OpenTree: A Python Package for Accessing and Analyzing Data from the Open Tree of Life.

The Open Tree of Life project constructs a comprehensive, dynamic, and digitally available tree of life by synthesizing published phylogenetic trees along with taxonomic data. Open Tree of Life provides web-service application programming interfaces (APIs) to make the tree estimate, unified taxonomy, and input phylogenetic data available to anyone. Here, we describe the Python package opentree, which provides a user friendly Python wrapper for these APIs and a set of scripts and tutorials for straightforward downstream data analyses. We demonstrate the utility of these tools by generating an estimate of the phylogenetic relationships of all bird families, and by capturing a phylogenetic estimate for all taxa observed at the University of California Merced Vernal Pools and Grassland Reserve.[Evolution; open science; phylogenetics; Python; taxonomy.].

Impact of Oxidative DNA Damage and the Role of DNA Glycosylases in Neurological Dysfunction.

The human brain requires a high rate of oxygen consumption to perform intense metabolic activities, accounting for 20% of total body oxygen consumption. This high oxygen uptake results in the generation of free radicals, including reactive oxygen species (ROS), which, at physiological levels, are beneficial to the proper functioning of fundamental cellular processes. At supraphysiological levels, however, ROS and associated lesions cause detrimental effects in brain cells, commonly observed in several neurodegenerative disorders. In this review, we focus on the impact of oxidative DNA base lesions and the role of DNA glycosylase enzymes repairing these lesions on brain function and disease. Furthermore, we discuss the role of DNA base oxidation as an epigenetic mechanism involved in brain diseases, as well as potential roles of DNA glycosylases in different epigenetic contexts. We provide a detailed overview of the impact of DNA glycosylases on brain metabolism, cognition, inflammation, tissue loss and regeneration, and age-related neurodegenerative diseases based on evidence collected from animal and human models lacking these enzymes, as well as post-mortem studies on patients with neurological disorders.

Loss of Neil3, the major DNA glycosylase activity for removal of hydantoins in single stranded DNA, reduces cellular proliferation and sensitizes cells to genotoxic stress.

7,8-Dihydro-8-oxoguanine (8-oxoG) is one of the most common oxidative base lesions in normal tissues induced by a variety of endogenous and exogenous agents. Hydantoins are products of 8-oxoG oxidation and as 8-oxoG, they have been shown to be mutagenic lesions. Oxidative DNA damage has been implicated in the etiology of various age-associated pathologies, such as cancer, cardiovascular diseases, arthritis, and several neurodegenerative diseases. The mammalian endonuclease VIII-like 3 (Neil3) is one of the four DNA glycosylases found to recognize and remove hydantoins in the first step of base excision repair (BER) pathway. We have generated mice lacking Neil3 and by using total cell extracts we demonstrate that Neil3 is the main DNA glycosylase that incises hydantoins in single stranded DNA in tissues. Using the neurosphere culture system as a model to study neural stem/progenitor (NSPC) cells we found that lack of Neil3 impaired self renewal but did not affect differentiation capacity. Proliferation was also reduced in mouse embryonic fibroblasts (MEFs) derived from Neil3(-/-) embryos and these cells were sensitive to both the oxidative toxicant paraquat and interstrand cross-link (ICL)-inducing agent cisplatin. Our data support the involvement of Neil3 in removal of replication blocks in proliferating cells.

Endonuclease VIII-like 3 (Neil3) DNA glycosylase promotes neurogenesis induced by hypoxia-ischemia.

Neural stem/progenitor cell proliferation and differentiation are required to replace damaged neurons and regain brain function after hypoxic-ischemic events. DNA base lesions accumulating during hypoxic-ischemic stress are removed by DNA glycosylases in the base-excision repair pathway to prevent cytotoxicity and mutagenesis. Expression of the DNA glycosylase endonuclease VIII-like 3 (Neil3) is confined to regenerative subregions in the embryonic and perinatal brains. Here we show profound neuropathology in Neil3-knockout mice characterized by a reduced number of microglia and loss of proliferating neuronal progenitors in the striatum after hypoxia-ischemia. In vitro expansion of Neil3-deficient neural stem/progenitor cells revealed an inability to augment neurogenesis and a reduced capacity to repair for oxidative base lesions in single-stranded DNA. We propose that Neil3 exercises a highly specialized function through accurate molecular repair of DNA in rapidly proliferating cells.

