Arne Klungland

• Lier, Silje; Markusson, Sara Bachmann; Kocijančič, Anja; Narum, Martine; Lund, Solveig Osnes & Böllering, Bianka [Show all 30 contributors for this article] (2026). CDK12/CDK13 inhibition disrupts transcriptional elongation and replication fork progression in glioblastoma. EMBO Molecular Medicine. ISSN 1757-4676. 18, p. 1592–1624. doi: 10.1038/s44321-026-00393-w. Full text in Research Archive Show summary

  Glioblastomas are the most prevalent and aggressive malignant brain tumors, characterized by hypertranscription and dependence on neurodevelopmental transcription factors. The transcriptional cycle is regulated by phosphorylation of the C-terminal domain (CTD) of RNA polymerase II (RNAPII) by transcriptional cyclin-dependent kinases (tCDKs), including CDK7, CDK9, CDK12, and CDK13. Here we find that glioblastoma stem cells (GSCs) are selectively sensitive to CDK12/CDK13 inhibition, whereas CDK7 and CDK9 inhibition cause non-specific cytotoxicity. This selective targeting halts GSC and organoid proliferation, curtails GSC invasion and suppresses tumor growth in a xenograft mouse model. In GSCs, CDK12/CDK13 inhibition leads to a rapid and genome-wide loss of serine-2 phosphorylation (pSer2) of the RNAPII CTD, abolishing transcriptional elongation and a transcriptional program sustained by key neurodevelopmental transcription factors. CDK12/CDK13 inhibition unexpectedly arrests DNA replication and replication fork progression in a manner distinct from the effect of inhibiting other tCDKs. This dramatic arrest precedes DNA damage response activation and cell cycle arrest, directly linking RNAPII elongation to replication fork dynamics and revealing a previously unrecognized dependence of DNA replication on CDK12/CDK13-RNAPII regulation.

• Fosslie, Madeleine Lystad; Ilaslan, Erkut; Skuland, Trine; Choudalakis, Michel; Søegaard, Mirra Louise Cicilie & Halliwell, Jason Alexander [Show all 23 contributors for this article] (2025). Major waves of H2A.Z incorporation during mouse oogenesis and preimplantation embryo development. Nature Communications. 17(1). doi: 10.1038/s41467-025-66919-x. Full text in Research Archive Show summary

  Abstract Epigenomes of mammalian oocytes and embryos undergo major transitions essential for successful development. Here, we provide genome-wide maps of histone variant H2A.Z during twelve stages of mouse oogenesis and preimplantation embryo development and relate it to histone marks and genomic features. This revealed that major waves of H2A.Z incorporation occur early in growing oocytes, forming distinct patterns of maternal, embryonic, and persistent H2A.Z enrichment. Late maternal enrichment is inherited by the zygote and precedes reduced formation of lamina associated domains and early replication in the maternal genome of 2-cell embryos. Persistent H2A.Z enrichment is strongly associated with CpG islands and H3K4me3 near transcription start sites of active genes, but thousands of maternal and embryonic H2A.Z incorporation sites exist elsewhere, frequently at transposable elements. The persisting H2A.Z enrichments across related developmental stages enable preservation of epigenetic information despite major concurrent changes in H3K4me3, H3K27me3, and DNA methylation. Altogether, this advances our understanding of how histone variants contribute to epigenetic reprogramming during mammalian oogenesis and early development.

• Desgarnier, Marie-Catherine Anne Danielle Drigeard; Monshaugen, Ida; Ougland, Rune & Klungland, Arne (2025). The 6-methyladenine erasers ALKBH5 and FTO influence chemotherapy efficiency in bladder cancer cell lines. Annals of Translational Medicine. ISSN 2305-5839. 13(3). doi: 10.21037/atm-25-7. Full text in Research Archive
• Luan, Yan; Zhang, Hanyue; Liu, Yingfei; Zhou, Jing; Li, Wen & Ma, Kaige [Show all 14 contributors for this article] (2025). Inverse and dynamic levels of H3K4me3 and H3K27me3 regulate mouse postnatal dental gyrus development. Cell Death and Differentiation. ISSN 1350-9047. 33, p. 219–235. doi: 10.1038/s41418-025-01563-y. Full text in Research Archive Show summary

  The dentate gyrus (DG), a crucial region of the hippocampus responsible for learning, spatial encoding, and memory formation, undergoes its main development and maturation after birth. Despite its importance, the regulatory mechanisms underlying postnatal DG development remain poorly understood. Thi …

• Li, Yanjiao; Wang, Yunhao; Cengiz, Aylin; Jin, Kangxuan; castroviejo, Blanca Corral & Lin, Xiaolin [Show all 19 contributors for this article] (2025). The RNA m6A landscape during human oocyte-to-embryo transition. EMBO Journal. ISSN 0261-4189. 44(14), p. 4150–4180. doi: 10.1038/s44318-025-00474-5. Full text in Research Archive Show summary

  RNA N6-methyladenosine (m6A, m6A) modification is a critical regulator for a range of physiological processes. However, the dynamic m6A profiles within human preimplantation embryos remain uncharacterized. Here, we present the first RNA m6A landscape of single human oocytes and early embryos. Comparative analyses with mouse data reveal an intriguing divergence during the window of zygotic genome activation. m6A-modified genes are involved in regulation of gene transcription, while unmodified genes are mainly associated with basic metabolic processes. Maternal decay mRNAs exhibit a propensity for m6A modifications, and these genes are targeted by miRNAs. m6A modified genes that are constantly expressed across all stages demonstrate higher translation efficiency. Moreover, we observe frequent m6A enrichment on stage-specifically expressed retrotransposons, particularly within young subfamilies. m6A inhibitor leads to m6A erasure on massive retrotransposons. In summary, this study provides a resource to broaden our understanding about the regulatory roles of m6A during early human embryo development.

• Li, Yanjiao; Wang, Yunhao; Vera-Rodriguez, Maria; Lindeman, Leif Christopher; Skuggen, Linda Ellevog & Rasmussen, Erik Magne Koscielniak [Show all 21 contributors for this article] (2023). Single-cell m6A mapping in vivo using picoMeRIP–seq. Nature Biotechnology. ISSN 1087-0156. doi: 10.1038/s41587-023-01831-7. Full text in Research Archive Show summary

  Abstract Current N 6 -methyladenosine (m 6 A) mapping methods need large amounts of RNA or are limited to cultured cells. Through optimized sample recovery and signal-to-noise ratio, we developed picogram-scale m 6 A RNA immunoprecipitation and sequencing (picoMeRIP–seq) for studying m 6 A in vivo in single cells and scarce cell types using standard laboratory equipment. We benchmark m 6 A mapping on titrations of poly(A) RNA and embryonic stem cells and in single zebrafish zygotes, mouse oocytes and embryos.

• Monshaugen, Ida; Luna, Luisa; Rhodes, Jayden; Kristiansen, Felicia Iselin Svensson; Lång, Anna Ulrika & Bøe, Stig Ove [Show all 10 contributors for this article] (2023). Depletion of the m1A writer TRMT6/TRMT61A reduces proliferation and resistance against cellular stress in bladder cancer. Frontiers in Oncology. 13, p. 1–14. doi: 10.3389/fonc.2023.1334112. Full text in Research Archive Show summary

  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.

• Wang, Yunhao; Li, Yanjiao; Skuland, Trine; Zhou, Chengjie; Li, Aifu & Hashim, Adnan [Show all 13 contributors for this article] (2023). The RNA m6A landscape of mouse oocytes and preimplantation embryos. Nature Structural & Molecular Biology. ISSN 1545-9993. 30(5), p. 703–709. doi: 10.1038/s41594-023-00969-x. Full text in Research Archive Show summary

  Despite the significance of N6-methyladenosine (m6A) in gene regulation, the requirement for large amounts of RNA has hindered m6A profiling in mammalian early embryos. Here we apply low-input methyl RNA immunoprecipitation and sequencing to map m6A in mouse oocytes and preimplantation embryos. We define the landscape of m6A during the maternal-to-zygotic transition, including stage-specifically expressed transcription factors essential for cell fate determination. Both the maternally inherited transcripts to be degraded post fertilization and the zygotically activated genes during zygotic genome activation are widely marked by m6A. In contrast to m6A-marked zygotic ally-activated genes, m6A-marked maternally inherited transcripts have a higher tendency to be targeted by microRNAs. Moreover, RNAs derived from retrotransposons, such as MTA that is maternally expressed and MERVL that is transcriptionally activated at the two-cell stage, are largely marked by m6A. Our results provide a foundation for future studies exploring the regulatory roles of m6A in mammalian early embryonic development.

• Su, Zhangli; Monshaugen, Ida; Klungland, Arne; Ougland, Rune & Dutta, Anindya (2022). Characterization of novel small non-coding RNAs and their modifications in bladder cancer using an updated small RNA-seq workflow. Frontiers in Molecular Biosciences. 9. doi: 10.3389/fmolb.2022.887686. Full text in Research Archive Show summary

  Background: Bladder cancer (BLCA) is one of the most common cancer types worldwide. The disease is responsible for about 200,000 deaths annually, thus improved diagnostics and therapy is needed. A large body of evidence reveal that small RNAs of less than 40 nucleotides may act as tumor suppressors, oncogenes, and disease biomarkers, with a major focus on microRNAs. However, the role of other families of small RNAs is not yet deciphered. Recent results suggest that small RNAs and their modification status, play a role in BLCA development and are promising biomarkers due to their high abundance in the exomes and body fluids (including urine). Moreover, free modified nucleosides have been detected at elevated levels from the urine of BLCA patients. A genome-wide view of small RNAs, and their modifications, will help pinpoint the molecules that could be used as biomarker or has important biology in BLCA development. Methods: BLCA tumor tissue specimens were obtained from 12 patients undergoing transurethral resection of non-muscle invasive papillary urothelial carcinomas. Genome-wide profiling of small RNAs less than 40 bases long was performed by a modified protocol with TGIRT (thermostable group II reverse transcriptase) to identify novel small RNAs and their modification status. Results: Comprehensive analysis identified not only microRNAs. Intriguingly, 57 ± 15% (mean ± S.D.) of sequencing reads mapped to non-microRNA-small RNAs including tRNA-derived fragments (tRFs), ribosomal RNA-derived fragments (rRFs) and YRNA-derived fragments (YRFs). Misincorporation (mismatch) sites identified potential base modification positions on the small RNAs, especially on tRFs, corresponding to m 1 A (N 1 -methyladenosine), m 1 G (N 1 -methylguanosine) and m 2 2 G (N 2 , N 2 -dimethylguanosine). We also detected mismatch sites on rRFs corresponding to known modifications on 28 and 18S rRNA. Conclusion: We found abundant non-microRNA-small RNAs in BLCA tumor samples. Small RNAs, especially tRFs and rRFs, contain modifications that can be captured as mismatch by TGIRT sequencing. Both the modifications and the non-microRNA-small RNAs should be explored as a biomarker for BLCA detection or follow-up.

