological experiments in vivo and in vitro.…”

Section: Methodsmentioning

confidence: 99%

Site‐Specific 5‐Formyl Cytosine Mediated DNA‐Histone Cross‐Links: Synthesis and Polymerase Bypass by Human DNA Polymerase η

Pujari

Thomforde

et al. 2021

Angew Chem Int Ed

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DNA-protein cross-links (DPCs) between DNA epigenetic mark 5-formylC and lysine residues of histone proteins spontaneously form in human cells. Such conjugates are likely to influence chromatin structure and mediate DNA replication, transcription, and repair, but are challenging to study due to their reversible nature. Here we report the construction of site specific, hydrolytically stable DPCs between 5fdC in DNA and K4 of histone H3 and an investigation of their effects on DNA replication. Our approach employs oxime ligation, allowing for site-specific conjugation of histones to DNA under physiological conditions. Primer extension experiments revealed that histone H3-DNA crosslinks blocked DNA synthesis by hPol h polymerase, but were bypassed following proteolytic processing.Irreversible entrapment of cellular proteins on genomic DNA following exposure to bis-electrophiles, transition metals, and free radical species gives rise to DNA-protein cross-links (DPCs). [1] DPCs are unusually bulky DNA lesions that have the ability to block DNA transactions including transcription, replication, and repair, potentially leading to genomic instability and cell death. [2] DPCs accumulate in the heart and brain tissues with age and are hypothesized to play an important role in aging, cancer and neurodegenerative diseases. [3] Individuals deficient in SPRTN, a critical gene required for DPC repair, develop the Ruijs-Aalfs syndrome, a human autosomal recessive disorder characterized by accelerated aging, genomic instability, and early-onset of hepatocellular carcinoma. [4] Scheme 1. Reversible histone-DNA cross-linking and their stabilization via reduction of the resulting Schiff bases.

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Section: Methodsmentioning

confidence: 99%

Site‐Specific 5‐Formyl Cytosine Mediated DNA‐Histone Cross‐Links: Synthesis and Polymerase Bypass by Human DNA Polymerase η

Pujari

Thomforde

et al. 2021

Angew Chem Int Ed

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“…Once TLS Pols are in the vicinity of the damage site, Pol affinity for the DNA substrate becomes a driver of Pol selection. Accommodation of lesions by TLS Pols can be influenced by template-strand sequence and steric hindrance between bulkier lesions and Pol active sites ( Zhao and Washington, 2017 ; Thomforde et al, 2021 ). Sequence context has also been shown to affect Pol efficiency and accuracy during TLS opposite various lesions, however the molecular basis for this effect is not well understood ( Shriber et al, 2015 ; Bacurio et al, 2021 ).…”

Section: Prelesion: the First Translesion Synthesis Polymerase Substitutionmentioning

confidence: 99%

Beyond the Lesion: Back to High Fidelity DNA Synthesis

Kaszubowski

Trakselis

2022

Front. Mol. Biosci.

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High fidelity (HiFi) DNA polymerases (Pols) perform the bulk of DNA synthesis required to duplicate genomes in all forms of life. Their structural features, enzymatic mechanisms, and inherent properties are well-described over several decades of research. HiFi Pols are so accurate that they become stalled at sites of DNA damage or lesions that are not one of the four canonical DNA bases. Once stalled, the replisome becomes compromised and vulnerable to further DNA damage. One mechanism to relieve stalling is to recruit a translesion synthesis (TLS) Pol to rapidly synthesize over and past the damage. These TLS Pols have good specificities for the lesion but are less accurate when synthesizing opposite undamaged DNA, and so, mechanisms are needed to limit TLS Pol synthesis and recruit back a HiFi Pol to reestablish the replisome. The overall TLS process can be complicated with several cellular Pols, multifaceted protein contacts, and variable nucleotide incorporation kinetics all contributing to several discrete substitution (or template hand-off) steps. In this review, we highlight the mechanistic differences between distributive equilibrium exchange events and concerted contact-dependent switching by DNA Pols for insertion, extension, and resumption of high-fidelity synthesis beyond the lesion.

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“…Cross-linking targets multiple sites of DNA, including the C-5 methyl group of thymine, the N7 of guanine, and exocyclic amines of adenine, cytosine, and guanine . If not repaired, DPCs constitute a complete block to replication and transcription machinery and thus represent a serious threat to cell viability. ,− With the exception of topoisomerase DPCs that can be directly reversed by tyrosyl-DNA phosphodiesterases (TDP1 and TDP2), cellular repair of DPCs requires the activity of proteolytic enzymes to break the protein down to smaller peptides. ,− The resulting DNA–peptide cross-links (DpC) can be bypassed by DNA polymerases ,− and are subject to canonical DNA repair pathways. − Proteolytic cleavage can be accomplished through the activity of SPRTN metalloprotease or the proteasome . DNA repair pathways participating in DpC tolerance and repair include translesion synthesis polymerases , and the nucleotide excision pathway (NER). , …”

Section: Introductionmentioning

confidence: 99%

Endogenous Cellular Metabolite Methylglyoxal Induces DNA–Protein Cross-Links in Living Cells

Hurben,

Zhang,

Galligan

et al. 2024

ACS Chem. Biol.

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Methylglyoxal (MGO) is an electrophilic α-oxoaldehyde generated endogenously through metabolism of carbohydrates and exogenously due to autoxidation of sugars, degradation of lipids, and fermentation during food and drink processing. MGO can react with nucleophilic sites within proteins and DNA to form covalent adducts. MGO-induced advanced glycation end-products such as protein and DNA adducts are thought to be involved in oxidative stress, inflammation, diabetes, cancer, renal failure, and neurodegenerative diseases. Additionally, MGO has been hypothesized to form toxic DNA−protein crosslinks (DPC), but the identities of proteins participating in such cross-linking in cells have not been determined. In the present work, we quantified DPC formation in human cells exposed to MGO and identified proteins trapped on DNA upon MGO exposure using mass spectrometry-based proteomics. A total of 265 proteins were found to participate in MGO-derived DPC formation including gene products engaged in telomere organization, nucleosome assembly, and gene expression. In vitro experiments confirmed DPC formation between DNA and glyceraldehyde-3-phosphate dehydrogenase (GAPDH), as well as histone proteins H3.1 and H4. Collectively, our study provides the first evidence for MGO-mediated DNA−protein cross-linking in living cells, prompting future studies regarding the relevance of these toxic lesions in cancer, diabetes, and other diseases linked to elevated MGO levels.

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Translesion Synthesis Past 5-Formylcytosine-Mediated DNA–Peptide Cross-Links by hPolη Is Dependent on the Local DNA Sequence

Cited by 9 publications

References 49 publications

Site‐Specific 5‐Formyl Cytosine Mediated DNA‐Histone Cross‐Links: Synthesis and Polymerase Bypass by Human DNA Polymerase η

Site‐Specific 5‐Formyl Cytosine Mediated DNA‐Histone Cross‐Links: Synthesis and Polymerase Bypass by Human DNA Polymerase η

Beyond the Lesion: Back to High Fidelity DNA Synthesis

Endogenous Cellular Metabolite Methylglyoxal Induces DNA–Protein Cross-Links in Living Cells

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