Enhanced glutathione levels confer resistance to apoptotic and ferroptotic programmed cell death in NEIL DNA glycosylase deficient HAP1 cells.

The NTHL1 and NEIL1-3 DNA glycosylases are major enzymes in the removal of oxidative DNA base lesions, via the base excision repair (BER) pathway. It is expected that lack of these DNA glycosylases activities would render cells vulnerable to oxidative stress, promoting cell death. Intriguingly, we found that single, double, triple, and quadruple DNA glycosylase knockout HAP1 cells are, however, more resistant to oxidative stress caused by genotoxic agents than wild type cells. Furthermore, glutathione depletion in NEIL deficient cells further enhances resistance to cell death induced via apoptosis and ferroptosis. Finally, we observed higher basal level of glutathione and differential expression of NRF2-regulated genes associated with glutathione homeostasis in the NEIL triple KO cells. We propose that lack of NEIL DNA glycosylases causes aberrant transcription and subsequent errors in protein synthesis. This leads to increased endoplasmic reticulum stress and proteotoxic stress. To counteract the elevated intracellular stress, an adaptive response mediated by increased glutathione basal levels, rises in these cells. This study reveals an unforeseen role of NEIL glycosylases in regulation of resistance to oxidative stress, suggesting that modulation of NEIL glycosylase activities is a potential approach to improve the efficacy of e.g. anti-inflammatory therapies.

Depletion of the m1A writer TRMT6/TRMT61A reduces proliferation and resistance against cellular stress in bladder cancer.

Background: Bladder cancer (BLCA) is a common and deadly disease that results in a reduced quality of life for the patients and a significant economic burden on society. A better understanding of tumorigenesis is needed to improve clinical outcomes. Recent evidence places the RNA modification m1A and its regulatory proteins TRMT6/TRMT61A and ALKBH3 in BLCA pathogenesis. Methods: TRMT6/TRMT61A, ALKBH1, and ALKBH3 expression was examined in human BLCA cell lines and a normal urinary tract epithelium cell line through qRT-PCR and western blot analysis. Prestoblue Cell Viability Reagent, wound-healing assay, and live-cell imaging-based cell displacement analysis, were conducted to assess proliferation, migration, and displacement of this BLCA cell line panel. Cell survival was assessed after inducing cellular stress and activating the unfolded protein response (UPR) with tunicamycin. Moreover, siRNA-mediated gene silencing in two BLCA cell lines (5637 and HT1197) was conducted to investigate the biological roles of TRMT6/TRMT61A. Results: Heterogeneous morphology, proliferation, displacement, tunicamycin sensitivity, and expression levels of m1A regulators were observed among the panel of cell lines examined. In general, TRMT61A expression was increased in BLCA cell lines when compared to SV-HUC-1. Depletion of TRMT6/TRMT61A reduced proliferation capacity in both 5637 and HT1197 cell lines. The average cell displacement of 5637 was also reduced upon TRMT6/TRMT61A depletion. Interestingly, TRMT6/TRMT61A depletion decreased mRNA expression of targets associated with the ATF6-branch of the UPR in 5637 but not in HT1197. Moreover, cell survival after induction of cellular stress was compromised after TRMT6/TRMT61A knockdown in 5637 but not in HT1197 cells. Conclusion: The findings suggest that TRMT6/TRMT61A plays an oncogenic role in BLCA and is involved in desensitizing BLCA cells against cellular stress. Further investigation into the regulation of TRMT6/TRMT61A expression and its impact on cellular stress tolerance may provide insights for future BLCA treatment.

NEIL1 and NEIL2 DNA glycosylases modulate anxiety and learning in a cooperative manner in mice.

Oxidative DNA damage in the brain has been implicated in neurodegeneration and cognitive decline. DNA glycosylases initiate base excision repair (BER), the main pathway for oxidative DNA base lesion repair. NEIL1 and NEIL3 DNA glycosylases affect cognition in mice, while the role of NEIL2 remains unclear. Here, we investigate the impact of NEIL2 and its potential overlap with NEIL1 on behavior in knockout mouse models. Neil1-/-Neil2-/- mice display hyperactivity, reduced anxiety and improved learning. Hippocampal oxidative DNA base lesion levels are comparable between genotypes and no mutator phenotype is found. Thus, impaired canonical repair is not likely to explain the altered behavior. Electrophysiology suggests reduced axonal activation in the hippocampal CA1 region in Neil1-/-Neil2-/- mice and lack of NEIL1 and NEIL2 causes dysregulation of genes in CA1 relevant for synaptic function. We postulate a cooperative function of NEIL1 and NEIL2 in genome regulation, beyond canonical BER, modulating behavior in mice.