• Sui, Xuesong; Klungland, Arne & Gao, Lu (2022). RNA m6A modifications in mammalian gametogenesis and pregnancy. Reproduction. ISSN 1470-1626. 165(1), p. R1–R8. doi: 10.1530/REP-22-0112. Full text in Research Archive
• Khodeer, Sherif; Klungland, Arne & Dahl, John Arne (2022). ALKBH5 regulates somatic cell reprogramming in a phase-specific manner. Journal of Cell Science. ISSN 0021-9533. 135(11). doi: 10.1242/jcs.259824. Full text in Research Archive Show summary

  ABSTRACT Establishment of the pluripotency regulatory network in somatic cells by introducing four transcription factors [octamer binding transcription factor 4 (OCT4; also known as POU5F1), sex determining region Y (SRY)-box 2 (SOX2), Kruppel-like factor 4 (KLF4) and cellular myelocytomatosis (c-MYC)] provides a promising tool for cell-based therapies in regenerative medicine. Nevertheless, the mechanisms at play when generating induced pluripotent stem cells from somatic cells are only partly understood. Here, we show that the RNA-specific N6-methyladenosine (m6A) demethylase ALKBH5 regulates somatic cell reprogramming in a stage-specific manner through its catalytic activity. Knockdown or knockout of Alkbh5 in the early reprogramming phase impairs reprogramming efficiency by reducing the proliferation rate through arresting the cells at G2/M phase and decreasing the upregulation of epithelial markers. On the other hand, ALKBH5 overexpression at the early reprogramming phase has no significant impact on reprogramming efficiency, whereas overexpression at the late phase enhances reprogramming by stabilizing Nanog transcripts, resulting in upregulated Nanog expression. Our study provides mechanistic insight into the crucial dynamic role of ALKBH5, mediated through its catalytic activity, in regulating somatic cell reprogramming at the post-transcriptional level. This article has an associated First Person interview with the first author of the paper.

• Su, Zhangli; Monshaugen, Ida; Wilson, Briana; Wang, Fengbin; Klungland, Arne & Ougland, Rune [Show all 7 contributors for this article] (2022). TRMT6/61A-dependent base methylation of tRNA-derived fragments regulates gene-silencing activity and the unfolded protein response in bladder cancer. Nature Communications. 13(1). doi: 10.1038/s41467-022-29790-8. Full text in Research Archive Show summary

  Abstract RNA modifications are important regulatory elements of RNA functions. However, most genome-wide mapping of RNA modifications has focused on messenger RNAs and transfer RNAs, but such datasets have been lacking for small RNAs. Here we mapped N 1 -methyladenosine (m 1 A) in the cellular small RNA space. Benchmarked with synthetic m 1 A RNAs, our workflow identified specific groups of m 1 A-containing small RNAs, which are otherwise disproportionally under-represented. In particular, 22-nucleotides long 3′ tRNA-fragments are highly enriched for TRMT6/61A-dependent m 1 A located within the seed region. TRMT6/61A-dependent m 1 A negatively affects gene silencing by tRF-3s. In urothelial carcinoma of the bladder, where TRMT6/61A is over-expressed, higher m 1 A modification on tRFs is detected, correlated with a dysregulation of tRF targetome. Lastly, TRMT6/61A regulates tRF-3 targets involved in unfolded protein response. Together, our results reveal a mechanism of regulating gene expression via base modification of small RNA.

• Zhao, Xu; Hendriks, Ivo A; Gras, Stephanie Le; Ye, Tao; Alonso, Lucia Ramos & P, Aurélie Nguéa [Show all 13 contributors for this article] (2022). Waves of sumoylation support transcription dynamics during adipocyte differentiation. Nucleic Acids Research (NAR). ISSN 0305-1048. 50(3), p. 1351–1369. doi: 10.1093/nar/gkac027. Full text in Research Archive Show summary

  Abstract Tight control of gene expression networks required for adipose tissue formation and plasticity is essential for adaptation to energy needs and environmental cues. However, the mechanisms that orchestrate the global and dramatic transcriptional changes leading to adipocyte differentiation remain to be fully unraveled. We investigated the regulation of nascent transcription by the sumoylation pathway during adipocyte differentiation using SLAMseq and ChIPseq. We discovered that the sumoylation pathway has a dual function in differentiation; it supports the initial downregulation of pre-adipocyte-specific genes, while it promotes the establishment of the mature adipocyte transcriptional program. By characterizing endogenous sumoylome dynamics in differentiating adipocytes by mass spectrometry, we found that sumoylation of specific transcription factors like PPARγ/RXR and their co-factors are associated with the transcription of adipogenic genes. Finally, using RXR as a model, we found that sumoylation may regulate adipogenic transcription by supporting the chromatin occurrence of transcription factors. Our data demonstrate that the sumoylation pathway supports the rewiring of transcriptional networks required for formation of functional adipocytes. This study also provides the scientists in the field of cellular differentiation and development with an in-depth resource of the dynamics of the SUMO-chromatin landscape, SUMO-regulated transcription and endogenous sumoylation sites during adipocyte differentiation.

• Lång, Emma Helena; Pedersen, Christian; Lång, Anna Ulrika; Blicher, Pernille; Klungland, Arne & Carlson, Andreas [Show all 7 contributors for this article] (2022). Mechanical coupling of supracellular stress amplification and tissue fluidization during exit from quiescence. Proceedings of the National Academy of Sciences of the United States of America (PNAS). ISSN 0027-8424. 119(32). doi: 10.1073/pnas.2201328119. Full text in Research Archive Show summary

  Cellular quiescence is a state of reversible cell cycle arrest that is associated with tissue dormancy. Timely regulated entry into and exit from quiescence is important for processes such as tissue homeostasis, tissue repair, stem cell maintenance, developmental processes, and immunity. However, little is known about processes that control the mechanical adaption to cell behavior changes during the transition from quiescence to proliferation. Here, we show that quiescent human keratinocyte monolayers sustain an actinomyosin-based system that facilitates global cell sheet displacements upon serum-stimulated exit from quiescence. Mechanistically, exposure of quiescent cells to serum-borne mitogens leads to rapid amplification of preexisting contractile sites, leading to a burst in monolayer tension that subsequently drives large-scale displacements of otherwise motility-restricted monolayers. The stress level after quiescence exit correlates with the level of quiescence depth at the time of activation, and a critical stress magnitude must be reached to overcome the cell sheet displacement barrier. The study shows that static quiescent cell monolayers are mechanically poised for motility, and it identifies global stress amplification as a mechanism for overcoming motility restrictions in confined confluent cell monolayers.

• Hildrestrand, Gunn Annette; Rolseth, Veslemøy; Kunath, Nicolas; Suganthan, Rajikala; Jensen, Vidar & Bugaj, Anna Maria [Show all 24 contributors for this article] (2021). NEIL1 and NEIL2 DNA glycosylases modulate anxiety and learning in a cooperative manner in mice. Communications Biology. 4. doi: 10.1038/s42003-021-02864-x. Full text in Research Archive Show summary

  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.

• Alexeeva, Marina; Moen, Marivi Nabong; Xu, Xiang Ming; Rasmussen, Anette; Leiros, Ingar & Kirpekar, Finn [Show all 10 contributors for this article] (2021). Intrinsic Strand-Incision Activity of Human UNG: Implications for Nick Generation in Immunoglobulin Gene Diversification. Frontiers in Immunology. 12, p. 13–13. doi: 10.3389/fimmu.2021.762032. Full text in Research Archive Show summary

  Uracil arises in cellular DNA by cytosine (C) deamination and erroneous replicative incorporation of deoxyuridine monophosphate opposite adenine. The former generates C ! thymine transition mutations if uracil is not removed by uracil-DNA glycosylase (UDG) and replaced by C by the base excision repair (BER) pathway. The primary human UDG is hUNG. During immunoglobulin gene diversification in activated B cells, targeted cytosine deamination by activation-induced cytidine deaminase followed by uracil excision by hUNG is important for class switch recombination (CSR) and somatic hypermutation by providing the substrate for DNA double-strand breaks and mutagenesis, respectively. However, considerable uncertainty remains regarding the mechanisms leading to DNA incision following uracil excision: based on the general BER scheme, apurinic/apyrimidinic (AP) endonuclease (APE1 and/or APE2) is believed to generate the strand break by incising the AP site generated by hUNG. We report here that hUNG may incise the DNA backbone subsequent to uracil excision resulting in a 3´-a,b-unsaturated aldehyde designated uracil-DNA incision product (UIP), and a 5´-phosphate. The formation of UIP accords with an elimination (E2) reaction where deprotonation of C2´ occurs via the formation of a C1´ enolate intermediate. UIP is removed from the 3´-end by hAPE1. This shows that the first two steps in uracil BER can be performed by hUNG, which might explain the significant residual CSR activity in cells deficient in APE1 and APE2.

