Trinucleotide repeat (TNR) expansion is the cause of over 40 neurodegenerative diseases, including Huntington\'s disease and Friedreich\'s ataxia (FRDA). There are no effective treatments for these diseases due to the poor understanding of molecular mechanisms underlying somatic TNR expansion and contraction in neural systems. We and others have found that DNA base excision repair (BER) actively modulates TNR instability, shedding light on the development of effective treatments for the diseases by contracting expanded repeats through DNA repair. In this study, temozolomide (TMZ) was employed as a model DNA base damaging agent to reveal the mechanisms of the BER pathway in modulating GAA repeat instability at the frataxin (FXN) gene in FRDA neural cells and transgenic mouse mice. We found that TMZ induced large GAA repeat contraction in FRDA mouse brain tissue, neurons, and FRDA iPSC-differentiated neural cells, increasing frataxin protein levels in FRDA mouse brain and neural cells. Surprisingly, we found that TMZ could also inhibit H3K9 methyltransferases, leading to open chromatin and increasing ssDNA breaks and recruitment of the key BER enzyme, pol β, on the repeats in FRDA neural cells. We further demonstrated that the H3K9 methyltransferase inhibitor BIX01294 also induced the contraction of the expanded repeats and increased frataxin protein in FRDA neural cells by opening the chromatin and increasing the endogenous ssDNA breaks and recruitment of pol β on the repeats. Our study provides new mechanistic insight illustrating that inhibition of H3K9 methylation can crosstalk with BER to induce GAA repeat contraction in FRDA. Our results will open a new avenue for developing novel gene therapy by targeting histone methylation and the BER pathway for repeat expansion diseases.

Non-membrane compartments or biomolecular condensates play an important role in the regulation of cellular processes including DNA repair. Here, an ability of XRCC1, a scaffold protein involved in DNA base excision repair (BER) and single-strand break repair, to form protein-rich microphases in the presence of DNA duplexes was discovered. We also showed that the gap-filling activity of BER-related DNA polymerase λ (Pol λ) is significantly increased by the presence of XRCC1. The stimulation of the Pol λ activity was observed only at micromolar XRCC1 concentrations, which were well above the nanomolar dissociation constant determined for the XRCC1-Pol λ complex and pointed to the presence of an auxiliary stimulatory factor in addition to protein-protein interactions. Indeed, according to dynamic light scattering measurements, the stimulation of the Pol λ activity by XRCC1 was coupled with microphase separation in a protein-DNA mixture. Fluorescence microscopy revealed colocalization of Pol λ, XRCC1, and gapped DNA within the microphases. Thus, stimulation of Pol λ activity is caused both by its interaction with XRCC1 and by specific conditions of microphase separation; this phenomenon is shown for the first time.

Mutations in isocitrate dehydrogenase isoform 1 (IDH1) are primarily found in secondary glioblastoma (GBM) and low-grade glioma but are rare in primary GBM. The standard treatment for GBM includes radiation combined with temozolomide, an alkylating agent. Fortunately, IDH1 mutant gliomas are sensitive to this treatment, resulting in a more favorable prognosis. However, it\'s estimated that up to 75 % of IDH1 mutant gliomas will progress to WHO grade IV over time and develop resistance to alkylating agents. Therefore, understanding the mechanism(s) by which IDH1 mutant gliomas confer sensitivity to alkylating agents is crucial for developing targeted chemotherapeutic approaches. The base excision repair (BER) pathway is responsible for repairing most base damage induced by alkylating agents. Defects in this pathway can lead to hypersensitivity to these agents due to unresolved DNA damage. The coordinated assembly and disassembly of BER protein complexes are essential for cell survival and for maintaining genomic integrity following alkylating agent exposure. These complexes rely on poly-ADP-ribose formation, an NAD+-dependent post-translational modification synthesized by PARP1 and PARP2 during the BER process. At the lesion site, poly-ADP-ribose facilitates the recruitment of XRCC1. This scaffold protein helps assemble BER proteins like DNA polymerase beta (Polβ), a bifunctional DNA polymerase containing both DNA synthesis and 5\'-deoxyribose-phosphate lyase (5\'dRP lyase) activity. Here, we confirm that IDH1 mutant glioma cells have defective NAD+ metabolism, but still produce sufficient nuclear NAD+ for robust PARP1 activation and BER complex formation in response to DNA damage. However, the overproduction of 2-hydroxyglutarate, an oncometabolite produced by the IDH1 R132H mutant protein, suppresses BER capacity by reducing Polβ protein levels. This defines a novel mechanism by which the IDH1 mutation in gliomas confers cellular sensitivity to alkylating agents and to inhibitors of the poly-ADP-ribose glycohydrolase, PARG.