Modulation of DNA glycosylase activities in mesenchymal stem cells.

Adipose-tissue derived mesenchymal stem cells (AT-MSCs) are a promising tool for use in cell-based therapies. However, in vitro expansion is required to obtain clinically relevant cell numbers, and this might increase the chance of genomic instability. DNA repair is crucial for maintaining DNA integrity. Here we have compared the initial step of base excision repair in uncultured and cultured AT-MSCs by analysis of base removal activities and expression levels of relevant DNA glycosylases. Uracil, 5-hydroxyuracil and ethenoadenine removal activities were upregulated in cultured cells compared to uncultured cells. In contrast, both the 8-oxo-7,8-dihydroguanine (8-oxoG) removal activity and the concentration of 8-oxoG bases in the DNA were reduced in the cultured cells. Gene expression analysis showed no substantial changes in mRNA expression. The glycosylase activities remained stable through at least 12 passages, suggesting that DNA repair is proficient through the period required for in vitro expansion of AT-MSCs to clinically relevant numbers.

OXR1A, a Coactivator of PRMT5 Regulating Histone Arginine Methylation.

Oxidation resistance gene 1 (OXR1) protects cells against oxidative stress. We find that male mice with brain-specific isoform A knockout (Oxr1A-/-) develop fatty liver. RNA sequencing of male Oxr1A-/- liver indicates decreased growth hormone (GH) signaling, which is known to affect liver metabolism. Indeed, Gh expression is reduced in male mice Oxr1A-/- pituitary gland and in rat Oxr1A-/- pituitary adenoma cell-line GH3. Oxr1A-/- male mice show reduced fasting-blood GH levels. Pull-down and proximity ligation assays reveal that OXR1A is associated with arginine methyl transferase PRMT5. OXR1A-depleted GH3 cells show reduced symmetrical dimethylation of histone H3 arginine 2 (H3R2me2s), a product of PRMT5 catalyzed methylation, and chromatin immunoprecipitation (ChIP) of H3R2me2s shows reduced Gh promoter enrichment. Finally, we demonstrate with purified proteins that OXR1A stimulates PRMT5/MEP50-catalyzed H3R2me2s. Our data suggest that OXR1A is a coactivator of PRMT5, regulating histone arginine methylation and thereby GH production within the pituitary gland.

The carotenoid beta-cryptoxanthin stimulates the repair of DNA oxidation damage in addition to acting as an antioxidant in human cells.

The role of dietary antioxidants in human health remains controversial. Fruits and vegetables in the diet are associated with lower rates of chronic disease, and this is often attributed to their content of antioxidants, and a resulting protection against oxidative stress. However, large-scale human trials with antioxidant supplements have shown, if anything, an increase in mortality. We have investigated the biological properties of beta-cryptoxanthin, a common carotenoid, in cell culture model systems, using the comet assay to measure DNA damage. At low concentrations, close to those found in plasma, beta-cryptoxanthin does not itself cause damage, but protects transformed human cells (HeLa and Caco-2) from damage induced by H(2)O(2) or by visible light in the presence of a photosensitizer. In addition, it has a striking effect on DNA repair, measured in different ways. Incubation of H(2)O(2)-treated cells with beta-cryptoxanthin led to a doubling of the rate of rejoining of strand breaks and had a similar effect on the rate of removal of oxidized purines by base excision repair. The latter effect was confirmed with an in vitro assay: cells were incubated with or without beta-cryptoxanthin before preparing an extract, which was then incubated with substrate DNA containing 8-oxo-7,8-dihydroguanine; incision was more rapid with the extract prepared from carotenoid-preincubated cells. No significant increases were seen in protein content of human 8-oxoguanine DNA glycosylase 1 or apurinic endonuclease 1. The apparent cancer-preventive effects of dietary carotenoids may depend on the enhancement of DNA repair as well as antioxidant protection against damage.

Widespread distribution of DNA glycosylases removing oxidative DNA lesions in human and rodent brains.