• Zhang, Li-Sheng; Xiong, Qing-Ping; Pérez, Sonia Peña; Liu, Chang; Wei, Jiangbo & Le, Cassy [Show all 19 contributors for this article] (2021). ALKBH7-mediated demethylation regulates mitochondrial polycistronic RNA processing. Nature Cell Biology. ISSN 1465-7392. 23(7), p. 684–691. doi: 10.1038/s41556-021-00709-7. Full text in Research Archive
• Jin, Kang-Xuan; Zuo, Rujuan; Anastassiadis, Konstantinos; Klungland, Arne; Marr, Carsten & Filipczyk, Adam (2021). N6-methyladenosine (m6A) depletion regulates pluripotency exit by activating signaling pathways in embryonic stem cells. Proceedings of the National Academy of Sciences of the United States of America (PNAS). ISSN 0027-8424. 118(51). doi: 10.1073/pnas.2105192118. Full text in Research Archive Show summary

  N6-methyladenosine (m 6 A) deposition on messenger RNA (mRNA) controls embryonic stem cell (ESC) fate by regulating the mRNA stabilities of pluripotency and lineage transcription factors (TFs) [P. J. Batista et al., Cell Stem Cell 15, 707–719 (2014); Y. Wang et al., Nat. Cell Biol. 16, 191–198 (2014); and S. Geula et al., Science 347, 1002–1006 (2015)]. If the mRNAs of these two TF groups become stabilized, it remains unclear how the pluripotency or lineage commitment decision is implemented. We performed noninvasive quantification of Nanog and Oct4 TF protein levels in reporter ESCs to define cell-state dynamics at single-cell resolution. Long-term single-cell tracking shows that immediate m 6 A depletion by Mettl3 knock-down in serum/leukemia inhibitory factor supports both pluripotency maintenance and its departure. This is mediated by differential and opposing signaling pathways. Increased FGF5 mRNA stability activates pErk, leading to Nanog down-regulation. FGF5-mediated coactivation of pAkt reenforces Nanog expression. In formative stem cells poised toward differentiation, m 6 A depletion activates both pErk and pAkt, increasing the propensity for mesendodermal lineage induction. Stable m 6 A depletion by Mettl3 knock-out also promotes pErk activation. Higher pErk counteracts the pluripotency exit delay exhibited by stably m 6 A-depleted cells upon differentiation. At single-cell resolution, we illustrate that decreasing m 6 A abundances activates pErk and pAkt-signaling, regulating pluripotency departure.

• Tesfahun, Almaz Nigatu; Alexeeva, Marina; Tomkuvienė, Miglė; Arshad, Aysha; Guragain, Prashanna & Klungland, Arne [Show all 9 contributors for this article] (2021). Alleviation of C⋅C Mismatches in DNA by the Escherichia coli Fpg Protein. Frontiers in Microbiology. 12. doi: 10.3389/fmicb.2021.608839. Full text in Research Archive Show summary

  DNA polymerase III mis-insertion may, where not corrected by its 3′→ 5′ exonuclease or the mismatch repair (MMR) function, result in all possible non-cognate base pairs in DNA generating base substitutions. The most thermodynamically unstable base pair, the cytosine (C)⋅C mismatch, destabilizes adjacent base pairs, is resistant to correction by MMR in Escherichia coli, and its repair mechanism remains elusive. We present here in vitro evidence that C⋅C mismatch can be processed by base excision repair initiated by the E. coli formamidopyrimidine-DNA glycosylase (Fpg) protein. The kcat for C⋅C is, however, 2.5 to 10 times lower than for its primary substrate 8-oxoguanine (oxo8G)⋅C, but approaches those for 5,6-dihydrothymine (dHT)⋅C and thymine glycol (Tg)⋅C. The KM values are all in the same range, which indicates efficient recognition of C⋅C mismatches in DNA. Fpg activity was also exhibited for the thymine (T)⋅T mismatch and for N4- and/or 5-methylated C opposite C or T, Fpg activity being enabled on a broad spectrum of DNA lesions and mismatches by the flexibility of the active site loop. We hypothesize that Fpg plays a role in resolving C⋅C in particular, but also other pyrimidine⋅pyrimidine mismatches, which increases survival at the cost of some mutagenesis.

• Yang, Mingyi; Lin, Xiaolin; Segers, Filip; Suganthan, Rajikala; Hildrestrand, Gunn Annette & Rinholm, Johanne Egge [Show all 21 contributors for this article] (2020). OXR1A, a Coactivator of PRMT5 Regulating Histone Arginine Methylation. Cell reports. 30(12), p. 4165–4178. doi: 10.1016/j.celrep.2020.02.063. Full text in Research Archive Show summary

  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.

• Grøsvik, Kristin; Tesfahun, Almaz Nigatu; Muruzábal-Lecumberri, Izaskun; Haugland, Gyri Teien; Leiros, Ingar & Ruoff, Peter [Show all 15 contributors for this article] (2020). The Escherichia coli alkA Gene Is Activated to Alleviate Mutagenesis by an Oxidized Deoxynucleoside. Frontiers in Microbiology. 11. doi: 10.3389/fmicb.2020.00263. Full text in Research Archive Show summary

  The cellular methyl donor S-adenosylmethionine (SAM) and other endo/exogenous agents methylate DNA bases non-enzymatically into products interfering with replication and transcription. An important product is 3-methyladenine (m3A), which in Escherichia coli is removed by m3A-DNA glycosylase I (Tag) and II (AlkA). The tag gene is constitutively expressed, while alkA is induced by sub-lethal concentrations of methylating agents. We previously found that AlkA exhibits activity for the reactive oxygen-induced thymine (T) lesion 5-formyluracil (fU) in vitro. Here, we provide evidence for AlkA involvement in the repair of oxidized bases by showing that the adenine (A) ⋅ T → guanine (G) ⋅ cytosine (C) mutation rate increased 10-fold in E. coli wild-type and alkA– cells exposed to 0.1 mM 5-formyl-2′-deoxyuridine (fdU) compared to a wild-type specific reduction of the mutation rate at 0.2 mM fdU, which correlated with alkA gene induction. G⋅C → A⋅T alleviation occurred without alkA induction (at 0.1 mM fdU), correlating with a much higher AlkA efficiency for fU opposite to G than for that to A. The common keto form of fU is the AlkA substrate. Mispairing with G by ionized fU is favored by its exclusion from the AlkA active site.

• Kong, Xiang Yi; Vik, Erik Sebastian; Nawaz, Meh Sameen; Berges, Natalia; Dahl, Tuva Børresdatter & Vågbø, Cathrine [Show all 17 contributors for this article] (2020). Deletion of Endonuclease V suppresses chemically induced hepatocellular carcinoma. Nucleic Acids Research (NAR). ISSN 0305-1048. 48(8), p. 4463–4479. doi: 10.1093/nar/gkaa115. Full text in Research Archive Show summary

  Endonuclease V (EndoV) is a conserved inosine-specific ribonuclease with unknown biological function. Here, we present the first mouse model lacking EndoV, which is viable without visible abnormalities. We show that endogenous murine EndoV cleaves inosine-containing RNA in vitro, nevertheless a series of experiments fails to link an in vivo function to processing of such transcripts. As inosine levels and adenosine-to-inosine editing often are dysregulated in hepatocellular carcinoma (HCC), we chemically induced HCC in mice. All mice developed liver cancer, however, EndoV−/− tumors were significantly fewer and smaller than wild type tumors. Opposed to human HCC, adenosine deaminase mRNA expression and site-specific editing were unaltered in our model. Loss of EndoV did not affect editing levels in liver tumors, however mRNA expression of a selection of cancer related genes were reduced. Inosines are also found in certain tRNAs and tRNAs are cleaved during stress to produce signaling entities. tRNA fragmentation was dysregulated in EndoV−/− livers and apparently, inosine-independent. We speculate that the inosine-ribonuclease activity of EndoV is disabled in vivo, but RNA binding allowed to promote stabilization of transcripts or recruitment of proteins to fine-tune gene expression. The EndoV−/− tumor suppressive phenotype calls for related studies in human HCC.

• Sankar, Aditya; Lerdrup, Mads; Manaf, Adeel; Johansen, Jens Vilstrup; Gonzalez, Javier Martin & Borup, Rehannah [Show all 13 contributors for this article] (2020). KDM4A regulates the maternal-to-zygotic transition by protecting broad H3K4me3 domains from H3K9me3 invasion in oocytes. Nature Cell Biology. ISSN 1465-7392. 22(4), p. 380–388. doi: 10.1038/s41556-020-0494-z. Full text in Research Archive
• Li, Miaomiao & Klungland, Arne (2020). Modifications and interactions at the R-loop. DNA Repair. ISSN 1568-7864. 96. doi: 10.1016/j.dnarep.2020.102958. Full text in Research Archive
• Pilžys, Tomaš; Marcinkowski, Michał; Kukwa, Wojciech; Garbicz, Damian; Dylewska, Małgorzata & Ferenc, Karolina [Show all 15 contributors for this article] (2019). ALKBH overexpression in head and neck cancer: potential target for novel anticancer therapy. Scientific Reports. 9(1). doi: 10.1038/s41598-019-49550-x. Full text in Research Archive
• Jarøy, Emilie Gimnes; Jiménez, Lourdes Patricia Acosta; Hotta, Ryo; Goldstein, Allan M.; Emblem, Ragnhild & Klungland, Arne [Show all 7 contributors for this article] (2019). "Too much guts and not enough brains": (epi)genetic mechanisms and future therapies of Hirschsprung disease - a review. Clinical Epigenetics. 11. doi: 10.1186/s13148-019-0718-x. Full text in Research Archive Show summary

  Hirschsprung disease is a neurocristopathy, characterized by aganglionosis in the distal bowel. It is caused by failure of the enteric nervous system progenitors to migrate, proliferate, and differentiate in the gut. Development of an enteric nervous system is a tightly regulated process. Both the neural crest cells and the surrounding environment are regulated by different genes, signaling pathways, and morphogens. For this process to be successful, the timing of gene expression is crucial. Hence, alterations in expression of genes specific for the enteric nervous system may contribute to the pathogenesis of Hirschsprung’s disease. Several epigenetic mechanisms contribute to regulate gene expression, such as modifications of DNA and RNA, histone modifications, and microRNAs. Here, we review the current knowledge of epigenetic and epitranscriptomic regulation in the development of the enteric nervous system and its potential significance for the pathogenesis of Hirschsprung’s disease. We also discuss possible future therapies and how targeting epigenetic and epitranscriptomic mechanisms may open new avenues for novel treatment.

• Zhang, Zhichao; Manaf, Adeel; Li, Yanjiao; Pérez, Sonia Peña; Suganthan, Rajikala & Dahl, John Arne [Show all 8 contributors for this article] (2019). Histone methylations define neural stem/progenitor cell subtypes in the mouse subventricular zone. Molecular Neurobiology. ISSN 0893-7648. doi: 10.1007/s12035-019-01777-5. Full text in Research Archive Show summary

  Neural stem/progenitor cells (NSPCs) persist in the mammalian brain throughout life and can be activated in response to the physiological and pathophysiological stimuli. Epigenetic reprogramming of NPSC represents a novel strategy for enhancing the intrinsic potential of the brain to regenerate after brain injury. Therefore, defining the epigenetic features of NSPCs is important for developing epigenetic therapies for targeted reprogramming of NSPCs to rescue neurologic function after injury. In this study, we aimed at defining different subtypes of NSPCs by individual histone methylations. We found the three histone marks, histone H3 lysine 4 trimethylation (H3K4me3), histone H3 lysine 27 trimethylation (H3K27me3), and histone H3 lysine 36 trimethylation (H3K36me3), to nicely and dynamically portray individual cell types during neurodevelopment. First, we found all three marks co-stained with NSPC marker SOX2 in mouse subventricular zone. Then, CD133, Id1, Mash1, and DCX immunostaining were used to define NSPC subtypes. Type E/B, B/C, and C/A cells showed high levels of H3K27me3, H3K36me3, and H3K4me3, respectively. Our results reveal defined histone methylations of NSPC subtypes supporting that epigenetic regulation is critical for neurogenesis and for maintaining NSPCs.