The base excision repair (BER) pathway is a precise and versatile mechanism of DNA repair that is initiated by DNA glycosylases. Endonuclease VIII-like 1 (NEIL1) is a bifunctional glycosylase/abasic site (AP) lyase that excises a damaged base and subsequently cleaves the phosphodiester backbone. NEIL1 is able to recognize and hydrolyze a broad range of oxidatively-induced base lesions and substituted ring-fragmented guanines, including aflatoxin-induced 8,9-dihydro-8-(2,6-diamino-4-oxo-3,4-dihydropyrimid-5-yl-formamido)-9-hydroxyaflatoxin B1 (AFB1-FapyGua). Due to NEIL1\'s protective role against these and other pro-mutagenic lesions, it was hypothesized that naturally occurring single nucleotide polymorphic (SNP) variants of NEIL1 could increase human risk for aflatoxin-induced hepatocellular carcinoma (HCC). Given that populations in South Asia experience high levels of dietary aflatoxin exposures and hepatitis B viral infections that induce oxidative stress, investigations on SNP variants of NEIL1 that occur in this region may have clinical implications. In this study, the most common South Asian variants of NEIL1 were expressed, purified, and functionally characterized. All tested variants exhibited activities and substrate specificities similar to wild type (wt)-NEIL1 on high-molecular weight DNA containing an array of oxidatively-induced base lesions. On short oligodeoxynucleotides (17-mers) containing either a site-specific apurinic/apyrimidinic (AP) site, thymine glycol (ThyGly), or AFB1-FapyGua, P206L-NEIL1 was catalytically comparable to wt-NEIL1, while the activities of NEIL1 variants Q67K and T278I on these substrates were ≈2-fold reduced. Variant T103A had a greatly diminished ability to bind to 17-mer DNAs, limiting the subsequent glycosylase and lyase reactions. Consistent with this observation, the rate of excision by T103A on 17-mer oligodeoxynucleotides containing ThyGly or AFB1-FapyGua could not be measured. However, the ability of T103A to excise ThyGly was improved on longer oligodeoxynucleotides (51-mers), with ≈7-fold reduced activity compared to wt-NEIL1. Our studies suggest that NEIL1 variant T103A may present a pathogenic phenotype that is limited in damage recognition, potentially increasing human risk for HCC.

BACKGROUND: High mobility group box 2 (HMGB2) is considered as a biomarker of poor prognosis in various cancers.This study aims to investigate the effect and mechanism of HMGB2 in gliomas.
METHODS: With the glioma related on-line and our local hospital databases, the expression differences of HMGB2,Kaplan-Meier survival analysis and COX regression analysis were performed.The correlation analysis between the clinicopathological features and imaging parameters with the HMGB2 expression had been done. Then GSEA and PPI networks were carried out to find out the most significant pathway. The pathway inhibitor was applied to verify HMGB2\'s participation. CCK8,EDU assays,γ-H2AX immunofluorescence staining and colony formation assay were conducted to observe effects on glioma cells.
RESULTS: Available datasets showed that HMGB2 was highly expressed in glioma and patients with high expression of HMGB2 had poorer prognosis and molecular characteristics. Protein level evidence of western blot and immunohistochemistry from our center supported the conclusions above. Analysis on imaging features suggested that HMGB2 expression level had an inverse association with ADCmean but positively with the thickness of enhancing margin. Results from GSEA and PPI network analysis exhibited that HMGB2 was involved in base excision repair (BER) signaling pathway. Experimental evidence demonstrated that the overexpression of HMGB2 promoted the proliferation of glioma cells and enhanced the radio-resistance.
CONCLUSIONS: HMGB2 could promote glioma development and enhance the radioresistance of glioma cells, potentially related to the BER pathway, suggesting it may serve as an underlying biomarker for patients with glioma.

Base excision repair (BER) requires a tight coordination between the repair enzymes through protein-protein interactions and involves gap filling by DNA polymerase (pol) β and subsequent nick sealing by DNA ligase (LIG) 1 or LIGIIIα at the downstream steps. Apurinic/apyrimidinic-endonuclease 1 (APE1), by its exonuclease activity, proofreads 3\' mismatches incorporated by polβ during BER. We previously reported that the interruptions in the functional interplay between polβ and the BER ligases result in faulty repair events. Yet, how the protein interactions of LIG1 and LIGIIIα could affect the repair pathway coordination during nick sealing at the final steps remains unknown. Here, we demonstrate that LIGIIIα interacts more tightly with polβ and APE1 than LIG1, and the N-terminal noncatalytic region of LIG1 as well as the catalytic core and BRCT domain of LIGIIIα mediate interactions with both proteins. Our results demonstrated less efficient nick sealing of polβ nucleotide insertion products in the absence of LIGIIIα zinc-finger domain and LIG1 N-terminal region. Furthermore, we showed a coordination between APE1 and LIG1/LIGIIIα during the removal of 3\' mismatches from the nick repair intermediate on which both BER ligases can seal noncanonical ends or gap repair intermediate leading to products of single deletion mutagenesis. Overall results demonstrate the importance of functional coordination from gap filling by polβ coupled to nick sealing by LIG1/LIGIIIα in the presence of proofreading by APE1, which is mainly governed by protein-protein interactions and protein-DNA intermediate communications, to maintain repair efficiency at the downstream steps of the BER pathway.