High metabolic activity and low levels of antioxidant enzymes make neurons particularly prone to damage by reactive oxygen species. Thus, repair of oxidative DNA damage is essential for normal brain function. Base excision repair is the major pathway for repair of oxidative DNA damage, and is initiated by DNA glycosylases recognizing and removing the damaged base. In mammalian cells at least five different DNA glycosylases with overlapping substrate specificity, NEIL1, NEIL2, NEIL3, OGG1 and NTH1, remove oxidative DNA base lesions. Here we report mRNA expression and distribution of these five DNA glycosylases in human and rodent brains using in situ hybridization and Northern blotting supported by glycosylase activity assays. NEIL1, NEIL2, OGG1 and NTH1 showed widespread expression at all ages. In situ hybridization studies in mouse brain showed that expression of mNeil1 increased with age. In newborn mouse brain, mNeil3 revealed a discrete expression pattern in brain regions known to harbour stem cell populations, i.e., the subventricular zone, the rostral migratory stream, and the hilar region of the hippocampal formation. Expression of mNeil3 decreased with age, and in old mice brains could be detected only in layer V of neocortex. MNth1 was constitutively expressed during lifespan. In Northern blots, mOgg1 expression showed a transient decrease followed by an increase after 8 weeks of age. Assays for faPy DNA glycosylase activity revealed increased activity level with age in all brain regions analyzed. The widespread but differential expression of the DNA glycosylases recognizing oxidative base lesions suggests distinct and age dependent roles of these enzymes in genome maintenance in brain. The distribution of mNeil3 is particularly intriguing and points to a specific role of this enzyme in stem cell differentiation.

Base excision repair activities in organotypic hippocampal slice cultures exposed to oxygen and glucose deprivation.

The capacity for DNA repair is likely to be one of the factors that determine the vulnerability of neurons to ischemic stress and may influence the pathological outcome of stroke. In this report, initiation of base excision repair (BER) was assessed by analysis of enzyme activity and gene expression level of DNA glycosylases and AP-endonucleases in rat organotypic hippocampal slice cultures exposed to oxygen and glucose deprivation (OGD) - an in vitro model of stroke. Under basal conditions, AP-endonuclease activity and base removal of ethenoadenine and 8-oxoguanine (8-oxoG) were higher (by approximately 20-35 %) in CA3/fascia dentata (FD) than in CA1. Base removal of uracil did not differ between the two hippocampal regions, while removal of 5-hydroxyuracil (5-OHU) was slightly less efficient in CA3/FD than in CA1. Analyses performed immediately after 30 min of OGD revealed a decreased AP-endonuclease activity (by approximately 20%) in CA1 as well as CA3/FD, and an increased ethenoadenine activity (by approximately 25%) in CA1. Activities for 8-oxoG, 5-OHU and uracil showed no significant changes at this time point. At 8h after OGD, none of the enzyme activities differed from control values. Real-time RT-PCR showed that transcription of DNA glycosylases, including Ogg1, Nth1, Ung, Aag, Neil1 and Neil2 were not changed in response to OGD treatment (t=0 h). The hippocampal expression of Neil2 was low compared with the other DNA glycosylases. These data indicate that CA1 has a lower capacity than CA3/FD for removal of base lesions under basal conditions. The relatively low capacity for BER in basal conditions and the apparent failure to upregulate repair of oxidative damage after OGD might contribute to the high vulnerability of CA1 to ischemic injury.

Expression patterns of Neil3 during embryonic brain development and neoplasia.

BACKGROUND: The base excision repair pathway is responsible for repairing small DNA base lesions caused by endogenous and exogenous damaging agents. Repair is initiated by DNA glycosylases that recognize and remove the lesions. NEIL3 is one of 11 mammalian DNA glycosylases identified to date and it was discovered on the basis of sequence homology to the E. coli Fpg and Nei glycosylases. Difficulties in purifying the protein have limited its biochemical characterization and in contrast to the other glycosylases, its function remains unclear. RESULTS: In this study we describe the expression pattern of Neil3 during mouse embryonic development with special focus on brain development. We have also looked at the expression of NEIL3 in several normal and tumor tissues. Quantitative real-time PCR and in situ hybridization revealed that Neil3 was highly expressed at embryonic days 12-13, when neurogenesis starts. The expression decreased during development and in the adult brain,Neil3 could not be detected in any of the brain areas examined by quantitative real-time PCR. During embryogenesis and in newborn mice specific expression was observed in areas known to harbour neural stem and progenitor cells such as the subventricular zone and the dentate gyrus. Finally, NEIL3 expression was higher in tumors compared to normal tissues, except for testis and pancreas. CONCLUSION: Our findings indicate that mammalian NEIL3 is specifically expressed in brain areas where neurogenesis takes place during development and that its expression is tightly regulated both temporally and spatially. In addition, NEIL3 seems to be upregulated in tumor tissues compared to normal tissues. Altogether, mammalian NEIL3 seems to be highly expressed in cells with high proliferative potential.