• Abakir, Abdulkadir; Giles, Tom C.; Cristini, Agnese; Foster, Jeremy M.; Dai, Nan & Starczak, Marta [Show all 22 contributors for this article] (2019). N6-methyladenosine regulates the stability of RNA:DNA hybrids in human cells. Nature Genetics. ISSN 1061-4036. 52(1), p. 48–55. doi: 10.1038/s41588-019-0549-x. Full text in Research Archive
• Prikrylova, Terezia; Robertson, Julia; Ferrucci, Francesca; Konorska, Dorota Joanna; Aanes, Håvard & Manaf, Adeel [Show all 16 contributors for this article] (2019). 5-hydroxymethylcytosine marks mammalian origins acting as a barrier to replication. Scientific Reports. 9. doi: 10.1038/s41598-019-47528-3. Full text in Research Archive Show summary

  In most mammalian cells, DNA replication occurs once, and only once between cell divisions. Replication initiation is a highly regulated process with redundant mechanisms that prevent errant initiation events. In lower eukaryotes, replication is initiated from a defined consensus sequence, whereas a consensus sequence delineating mammalian origin of replication has not been identified. Here we show that 5-hydroxymethylcytosine (5hmC) is present at mammalian replication origins. Our data support the hypothesis that 5hmC has a role in cell cycle regulation. We show that 5hmC level is inversely proportional to proliferation; indeed, 5hmC negatively influences cell division by increasing the time a cell resides in G1. Our data suggest that 5hmC recruits replication-licensing factors, then is removed prior to or during origin firing. Later we propose that TET2, the enzyme catalyzing 5mC to 5hmC conversion, acts as barrier to rereplication. In a broader context, our results significantly advance the understating of 5hmC involvement in cell proliferation and disease states.

• Gamage, Thilini Hansamali Gamage Yaddehi; Gunnes, Gjermund; Lee, Robert Hugh; Louch, William Edward; Holmgren, Asbjørn & Bruton, Joseph D. [Show all 19 contributors for this article] (2018). STIM1 R304W causes muscle degeneration and impaired platelet activation in mice. Cell Calcium. ISSN 0143-4160. 76, p. 87–100. doi: 10.1016/j.ceca.2018.10.001. Full text in Research Archive Show summary

  STIM1 and ORAI1 regulate store-operated Ca2+ entry (SOCE) in most cell types, and mutations in these proteins have deleterious and diverse effects. We established a mouse line expressing the STIM1 R304 W gain-of-function mutation causing Stormorken syndrome to explore effects on organ and cell physiology. While STIM1 R304 W was lethal in the homozygous state, surviving mice presented with reduced growth, skeletal muscle degeneration, and reduced exercise endurance. Variable STIM1 expression levels between tissues directly impacted cellular SOCE capacity. In contrast to patients with Stormorken syndrome, STIM1 was downregulated in fibroblasts from Stim1R304W/R304W mice, which maintained SOCE despite constitutive protein activity. In studies using foetal liver chimeras, STIM1 protein was undetectable in homozygous megakaryocytes and platelets, resulting in impaired platelet activation and absent SOCE. These data indicate that downregulation of STIM1 R304 W effectively opposes the gain-of-function phenotype associated with this mutation, and highlight the importance of STIM1 in skeletal muscle development and integrity.

• Alexeeva, Marina; Guragain, Prashanna; Tesfahun, Almaz Nigatu; Tomkuvienė, Miglė; Arshad, Aysha & Gerasimaitė, Rūta [Show all 13 contributors for this article] (2018). Excision of the doubly methylated base N4,5-dimethylcytosine from DNA by Escherichia coli Nei and Fpg proteins. Philosophical Transactions of the Royal Society of London. Biological Sciences. ISSN 0962-8436. 373(1748). doi: 10.1098/rstb.2017.0337. Full text in Research Archive Show summary

  Cytosine (C) in DNA is often modified to 5-methylcytosine (m5C) to execute important cellular functions. Despite the significance of m5C for epigenetic regulation in mammals, damage to m5C has received little attention. For instance, almost no studies exist on erroneous methylation of m5C by alkylating agents to doubly or triply methylated bases. Owing to chemical evidence, and because many prokaryotes express methyltransferases able to convert m5C into N4,5-dimethylcytosine (mN4,5C) in DNA, mN4,5C is probably present in vivo. We screened a series of glycosylases from prokaryotic to human and found significant DNA incision activity of the Escherichia coli Nei and Fpg proteins at mN4,5C residues in vitro. The activity of Nei was highest opposite cognate guanine followed by adenine, thymine (T) and C. Fpg-complemented Nei by exhibiting the highest activity opposite C followed by lower activity opposite T. To our knowledge, this is the first description of a repair enzyme activity at a further methylated m5C in DNA, as well as the first alkylated base allocated as a Nei or Fpg substrate. Based on our observed high sensitivity to nuclease S1 digestion, we suggest that mN4,5C occurs as a disturbing lesion in DNA and that Nei may serve as a major DNA glycosylase in E. coli to initiate its repair.

• Wei, Jiangbo; Liu, Fange; Lu, Zhike; Fei, Qili; Ai, Yuxi & He, P. Cody [Show all 13 contributors for this article] (2018). Differential m6A, m6Am, and m1A Demethylation Mediated by FTO in the Cell Nucleus and Cytoplasm. Molecular Cell. ISSN 1097-2765. 71(6), p. 973–985. doi: 10.1016/j.molcel.2018.08.011. Full text in Research Archive
• Askeland, Georgina; Rodinova, Marie; Štufková, Hana; Dosoudilova, Zaneta; Baxa, Monika & Smatlikova, Petra [Show all 15 contributors for this article] (2018). A transgenic minipig model of Huntington's disease shows early signs of behavioral and molecular pathologies. Disease Models and Mechanisms. ISSN 1754-8403. 11(10). doi: 10.1242/dmm.035949. Full text in Research Archive Show summary

  Huntington's disease (HD) is a monogenic, progressive, neurodegenerative disorder with currently no available treatment. The Libechov transgenic minipig model for HD (TgHD) displays neuroanatomical similarities to humans and exhibits slow disease progression, and is therefore more powerful than available mouse models for the development of therapy. The phenotypic characterization of this model is still ongoing, and it is essential to validate biomarkers to monitor disease progression and intervention. In this study, the behavioral phenotype (cognitive, motor and behavior) of the TgHD model was assessed, along with biomarkers for mitochondrial capacity, oxidative stress, DNA integrity and DNA repair at different ages (24, 36 and 48 months), and compared with age-matched controls. The TgHD minipigs showed progressive accumulation of the mutant huntingtin (mHTT) fragment in brain tissue and exhibited locomotor functional decline at 48 months. Interestingly, this neuropathology progressed without any significant age-dependent changes in any of the other biomarkers assessed. Rather, we observed genotype-specific effects on mitochondrial DNA (mtDNA) damage, mtDNA copy number, 8-oxoguanine DNA glycosylase activity and global level of the epigenetic marker 5-methylcytosine that we believe is indicative of a metabolic alteration that manifests in progressive neuropathology. Peripheral blood mononuclear cells (PBMCs) were relatively spared in the TgHD minipig, probably due to the lack of detectable mHTT. Our data demonstrate that neuropathology in the TgHD model has an age of onset of 48 months, and that oxidative damage and electron transport chain impairment represent later states of the disease that are not optimal for assessing interventions.

• Li, Miaomiao; Zhao, Xu; Wang, Wei; Shi, Hailing; Pan, Qingfei & Lu, Zhike [Show all 11 contributors for this article] (2018). Ythdf2-mediated m6A mRNA clearance modulates neural development in mice. Genome Biology. ISSN 1465-6906. 19(1). doi: 10.1186/s13059-018-1436-y. Full text in Research Archive Show summary

  Background N 6 -methyladenosine (m6A) modification in mRNAs was recently shown to be dynamically regulated, indicating a pivotal role in multiple developmental processes. Most recently, it was shown that the Mettl3-Mettl14 writer complex of this mark is required for the temporal control of cortical neurogenesis. The m6A reader protein Ythdf2 promotes mRNA degradation by recognizing m6A and recruiting the mRNA decay machinery. Results We show that the conditional depletion of the m6A reader protein Ythdf2 in mice causes lethality at late embryonic developmental stages, with embryos characterized by compromised neural development. We demonstrate that neural stem/progenitor cell (NSPC) self-renewal and spatiotemporal generation of neurons and other cell types are severely impacted by the loss of Ythdf2 in embryonic neocortex. Combining in vivo and in vitro assays, we show that the proliferation and differentiation capabilities of NSPCs decrease significantly in Ythdf2 −/− embryos. The Ythdf2 −/− neurons are unable to produce normally functioning neurites, leading to failure in recovery upon reactive oxygen species stimulation. Consistently, expression of genes enriched in neural development pathways is significantly disturbed. Detailed analysis of the m6A-methylomes of Ythdf2 −/− NSPCs identifies that the JAK-STAT cascade inhibitory genes contribute to neuroprotection and neurite outgrowths show increased expression and m6A enrichment. In agreement with the function of Ythdf2, delayed degradation of neuron differentiation-related m6A-containing mRNAs is seen in Ythdf2 −/− NSPCs. Conclusions We show that the m6A reader protein Ythdf2 modulates neural development by promoting m6A-dependent degradation of neural development-related mRNA targets.