UNASSIGNED: Head and neck squamous carcinoma (HNSCC) is caused by different exogenous risk factors including smoking cigarettes, alcohol consumption, and HPV infection. Base excision repair (BER) is the frontline to repair oxidative DNA damage, which is initiated by the DNA N-glycosylase proteins (OGG1) and other BER factors including DNA polymerase β (POLB).
UNASSIGNED: Explore whether BER genes\' (OGG1, POLB) overexpression in HNSCC alters genomic integrity, immunogenicity, and its role in prognostic value.
UNASSIGNED: RNA sequencing (RNA-Seq) and clinical information (age, gender, histological grade, survival status, and stage) of 530 patients of HNSCC were retrieved from the Cancer Genome Atlas. Patients\' data are categorized HPV positive or negative to analyze the tumor data including the tumor stage, POLB, and OGG1 gene expression.
UNASSIGNED: RNA-Seq of HNSCC data retrieved and mutation count and aneuploidy score were compared using an unpaired t-test. The TIMER algorithm was used to calculate the tumor abundance of six infiltrating immune cells (CD4+ T cells, CD8+ T cells, B cells, neutrophils, macrophages, and dendritic cells) based on RNA-Seq expression profile data. The correlation between the POLB, OGG1, and immune cells was calculated by Spearman correlation analysis using TIMER 2.0.
UNASSIGNED: Our data analysis reveals that BER genes frequently overexpressed in HNSCC tumors and increase mutation count. In addition, OGG1 and POLB overexpression are associated with low infiltration of immune cells, low immune checkpoint gene expression (PD-1, cytotoxic T-lymphocyte antigen 4, program death ligand 1, and program death ligand 2), and innate immune signaling genes. Furthermore, dysregulated BER factors in Human papillomavirus (HPV) positive tumors had better overall survival.
UNASSIGNED: Our analysis suggests that dysregulation of the BER genes panel might be a potential prognosis marker and/or an attractive target for an immune checkpoint blockade in HNSCC cancers. However, our observation still requires further experimental-based scientific validation studies.

DNA repair pathways play a critical role in genome stability, but in eukaryotic cells, they must operate to repair DNA lesions in the compact and tangled environment of chromatin. Previous studies have shown that the packaging of DNA into nucleosomes, which form the basic building block of chromatin, has a profound impact on DNA repair. In this review, we discuss the principles and mechanisms governing DNA repair in chromatin. We focus on the role of histone post-translational modifications (PTMs) in repair, as well as the molecular mechanisms by which histone mutants affect cellular sensitivity to DNA damage agents and repair activity in chromatin. Importantly, these mechanisms are thought to significantly impact somatic mutation rates in human cancers and potentially contribute to carcinogenesis and other human diseases. For example, a number of the histone mutants studied primarily in yeast have been identified as candidate oncohistone mutations in different cancers. This review highlights these connections and discusses the potential importance of DNA repair in chromatin to human health.

Endonuclease VIII-like 3 (NEIL3) is a versatile DNA glycosylase that repairs a diverse array of chemical modifications to DNA. Unlike other glycosylases, NEIL3 has a preference for lesions within single-strand DNA and at single/double-strand DNA junctions. Beyond its canonical role in base excision repair of oxidized DNA, NEIL3 initiates replication-dependent interstrand DNA crosslink repair as an alternative to the Fanconi Anemia pathway. This review outlines our current understanding of NEIL3\'s biological functions, role in disease, and three-dimensional structure as it pertains to substrate specificity and catalytic mechanism.

Gene knock-out (KO) mouse models for DNA polymerase beta (Polβ) revealed that loss of Polβ leads to neonatal lethality, highlighting the critical organismic role for this DNA polymerase. While biochemical analysis and gene KO cell lines have confirmed its biochemical role in base excision repair and in TET-mediated demethylation, more long-lived mouse models continue to be developed to further define its organismic role. The Polb-KO mouse was the first of the Cre-mediated tissue-specific KO mouse models. This technology was exploited to investigate roles for Polβ in V(D)J recombination (variable-diversity-joining rearrangement), DNA demethylation, gene complementation, SPO11-induced DNA double-strand break repair, germ cell genome stability, as well as neuronal differentiation, susceptibility to genotoxin-induced DNA damage, and cancer onset. The revolution in knock-in (KI) mouse models was made possible by CRISPR/cas9-mediated gene editing directly in C57BL/6 zygotes. This technology has helped identify phenotypes associated with germline or somatic mutants of Polβ. Such KI mouse models have helped uncover the importance of key Polβ active site residues or specific Polβ enzyme activities, such as the PolbY265C mouse that develops lupus symptoms. More recently, we have used this KI technology to mutate the Polb gene with two codon changes, yielding the PolbL301R/V303R mouse. In this KI mouse model, the expressed Polβ protein cannot bind to its obligate heterodimer partner, Xrcc1. Although the expressed mutant Polβ protein is proteolytically unstable and defective in recruitment to sites of DNA damage, the homozygous PolbL301R/V303R mouse is viable and fertile, yet small in stature. We expect that this and additional targeted mouse models under development are poised to reveal new biological and organismic roles for Polβ.