Expression and purification of NEIL3, a human DNA glycosylase homolog.

The base excision repair (BER) pathway is mainly responsible for the repair of a vast number of non-bulky lesions produced by alkylation, oxidation or deamination of bases. DNA glycosylases are the key enzymes that recognize damaged bases and initiate BER by catalyzing the cleavage of the N-glycosylic bond between the base and the sugar. Many of the mammalian DNA glycosylases have been identified by a combination of biochemical and bioinformatics analysis. Thus, a mammalian family of three proteins (NEIL1, NEIL2 and NEIL3) that showed homology to the Escherichia coli Fpg/Nei DNA glycosylases was identified. Two of the proteins, NEIL1 and NEIL2 have been thoroughly characterized and shown to initiate BER of a diverse number of oxidized lesions. However, much less is known about NEIL3. The biochemical properties of NEIL3 have not been elucidated. This is mainly due to the difficulty in the expression and purification of NEIL3. Here, we describe the expression and partial purification of full-length human NEIL3 and the expression, purification and characterization of a truncated human core-NEIL3 (amino acids 1-301) that contains the complete E. coli Fpg/Nei-like domain but lacks the C-terminal region.

Up-regulation of myocardial DNA base excision repair activities in experimental heart failure.

Base excision repair (BER) is the major pathway to counteract the genotoxic effect of endogenous DNA damaging agents. The present study investigated the enzymatic activities and gene transcription of DNA glycosylases initiating BER in an experimental heart failure (HF) model. Rats were subjected to myocardial infarction or sham-operated. Twenty-eight days after surgical intervention cell-free protein extracts, total RNA and genomic DNA were isolated to analyze DNA glycosylase and AP-endonuclease activities, transcript levels of DNA glycosylases and accumulation of oxidative DNA damage. The capacity to remove major oxidation products (e.g., formamidopyrimidine and 5-hydroxycytosine) was significantly increased in the border zone of infarcted area, while the capacity to remove the highly mutagenic 8-oxoguanine residue was enhanced both in non-infarcted and infarcted areas of left ventricle (LV). DNA glycosylase activities towards 3-methyladenine and uracil were up-regulated in infarcted and non-infarcted areas of LV, indicating that generation of alkylated and deaminated base lesions on DNA increase during HF. Finally, we found no difference in accumulation of oxidative DNA damage in myocardial tissue between rats with post-myocardial infarction and sham-operated rats. This up-regulation of activities, initiating the BER pathway, could play an important role during HF by counteracting the effect of genotoxic stress, structural damage of tissue and myocardial remodeling.

Human NEIL1 localizes with the centrosomes and condensed chromosomes during mitosis.

The DNA glycosylase hNEIL1 initiates base excision repair (BER) of a number of oxidized purines and pyrimidines in cellular DNA and is one of three mammalian orthologs of the Escherichia coli Nei/Fpg enzymes. Human NEIL1 has been purified and extensively characterized biochemically, however, not much is known about its intracellular distribution. In the present work, we have studied the cellular localization of hNEIL1 using both antibodies raised against the full-length recombinant protein and a stable HeLa cell line expressing hNEIL1 fused N-terminal to EGFP. The results presented reveal an intricate mitotic distribution of hNEIL1. Centrosomal localization of hNEIL1 was observed when mitotic HeLa cells were immunostained with hNEIL1 antibodies. This localization was confirmed when Western blots of isolated centrosomes from stably expressing hNEIL1-EGFP HeLa cells were probed with GFP or hNEIL1 antibodies, even though a fluorescent signal could not be detected in the centrosomes of these cells. Human NEIL1 was also shown to be associated with mitotic condensed chromosomes. Notably, the interaction of hNEIL1 with condensed chromatin was disrupted when cells were fixed with chemical fixatives that are regularly used in immunodetection techniques.

The capacity to remove 8-oxoG is enhanced in newborn neural stem/progenitor cells and decreases in juvenile mice and upon cell differentiation.