• Askeland, Georgina; Dosoudilova, Zaneta; Rodinova, Marie; Klempir, Jiri; Liskova, Irena & Kusnierczyk, Anna [Show all 11 contributors for this article] (2018). Increased nuclear DNA damage precedes mitochondrial dysfunction in peripheral blood mononuclear cells from Huntington's disease patients. Scientific Reports. 8(1). doi: 10.1038/s41598-018-27985-y. Full text in Research Archive Show summary

  Huntington’s disease (HD) is a progressive neurodegenerative disorder primarily affecting the basal ganglia and is caused by expanded CAG repeats in the huntingtin gene. Except for CAG sizing, mitochondrial and nuclear DNA (mtDNA and nDNA) parameters have not yet proven to be representative biomarkers for disease and future therapy. Here, we identified a general suppression of genes associated with aerobic metabolism in peripheral blood mononuclear cells (PBMCs) from HD patients compared to controls. In HD, the complex II subunit SDHB was lowered although not sufficiently to affect complex II activity. Nevertheless, we found decreased level of factors associated with mitochondrial biogenesis and an associated dampening of the mitochondrial DNA damage frequency in HD, implying an early defect in mitochondrial activity. In contrast to mtDNA, nDNA from HD patients was four-fold more modified than controls and demonstrated that nDNA integrity is severely reduced in HD. Interestingly, the level of nDNA damage correlated inversely with the total functional capacity (TFC) score; an established functional score of HD. Our data show that PBMCs are a promising source to monitor HD progression and highlights nDNA damage and diverging mitochondrial and nuclear genome responses representing early cellular impairments in HD.

• Ma, Chunhui; Chang, Mengqi; Lv, Hongyi; Zhang, Zhi-Wei; Zhang, Weilong & He, Xue [Show all 17 contributors for this article] (2018). RNA m6A methylation participates in regulation of postnatal development of the mouse cerebellum. Genome Biology. ISSN 1465-6906. 19(1). doi: 10.1186/s13059-018-1435-z. Full text in Research Archive Show summary

  Background N6-methyladenosine (m6A) is an important epitranscriptomic mark with high abundance in the brain. Recently, it has been found to be involved in the regulation of memory formation and mammalian cortical neurogenesis. However, while it is now established that m6A methylation occurs in a spatially restricted manner, its functions in specific brain regions still await elucidation. Results We identify widespread and dynamic RNA m6A methylation in the developing mouse cerebellum and further uncover distinct features of continuous and temporal-specific m6A methylation across the four postnatal developmental processes. Temporal-specific m6A peaks from P7 to P60 exhibit remarkable changes in their distribution patterns along the mRNA transcripts. We also show spatiotemporal-specific expression of m6A writers METTL3, METTL14, and WTAP and erasers ALKBH5 and FTO in the mouse cerebellum. Ectopic expression of METTL3 mediated by lentivirus infection leads to disorganized structure of both Purkinje and glial cells. In addition, under hypobaric hypoxia exposure, Alkbh5-deletion causes abnormal cell proliferation and differentiation in the cerebellum through disturbing the balance of RNA m6A methylation in different cell fate determination genes. Notably, nuclear export of the hypermethylated RNAs is enhanced in the cerebellum of Alkbh5-deficient mice exposed to hypobaric hypoxia. Conclusions Together, our findings provide strong evidence that RNA m6A methylation is controlled in a precise spatiotemporal manner and participates in the regulation of postnatal development of the mouse cerebellum.

• Alexeeva, Marina; Moen, Marivi Nabong; Grøsvik, Kristin; Tesfahun, Almaz Nigatu; Xu, Xiang Ming & Muruzábal-Lecumberri, Izaskun [Show all 12 contributors for this article] (2018). Excision of uracil from DNA by hSMUG1 includes strand incision and processing. Nucleic Acids Research (NAR). ISSN 0305-1048. 47(2), p. 779–793. doi: 10.1093/nar/gky1184. Full text in Research Archive Show summary

  Uracil arises in DNA by hydrolytic deamination of cytosine (C) and by erroneous incorporation of deoxyuridine monophosphate opposite adenine, where the former event is devastating by generation of C → thymine transitions. The base excision repair (BER) pathway replaces uracil by the correct base. In human cells two uracil-DNA glycosylases (UDGs) initiate BER by excising uracil from DNA; one is hSMUG1 (human single-strand-selective mono-functional UDG). We report that repair initiation by hSMUG1 involves strand incision at the uracil site resulting in a 3′-α,β-unsaturated aldehyde designated uracil-DNA incision product (UIP), and a 5′-phosphate. UIP is removed from the 3′-end by human apurinic/apyrimidinic (AP) endonuclease 1 preparing for single-nucleotide insertion. hSMUG1 also incises DNA or processes UIP to a 3′-phosphate designated uracil-DNA processing product (UPP). UIP and UPP were indirectly identified and quantified by polyacrylamide gel electrophoresis and chemically characterised by matrix-assisted laser desorption/ionisation time-of-flight mass-spectrometric analysis of DNA from enzyme reactions using 18O- or 16O-water. The formation of UIP accords with an elimination (E2) reaction where deprotonation of C2′ occurs via the formation of a C1′ enolate intermediate. A three-phase kinetic model explains rapid uracil excision in phase 1, slow unspecific enzyme adsorption/desorption to DNA in phase 2 and enzyme-dependent AP site incision in phase 3.

• Aas, Aleksander; Isakson, Pauline; Bindesbøll, Christian; Alemu, Endalkachew Ashenafi; Klungland, Arne & Simonsen, Anne (2017). Nucleocytoplasmic shuttling of FTO does not affect starvation-induced autophagy. PLOS ONE. 12(3). doi: 10.1371/journal.pone.0168182. Full text in Research Archive Show summary

  Polymorphic variants of the FTO (fat mass and obesity) gene associate with body mass index in humans, but the underlying molecular mechanisms have not been firmly determined. FTO is linked to energy homeostasis via amino acid sensing and is thought to activate the mammalian target of rapamycin complex 1, a negative regulator of autophagy. FTO localises both to the nucleus and the cytoplasm, and in this study we identify a functional nuclear localisation signal (NLS) in the N-terminus of FTO, as well as nuclear localization information in its very C-terminus. Inhibition of FTO nuclear transport has no effect on autophagy and in contrast to a previously proposed role of FTO in autophagy, we find no difference in starvation-induced autophagy in control cells compared to a panel of cell types depleted of FTO. Future studies that further characterise the cellular functions of FTO will be important to understand why variants in FTO are associated with body weight.

• Pawar, Tina; Bjørås, Magnar; Klungland, Arne & Eide, Lars (2017). Metabolism and DNA repair shape a specific modification pattern in mitochondrial DNA. Mitochondrion (Amsterdam. Print). ISSN 1567-7249. 40, p. 16–28. doi: 10.1016/j.mito.2017.09.002. Full text in Research Archive Show summary

  The mitochondrial DNA (mtDNA) resides in the vicinity of energy-rich reactions. Thus, chemical modifications of mtDNA might mirror mitochondrial processes and could serve as biomarkers of metabolic processes in the mitochondria. This hypothesis was tested by assessing modifications at 17 different sites in the mtDNA as a function of cell type, oxidative stress and mitochondrial activity. Two mouse mutants with a metabolic phenotype were compared to wild-type (WT) mice: the ogg1−/− mouse that lacks the 8-oxoguanine DNA glycosylase (OGG1), and the alkbh7−/− mouse missing the ALKBH7 protein that has been implicated in fatty acid oxidation. It was found that cell type, oxidative stress and mitochondrial complex activity shaped distinct modification patterns in mtDNA, and that OGG1 and ALKBH7 independently modulated these modification patterns. The modifications included ribonucleotides, which also accumulated in mtDNA with age. Interestingly, this age-dependent accumulation most likely involves DNA repair, as mtDNA from ogg1−/− mice did not accumulate modifications with age. On the other hand, alkbh7−/− mtDNA accumulated more modifications with age than WT mtDNA. Our results show that mtDNA is dynamically modified with metabolic activity and imply a novel synergy between metabolism and mtDNA repair proteins.

• Rolseth, Veslemøy; Gomez, Luisa Fernanda Luna; Olsen, Ann-Karin; Suganthan, Rajikala; Scheffler, Katja & Neurauter, Christine Gran [Show all 15 contributors for this article] (2017). No cancer predisposition or increased spontaneous mutation frequencies in NEIL DNA glycosylases-deficient mice. Scientific Reports. 7. doi: 10.1038/s41598-017-04472-4. Full text in Research Archive Show summary

  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.

• Tang, Chong; Klukovich, Rachel; Peng, Hongying; Wang, Zhuqing; Yu, Tian & Zhang, Ying [Show all 9 contributors for this article] (2017). ALKBH5-dependent m6A demethylation controls splicing and stability of long 3'-UTR mRNAs in male germ cells. Proceedings of the National Academy of Sciences of the United States of America (PNAS). ISSN 0027-8424. 115(2), p. E325–E333. doi: 10.1073/pnas.1717794115. Full text in Research Archive
• Chymkowitch, Pierre; P, Aurélie Nguéa; Aanes, Håvard; Robertson, Joseph; Klungland, Arne & Enserink, Jorrit (2017). TORC1-dependent sumoylation of Rpc82 promotes RNA polymerase III assembly and activity. Proceedings of the National Academy of Sciences of the United States of America (PNAS). ISSN 0027-8424. 114(5), p. 1039–1044. doi: 10.1073/pnas.1615093114. Full text in Research Archive
• Seifermann, Marco; Ulges, Alexander; Bopp, Tobias; Melcea, Svetlana; Schäfer, Andrea & Oka, Sugako [Show all 10 contributors for this article] (2017). Role of the DNA repair glycosylase OGG1 in the activation of murine splenocytes. DNA Repair. ISSN 1568-7864. 58, p. 13–20. doi: 10.1016/j.dnarep.2017.08.005. Full text in Research Archive
• Olsen, Maria Belland; Hildrestrand, Gunn Annette; Scheffler, Katja; Vinge, Leif Erik; Alfsnes, Katrine & Palibrk, Vuk [Show all 29 contributors for this article] (2017). NEIL3-Dependent Regulation of Cardiac Fibroblast Proliferation Prevents Myocardial Rupture. Cell reports. 18, p. 82–92. doi: 10.1016/j.celrep.2016.12.009. Full text in Research Archive Show summary

  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.

• Landfors, Miriam; Johansen, Jostein; Aronsen, Jan Magnus; Vågbø, Cathrine Broberg; Doré, Louis C. & He, Chuan [Show all 13 contributors for this article] (2017). Genome-wide profiling of DNA 5-hydroxymethylcytosine during rat Sertoli cell maturation. Cell Discovery. ISSN 2056-5968. 3. doi: 10.1038/celldisc.2017.13. Full text in Research Archive Show summary

  Sertoli cells have dual roles during the cells’ lifetime. In the juvenile mammal, Sertoli cells proliferate and create the structure of the testis, and during puberty they cease to proliferate and take on the adult role of supporting germ cells through spermatogenesis. Accordingly, many genes expressed in Sertoli cells during testis formation are repressed during spermatogenesis. 5-Hydroxymethylcytosine (5hmC) is a DNA modification enzymatically generated from 5mC and present in all investigated mammalian tissues at varying levels. Using mass spectrometry and immunofluorescence staining we identified a substantial Sertoli cell-specific global 5hmC increase during rat puberty. Chemical labeling, pull-down and sequencing of 5hmC-containing genomic DNA from juvenile and adult rat Sertoli cells revealed that genes that lose or gain 5hmC belong to different functional pathways and mirror the functions of the cells in the two different states. Loss of 5hmC is associated with genes involved in development and cell structure, whereas gain of 5hmC is associated with genes involved in cellular pathways pertaining to the function of the adult Sertoli cells. This redistribution during maturation shows that 5hmC is a dynamic nucleotide modification, correlated to gene expression.

• Kalikstad, Betty; Kultima, Hanna Goransson; Andersstuen, Terese; Klungland, Arne & Isaksson, Anders (2017). Gene expression profiles in preterm infants on continuous long-term oxygen therapy suggest reduced oxidative stress-dependent signaling during hypoxia. Molecular Medicine Reports. ISSN 1791-2997. 15(4), p. 1513–1526. doi: 10.3892/mmr.2017.6185. Full text in Research Archive Show summary

  Preterm infants are susceptible to neonatal inflammatory/infective diseases requiring drug therapy. The present study hypothesized that mRNA expression in the blood may be modulated by signaling pathways during treatment. The current study aimed to explore changes in global gene expression in the blood from preterm infants with the objective of identifying patterns or pathways of potential relevance to drug therapy. The infants involved were selected based on maternal criteria indicating increased risk for therapeutic intervention. Global mRNA expression was measured in 107 longitudinal whole blood samples using Affymetrix Human‑Genome‑U133 Plus 2.0‑arrays; samples were obtained from 20 preterm infants. Unsupervised clustering revealed a distinct homogeneous gene expression pattern in 13 samples derived from seven infants undergoing continuous oxygen therapy. At these sampling times, all but one of the seven infants exhibited severe drops in peripheral capillary saturation levels below 60%. The infants were reoxygenated with 100% inspired oxygen concentration. The other samples (n=94) represented the infants from the cohort at time points when they did not undergo continuous oxygen therapy. Comparing these two sets of samples identified a distinct gene expression pattern of 5,986 significantly differentially expressed genes, of which 5,167 genes exhibited reduced expression levels during transient hypoxia. This expression pattern was reversed when the infants became stable, i.e., when they were not continuously oxygenated and had no events of hypoxia. To identify signaling pathways involved in gene regulation, the Database for Annotation, Visualization and Integrated Discovery online tool was used. Mitogen‑activated protein kinases, which are normally induced by oxidative stress, exhibited reduced gene expression during hypoxia. In addition, nuclear factor erythroid 2‑related factor 2‑antioxidant response element target genes involved in oxidative stress protection were also expressed at lower levels, suggesting reduced transcription of this pathway. The findings of the present study suggest that oxidative stress‑dependent signaling is reduced during hypoxia. Understanding the molecular response in preterm infants during continuous oxygenation may aid in refining therapeutic strategies for oxygen therapy.

• Jordan, Jennifer J.; Chhim, Sophea; Margulies, Carrie M.; Allocca, Mariacarmela; Bronson, Roderick T. & Klungland, Arne [Show all 8 contributors for this article] (2017). ALKBH7 drives a tissue and sex-specific necrotic cell death response following alkylation-induced damage. Cell Death & Disease. 8. doi: 10.1038/cddis.2017.343. Full text in Research Archive
• Liu, Fange; Clark, Wesley; Luo, Guanzheng; Wang, Xiaoyun; Fu, Ye & Wei, Jiangbo [Show all 16 contributors for this article] (2016). ALKBH1-Mediated tRNA Demethylation Regulates Translation. Cell. ISSN 0092-8674. 167(3), p. 816–828. doi: 10.1016/j.cell.2016.09.038. Full text in Research Archive
• Mi, Shuangli; Klungland, Arne & Yang, Yun-Gui (2016). Base-excision repair and beyond —A short summary attributed to scientific achievements of Tomas Lindahl, Nobel Prize Laureate in Chemistry 2015. Science China Life Sciences. ISSN 1674-7305. 59(1), p. 89–92. doi: 10.1007/s11427-015-4983-4. Full text in Research Archive
• Landfors, Miriam; Nakken, Sigve; Fusser, Markus; Dahl, John Arne; Klungland, Arne & Fedorcsák, Péter (2016). Sequencing of FTO and ALKBH5 in men undergoing infertility work-up identifies an infertility-associated variant and two missense mutations. Fertility and Sterility. ISSN 0015-0282. 105(5), p. 1170–1179e5. doi: 10.1016/j.fertnstert.2016.01.002. Full text in Research Archive
• Tekpli, Xavier; Urbanucci, Alfonso; Hashim, Adnan; Vågbø, Cathrine Broberg; Lyle, Robert & Kringen, Marianne K. [Show all 11 contributors for this article] (2016). Changes of 5-hydroxymethylcytosine distribution during myeloid and lymphoid differentiation of CD34+ cells. Epigenetics & Chromatin. 9. doi: 10.1186/s13072-016-0070-8. Full text in Research Archive Show summary

  Background Hematopoietic stem cell renewal and differentiation are regulated through epigenetic processes. The conversion of 5-methylcytosine into 5-hydroxymethylcytosine (5hmC) by ten-eleven-translocation enzymes provides new insights into the epigenetic regulation of gene expression during development. Here, we studied the potential gene regulatory role of 5hmC during human hematopoiesis. Results We used reduced representation of 5-hydroxymethylcytosine profiling (RRHP) to characterize 5hmC distribution in CD34+ cells, CD4+ T cells, CD19+ B cells, CD14+ monocytes and granulocytes. In all analyzed blood cell types, the presence of 5hmC at gene bodies correlates positively with gene expression, and highest 5hmC levels are found around transcription start sites of highly expressed genes. In CD34+ cells, 5hmC primes for the expression of genes regulating myeloid and lymphoid lineage commitment. Throughout blood cell differentiation, intragenic 5hmC is maintained at genes that are highly expressed and required for acquisition of the mature blood cell phenotype. Moreover, in CD34+ cells, the presence of 5hmC at enhancers associates with increased binding of RUNX1 and FLI1, transcription factors essential for hematopoiesis. Conclusions Our study provides a comprehensive genome-wide overview of 5hmC distribution in human hematopoietic cells and new insights into the epigenetic regulation of gene expression during human hematopoiesis.

• Dahl, John Arne; Jung, Inkyung; Aanes, Håvard; Greggains, Gareth; Manaf, Adeel & Lerdrup, Mads [Show all 20 contributors for this article] (2016). Broad histone H3K4me3 domains in mouse oocytes modulate maternal-to-zygotic transition. Nature. ISSN 0028-0836. 537(7621), p. 548–552. doi: 10.1038/nature19360. Full text in Research Archive
• Klungland, Arne & Robertson, Adam Brian (2016). Oxidized C5-methyl cytosine bases in DNA: 5-Hydroxymethylcytosine; 5-formylcytosine; and 5-carboxycytosine. Free Radical Biology & Medicine. ISSN 0891-5849. 107, p. 62–68. doi: 10.1016/j.freeradbiomed.2016.11.038. Full text in Research Archive
• Klungland, Arne; Dahl, John Arne; Greggains, Gareth; Fedorcsák, Péter & Filipczyk, Adam (2016). Reversible RNA modifications in meiosis and pluripotency. Nature Methods. ISSN 1548-7091. 14(1), p. 18–22. doi: 10.1038/nmeth.4111. Full text in Research Archive
• Rasmussen, Erik; Vågbø, Cathrine Broberg; Münch, Daniel; Krokan, Hans Einar; Klungland, Arne & Amdam, Gro Vang [Show all 7 contributors for this article] (2016). DNA base modifications in honey bee and fruit fly genomes suggest an active demethylation machinery with species- and tissue-specific turnover rates. Biochemistry and Biophysics Reports. 6, p. 9–15. doi: 10.1016/j.bbrep.2016.02.011. Full text in Research Archive Show summary

  Well-known epigenetic DNA modifications in mammals include the addition of a methyl group and a hydroxyl group to cytosine, resulting in 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) respectively. In contrast, the abundance and the functional implications of these modifications in invertebrate model organisms such as the honey bee (Apis mellifera) and the fruit fly (Drosophila melanogaster) are not well understood. Here we show that both adult honey bees and fruit flies contain 5mC and also 5hmC. Using a highly sensitive liquid chromatography/tandem mass spectrometry (LC/MS/MS) technique, we quantified 5mC and 5hmC in different tissues of adult honey bee worker castes and in adult fruit flies. A comparison of our data with reports from human and mouse shed light on notable differences in 5mC and 5hmC levels between tissues and species.

• Weber, Alain R.; Krawczyk, Claudia; Robertson, Adam Brian; Kusnierczyk, Anna; Vågbø, Cathrine Broberg & Schürmann, David [Show all 8 contributors for this article] (2016). Biochemical reconstitution of TET1-TDG-BER-dependent active DNA demethylation reveals a highly coordinated mechanism. Nature Communications. 7. doi: 10.1038/ncomms10806. Full text in Research Archive Show summary

  Cytosine methylation in CpG dinucleotides is an epigenetic DNA modification dynamically established and maintained by DNA methyltransferases and demethylases. Molecular mechanisms of active DNA demethylation began to surface only recently with the discovery of the 5-methylcytosine (5mC)-directed hydroxylase and base excision activities of ten–eleven translocation (TET) proteins and thymine DNA glycosylase (TDG). This implicated a pathway operating through oxidation of 5mC by TET proteins, which generates substrates for TDG-dependent base excision repair (BER) that then replaces 5mC with C. Yet, direct evidence for a productive coupling of TET with BER has never been presented. Here we show that TET1 and TDG physically interact to oxidize and excise 5mC, and proof by biochemical reconstitution that the TET–TDG–BER system is capable of productive DNA demethylation. We show that the mechanism assures a sequential demethylation of symmetrically methylated CpGs, thereby avoiding DNA double-strand break formation but contributing to the mutability of methylated CpGs.