In mammalian cells, 8-oxoguanine DNA glycosylase-1 (OGG1) is the main DNA glycosylase for the removal of 8-oxoguanine (8-oxoG). 8-oxoG, one of the most common products of the oxidative attack of DNA, is a premutagenic lesion that accumulates spontaneously at high frequencies in the genome. In this study, Ogg1 mRNA expression was detected throughout embryonic development in mice. In situ hybridization showed that in the neonatal brain, Ogg1 expression was detected in a distinct layer of cells in the medial wall of the lateral ventricle, which may correspond to ependymal cells, and in some scattered cells in the subventricular zone (SVZ), a brain region rich in neural stem/progenitor cells. Using neurospheres as a model for the study of neural stem/progenitor cells, we found that both the expression and activity of Ogg1 were high in neurospheres derived from newborn mice and decreased in adults and upon induction of cell differentiation. Furthermore, Ogg1 was shown to be the major DNA glycosylase initiating 8-oxoG repair in neurospheres. Our results strongly indicate that enhanced DNA repair capacity is an important mechanism by which neural stem/progenitor cells maintain their genome.

Dynamic relocalization of hOGG1 during the cell cycle is disrupted in cells harbouring the hOGG1-Cys326 polymorphic variant.

Numerous lines of evidence support the role of oxidative stress in different types of cancer. A major DNA lesion, 8-oxo-7,8-dihydroguanine (8-oxoG), is formed by reactive oxygen species in the genome under physiological conditions. 8-OxoG is strongly mutagenic, generating G.C-->T.A transversions, a frequent somatic mutation in cancers. hOGG1 was cloned as a gene encoding a DNA glycosylase that specifically recognizes and removes 8-oxoG from 8-oxoG:C base pairs and suppresses G.C-->T.A transversions. In this study, we investigated the subcellular localization and expression of hOGG1 during the cell cycle. Northern blots showed cell-cycle-dependent mRNA expression of the two major hOGG1 isoforms. By using a cell line constitutively expressing hOGG1 fused to enhanced green fluorescence protein (EGFP), we observed a dynamic relocalization of EGFP-hOGG1 to the nucleoli during the S-phase of the cell cycle, and this localization was shown to be linked to transcription. A C/G change that results in an amino acid substitution from serine to cysteine in codon 326 has been reported as a genetic polymorphism and a risk allele for a variety of cancers. We investigated the cellular localization of the corresponding protein, hOGG1-Cys326, fused to EGFP and observed a dramatic effect on its localization that is explained by a change in the phosphorylation status of hOGG1.

No cancer predisposition or increased spontaneous mutation frequencies in NEIL DNA glycosylases-deficient mice.

Base excision repair (BER) is a major pathway for removal of DNA base lesions and maintenance of genomic stability, which is essential in cancer prevention. DNA glycosylases recognize and remove specific lesions in the first step of BER. The existence of a number of these enzymes with overlapping substrate specificities has been thought to be the reason why single knock-out models of individual DNA glycosylases are not cancer prone. In this work we have characterized DNA glycosylases NEIL1 and NEIL2 (Neil1 -/- /Neil2 -/-) double and NEIL1, NEIL2 and NEIL3 (Neil1 -/- /Neil2 -/- /Neil3 -/-) triple knock-out mouse models. Unexpectedly, our results show that these mice are not prone to cancer and have no elevated mutation frequencies under normal physiological conditions. Moreover, telomere length is not affected and there was no accumulation of oxidative DNA damage compared to wild-type mice. These results strengthen the hypothesis that the NEIL enzymes are not simply back-up enzymes for each other but enzymes that have distinct functions beyond canonical repair.

NEIL3-Dependent Regulation of Cardiac Fibroblast Proliferation Prevents Myocardial Rupture.

Myocardial infarction (MI) triggers a reparative response involving fibroblast proliferation and differentiation driving extracellular matrix modulation necessary to form a stabilizing scar. Recently, it was shown that a genetic variant of the base excision repair enzyme NEIL3 was associated with increased risk of MI in humans. Here, we report elevated myocardial NEIL3 expression in heart failure patients and marked myocardial upregulation of Neil3 after MI in mice, especially in a fibroblast-enriched cell fraction. Neil3-/- mice show increased mortality after MI caused by myocardial rupture. Genome-wide analysis of 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) reveals changes in the cardiac epigenome, including in genes related to the post-MI transcriptional response. Differentially methylated genes are enriched in pathways related to proliferation and myofibroblast differentiation. Accordingly, Neil3-/- ruptured hearts show increased proliferation of fibroblasts and myofibroblasts. We propose that NEIL3-dependent modulation of DNA methylation regulates cardiac fibroblast proliferation and thereby affects extracellular matrix modulation after MI.