• Vindedal, Gry Fluge; Thoren, Anna; Jensen, Vidar; Klungland, Arne; Zhang, Yong & Holtzman, Michael [Show all 8 contributors for this article] (2016). Removal of aquaporin-4 from glial and ependymal membranes causes brain water accumulation. Molecular and Cellular Neuroscience. ISSN 1044-7431. 77, p. 47–52. doi: 10.1016/j.mcn.2016.10.004. Full text in Research Archive
• Alemu, Endalkachew Ashenafi; He, Chuan & Klungland, Arne (2016). ALKBHs-facilitated RNA modifications and de-modifications. DNA Repair. ISSN 1568-7864. 44, p. 87–91. doi: 10.1016/j.dnarep.2016.05.026. Full text in Research Archive
• Møllersen, Linda; Moldestad, Olve; Rowe, Alexander D.; Bjølgerud, Anja; Holm, Ingunn & Tveterås, Linda Augustin [Show all 8 contributors for this article] (2016). Effects of anthocyanins on CAG repeat instability and behaviour in Huntington's disease R6/1 mice. PLOS Currents. 8(HuntingtonDisease, 2016). doi: 10.1371/currents.hd.58d04209ab6d5de0844db7ef5628ff67. Full text in Research Archive
• Ougland, Rune; Jonson, Ida; Moen, Marivi Nabong; Nesse, Gaute; Asker, Gry & Klungland, Arne [Show all 7 contributors for this article] (2016). Role of ALKBH1 in the Core Transcriptional Network of Embryonic Stem Cells. Cellular Physiology and Biochemistry. ISSN 1015-8987. 38(1), p. 173–184. doi: 10.1159/000438619. Full text in Research Archive
• Lauritzen, Knut Husø; Hasan-Olive, Md Mahdi; Regnell, Christine E; Kleppa, Liv; Knudsen, Morten Scheibye & Gjedde, Albert [Show all 10 contributors for this article] (2016). A ketogenic diet accelerates neurodegeneration in mice with induced mitochondrial DNA toxicity in the forebrain. Neurobiology of Aging. ISSN 0197-4580. 48, p. 34–47. doi: 10.1016/j.neurobiolaging.2016.08.005. Full text in Research Archive
• Fusser, Markus; Kernstock, Stefan; Aileni, Vinay Kumar; Jacobsen, Wolfgang Mathias; Falnes, Pål & Klungland, Arne (2015). Lysine methylation of the valosin-containing protein (VCP) is dispensable for development and survival of mice. PLOS ONE. 10(11). doi: 10.1371/journal.pone.0141472. Full text in Research Archive
• Chymkowitch, Pierre; P, Aurélie Nguéa; Aanes, Håvard; Koehler, Christian; Thiede, Bernd & Lorenz, Susanne [Show all 9 contributors for this article] (2015). Sumoylation of Rap1 mediates the recruitment of TFIID to promote transcription of ribosomal protein genes. Genome Research. ISSN 1088-9051. 25(6), p. 897–906. doi: 10.1101/gr.185793.114. Full text in Research Archive
• Bjørge, Monica Dahl; Hildrestrand, Gunn Annette; Scheffler, Katja; Suganthan, Rajikala; Rolseth, Veslemøy & Kusnierczyk, Anna [Show all 15 contributors for this article] (2015). Synergistic Actions of Ogg1 and Mutyh DNA Glycosylases Modulate Anxiety-like Behavior in Mice. Cell reports. 13(12), p. 2671–2678. doi: 10.1016/j.celrep.2015.12.001. Full text in Research Archive Show summary

  Ogg1 and Mutyh DNA glycosylases cooperate to prevent mutations caused by 8-oxoG, a major premutagenic DNA lesion associated with cognitive decline. We have examined behavior and cognitive function in mice deficient of these glycosylases. Ogg1−/−Mutyh−/− mice were more active and less anxious, with impaired learning ability. In contrast, Mutyh−/− mice showed moderately improved memory. We observed no apparent change in genomic 8-oxoG levels, suggesting that Ogg1 and Mutyh play minor roles in global repair in adult brain. Notably, transcriptome analysis of hippocampus revealed that differentially expressed genes in the mutants belong to pathways known to be involved in anxiety and cognition. Esr1 targets were upregulated, suggesting a role of Ogg1 and Mutyh in repression of Esr1 signaling. Thus, beyond their involvement in DNA repair, Ogg1 and Mutyh regulate hippocampal gene expression related to cognition and behavior, suggesting a role for the glycosylases in regulating adaptive behavior.

• Lauritzen, Knut Husø; Kleppa, Liv; Aronsen, Jan Magnus; Eide, Lars; Carlsen, Harald & Haugen, Øyvind Pernell [Show all 12 contributors for this article] (2015). Impaired dynamics and function of mitochondria caused by mtDNA toxicity leads to heart failure. American Journal of Physiology. Heart and Circulatory Physiology. ISSN 0363-6135. 309(3), p. H434–H449. doi: 10.1152/ajpheart.00253.2014. Full text in Research Archive
• Ke, Shengdong; Alemu, Endalkachew Ashenafi; Mertens, Claudia; Gantman, Emily Conn; Fak, John J. & Mele, Aldo [Show all 15 contributors for this article] (2015). A majority of m6A residues are in the last exons, allowing the potential for 3' UTR regulation. Genes & Development. ISSN 0890-9369. 29(19), p. 2037–2053. doi: 10.1101/gad.269415.115. Full text in Research Archive
• Ougland, Rune; Rognes, Torbjørn; Klungland, Arne & Larsen, Elisabeth (2015). Non-homologous functions of the AlkB homologs. Journal of Molecular Cell Biology. ISSN 1674-2788. 7(6), p. 494–504. doi: 10.1093/jmcb/mjv029. Full text in Research Archive
• Klungland, Arne & Dahl, John Arne (2014). Dynamic RNA modifications in disease. Current Opinion in Genetics and Development. ISSN 0959-437X. 26, p. 47–52. doi: 10.1016/j.gde.2014.05.006. Full text in Research Archive
• Robertson, Adam Brian; Robertson, Julia; Fusser, Markus & Klungland, Arne (2014). Endonuclease G preferentially cleaves 5-hydroxymethylcytosine-modified DNA creating a substrate for recombination. Nucleic Acids Research (NAR). ISSN 0305-1048. 42(21), p. 13280–13293. doi: 10.1093/nar/gku1032. Full text in Research Archive Show summary

  5-hydroxymethylcytosine (5hmC) has been suggested to be involved in various nucleic acid transactions and cellular processes, including transcriptional regulation, demethylation of 5-methylcytosine and stem cell pluripotency. We have identified an activity that preferentially catalyzes the cleavage of double-stranded 5hmC-modified DNA. Using biochemical methods we purified this activity from mouse liver extracts and demonstrate that the enzyme responsible for the cleavage of 5hmC-modified DNA is Endonuclease G (EndoG). We show that recombinant EndoG preferentially recognizes and cleaves a core sequence when one specific cytosine within that core sequence is hydroxymethylated. Additionally, we provide in vivo evidence that EndoG catalyzes the formation of double-stranded DNA breaks and that this cleavage is dependent upon the core sequence, EndoG and 5hmC. Finally, we demonstrate that the 5hmC modification can promote conservative recombination in an EndoG-dependent manner.

• Robertson, Adam Brian; Dahl, John Arne & Klungland, Arne (2014). DNA metabolism: bases of DNA repair and regulation. Nature Chemical Biology. ISSN 1552-4450. 10(7), p. 487–488. doi: 10.1038/nchembio.1553. Full text in Research Archive
• Nilsen, Anja; Fusser, Markus; Greggains, Gareth; Fedorcsak, Peter & Klungland, Arne (2014). ALKBH4 depletion in mice leads to spermatogenic defects. PLOS ONE. 9(8). doi: 10.1371/journal.pone.0105113. Full text in Research Archive
• Zdzalik, Daria; Vågbø, Cathrine Broberg; Kirpekar, F; Davydova, Erna; Puścian, Alicja & Maciejewska, Agnieszka M. [Show all 11 contributors for this article] (2014). Protozoan ALKBH8 Oxygenases Display both DNA Repair and tRNA Modification Activities. PLOS ONE. 9(6). doi: 10.1371/journal.pone.0098729. Full text in Research Archive Show summary

  The ALKBH family of Fe(II) and 2-oxoglutarate dependent oxygenases comprises enzymes that display sequence homology to AlkB from E. coli, a DNA repair enzyme that uses an oxidative mechanism to dealkylate methyl and etheno adducts on the nucleobases. Humans have nine different ALKBH proteins, ALKBH1–8 and FTO. Mammalian and plant ALKBH8 are tRNA hydroxylases targeting 5-methoxycarbonylmethyl-modified uridine (mcm5U) at the wobble position of tRNAGly(UCC). In contrast, the genomes of some bacteria encode a protein with strong sequence homology to ALKBH8, and robust DNA repair activity was previously demonstrated for one such protein. To further explore this apparent functional duality of the ALKBH8 proteins, we have here enzymatically characterized a panel of such proteins, originating from bacteria, protozoa and mimivirus. All the enzymes showed DNA repair activity in vitro, but, interestingly, two protozoan ALKBH8s also catalyzed wobble uridine modification of tRNA, thus displaying a dual in vitro activity. Also, we found the modification status of tRNAGly(UCC) to be unaltered in an ALKBH8 deficient mutant of Agrobacterium tumefaciens, indicating that bacterial ALKBH8s have a function different from that of their eukaryotic counterparts. The present study provides new insights on the function and evolution of the ALKBH8 family of proteins.

• Dahl, John Arne & Klungland, Arne (2014). Micro Chromatin Immunoprecipitation (μChIP) from Early Mammalian Embryos. Methods in molecular biology. ISSN 1064-3745. 1222, p. 227–245. doi: 10.1007/978-1-4939-1594-1_17. Full text in Research Archive
• Li, Ming-Ming; Nilsen, Anja; Shi, Yue; Fusser, Markus; Ding, Yue-He & Fu, Ye [Show all 19 contributors for this article] (2013). ALKBH4-dependent demethylation of actin regulates actomyosin dynamics. Nature Communications. 4. doi: 10.1038/ncomms2863. Full text in Research Archive
• Furu, Kari & Klungland, Arne (2013). Tzfp Represses the Androgen Receptor in Mouse Testis. PLOS ONE. 8(4). doi: 10.1371/journal.pone.0062314. Full text in Research Archive
• Zheng, Guanqun; Dahl, John Arne; Niu, Yamei; Fedorcsak, Peter; Huang, Chun-Min & Li, Charles J. [Show all 28 contributors for this article] (2013). ALKBH5 Is a Mammalian RNA Demethylase that Impacts RNA Metabolism and Mouse Fertility. Molecular Cell. ISSN 1097-2765. 49(1), p. 18–29. doi: 10.1016/j.molcel.2012.10.015. Full text in Research Archive
• Solberg, Anja; Robertson, Adam Brian; Aronsen, Jan Magnus; Rognmo, Øivind; Sjaastad, Ivar & Wisløff, Ulrik [Show all 7 contributors for this article] (2013). Deletion of mouse Alkbh7 leads to obesity. Journal of Molecular Cell Biology. ISSN 1674-2788. 5(3), p. 194–203. doi: 10.1093/jmcb/mjt012. Full text in Research Archive
• Jonson, Ida; Ougland, Rune; Klungland, Arne & Larsen, Elisabeth (2013). Oxidative stress causes DNA triplet expansion in Huntington's disease mouse embryonic stem cells. Stem Cell Research. ISSN 1873-5061. 11(3), p. 1264–1271. doi: 10.1016/j.scr.2013.08.010. Full text in Research Archive
• Nordstrand, Line Mari; Furu, Kari; Paulsen, Jonas; Rognes, Torbjørn & Klungland, Arne (2012). Alkbh1 and Tzfp repress a non-repeat piRNA cluster in pachytene spermatocytes. Nucleic Acids Research (NAR). ISSN 0305-1048. 40(21), p. 10950–10963. doi: 10.1093/nar/gks839. Full text in Research Archive Show summary

  Piwi proteins and Piwi-interacting small RNAs (piRNAs) have known functions in transposon silencing in the male germline of fetal and newborn mice. Both are also present in adult testes; however, their function here remains a mystery. Here, we confirm that most piRNAs in meiotic spermatocytes originate from clusters in non-repeat intergenic regions of DNA. The regulation of these piRNA clusters, including the processing of the precursor transcripts into individual piRNAs, is accomplished through mostly unknown processes. We present a possible regulatory mechanism for one such cluster, named cluster 1082B, located on chromosome 7 in the mouse genome. The 1082B precursor transcript and its 788 unique piRNAs are repressed by the Alkbh1 dioxygenase and the testis-specific transcription repressor Tzfp. We observe a remarkable >1000-fold upregulation of individual piRNAs in pachytene spermatocytes isolated from Alkbh1- and Tzfp -deficient murine testes. Repression of cluster 1082B is further supported by the identification of a 10-bp Tzfp recognition sequence contained within the precursor transcript. Downregulation of LINE1 and IAP transcripts in the Alkbh1- and Tzfp -deficient mice leads us to propose a potential role for the 1082B-encoded piRNAs in transposon control.

• Meza, Trine Johansen; Moen, Marivi; Vågbø, Cathrine Broberg; Krokan, Hans Einar; Klungland, Arne & Grini, Paul Eivind [Show all 7 contributors for this article] (2012). The DNA dioxygenase ALKBH2 protects Arabidopsis thaliana against methylation damage. Nucleic Acids Research (NAR). ISSN 0305-1048. 40(14), p. 6620–6631. doi: 10.1093/nar/gks327. Full text in Research Archive Show summary

  The Escherichia coli AlkB protein (EcAlkB) is a DNA repair enzyme which reverses methylation damage such as 1-methyladenine (1-meA) and 3-methylcytosine (3-meC). The mammalian AlkB homologues ALKBH2 and ALKBH3 display EcAlkB-like repair activity in vitro , but their substrate specificities are different, and ALKBH2 is the main DNA repair enzyme for 1-meA in vivo . The genome of the model plant Arabidopsis thaliana encodes several AlkB homologues, including the yet uncharacterized protein AT2G22260, which displays sequence similarity to both ALKBH2 and ALKBH3. We have here characterized protein AT2G22260, by us denoted ALKBH2, as both our functional studies and bioinformatics analysis suggest it to be an orthologue of mammalian ALKBH2. The Arabidopsis ALKBH2 protein displayed in vitro repair activities towards methyl and etheno adducts in DNA, and was able to complement corresponding repair deficiencies of the E. coli alkB mutant. Interestingly, alkbh2 knock-out plants were sensitive to the methylating agent methylmethanesulphonate (MMS), and seedlings from these plants developed abnormally when grown in the presence of MMS. The present study establishes ALKBH2 as an important enzyme for protecting Arabidopsis against methylation damage in DNA, and suggests its homologues in other plants to have a similar function.

• Kleppa, Liv; Mari, Pierre-Olivier; Larsen, Elisabeth; Lien, Guro Flor; Godon, Camille & Theil, Arjan F [Show all 11 contributors for this article] (2012). Kinetics of endogenous mouse FEN1 in base excision repair. Nucleic Acids Research (NAR). ISSN 0305-1048. 40(18), p. 9044–9059. doi: 10.1093/nar/gks673. Full text in Research Archive
• Møllersen, Linda; Rowe, Alexander D.; Illuzzi, Jennifer L.; Hildrestrand, Gunn Annette; Gerhold, Katharina J. & Tveterås, Linda Augustin [Show all 10 contributors for this article] (2012). Neil1 is a genetic modifier of somatic and germline CAG trinucleotide repeat instability in R6/1 mice. Human Molecular Genetics. ISSN 0964-6906. 21(22), p. 4939–4947. doi: 10.1093/hmg/dds337. Full text in Research Archive
• Ougland, Rune; Lando, David; Jonson, Ida; Dahl, John Arne; Moen, Marivi & Nordstrand, Line Mari [Show all 11 contributors for this article] (2012). ALKBH1 Is a Histone H2A Dioxygenase Involved in Neural Differentiation. Stem Cells. ISSN 1066-5099. 30(12), p. 2672–2682. doi: 10.1002/stem.1228. Full text in Research Archive Show summary

  AlkB homolog 1 (ALKBH1) is one of nine members of the family of mammalian AlkB homologs. Most Alkbh1−/− mice die during embryonic development, and survivors are characterized by defects in tissues originating from the ectodermal lineage. In this study, we show that deletion of Alkbh1 prolonged the expression of pluripotency markers in embryonic stem cells and delayed the induction of genes involved in early differentiation. In vitro differentiation to neural progenitor cells (NPCs) displayed an increased rate of apoptosis in the Alkbh1−/− NPCs when compared with wild-type cells. Whole-genome expression analysis and chromatin immunoprecipitation revealed that ALKBH1 regulates both directly and indirectly, a subset of genes required for neural development. Furthermore, our in vitro enzyme activity assays demonstrate that ALKBH1 is a histone dioxygenase that acts specifically on histone H2A. Mass spectrometric analysis demonstrated that histone H2A from Alkbh1−/− mice are improperly methylated. Our results suggest that ALKBH1 is involved in neural development by modifying the methylation status of histone H2A. STEM CELLS 2012;30:2672–2682 First published online in STEM CELLSEXPRESS November 7, 2012. available online without subscription through the open access option.

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• Holub, Elina; Jochems, Reina; Zeremichael, Teklu Tewoldebrhan; Klungland, Arne & Kommisrud, Elisabeth (2026). Reduced in vitro maturation time of bovine oocytes may increase blastocyst yield. Full text in Research Archive
• Kleve, Guro; Santa, Chiara Dalla; Kristiansen, Felicia Iselin Svensson; Tegnander, Marita; Buch, Victoria & Hagemann, Cecilie Steen [Show all 29 contributors for this article] (2025). Digital home-based multimodal prehabilitation of colorectal cancer patients prior to surgery (the dHOPE study): a non-inferiority clinical trial protocol. doi: 10.3389/fdgth.2025.1609678. Full text in Research Archive Show summary

  Cancer surgery is associated with risk of complications and loss of function. Vulnerability factors, such as advanced age, malnutrition, smoking, comorbidity, frailty, and low socioeconomic status increase the risk. Lifestyle intervention prior to surgery, known as prehabilitation, often include physical activity, nutritional support, psychological coaching, and smoking cessation, increase functional reserves and reduce postoperative complications. Most importantly, it prevents loss of functional capacity and dependence. Methods The dHOPE study is a three-armed, open-labelled, parallel-group randomized controlled trial (RCT) with non-inferiority design to compare a digital home-based prehabilitation program with a hospital-based program or no organized prehabilitation. In addition, the dHOPE study aims to identify measurable parameters reflecting the effect of prehabilitation, thus preparing for future personalization of the prehabilitation programs. Discussion The feasibility of multimodal prehabilitation is threatened by low compliance to hospital-based programs due to burdensome commuting, even in central and metropolitan areas. In sparsely populated countries, this challenge is even more pronounced. To ensure equal healthcare to all citizens regardless of address or economic situation, there is a need to transfer the prehabilitation program to the patients’ homes. Thus, the primary hypothesis of dHOPE is that a digital home-based program is not inferior to a hospital-based program. Moreover, given the patient diversity, prehabilitation must be personalized to meet individual profiles or needs. An exploratory subtask of dHOPE is to confirm the utility of clinical, genetic, and molecular factors in evaluating prehabilitation response ultimately to identify new biomarkers and develop medical software for individual risk stratification and development of personalized prehabilitation programs. Clinical Trial registration https://clinicaltrials.gov/ , identifier (NCT06231576).

• Li, Miaomiao; Klungland, Arne & Dalhus, Bjørn (2022). Studies on Protein-RNA:DNA Hybrid Interactions by Microscale Thermophoresis (MST). Methods in molecular biology. ISSN 1064-3745. 2528, p. 239–251. doi: 10.1007/978-1-0716-2477-7_15. Full text in Research Archive
• Tesfahun, Almaz Nigatu; Alexeeva, Marina; Tomkuviene, Migle; Laerdahl, Jon Kristen; Klungland, Arne & Klimašauskas, Saulius [Show all 7 contributors for this article] (2018). Formamidopyrimidine-DNA glycosylase is involved in C:C mismatch repair. Full text in Research Archive
• Tesfahun, Almaz Nigatu; Alexeeva, Marina; Guragain, Prashanna; Tomkuviene, Migle; Arshad, Aysha & Klungland, Arne [Show all 8 contributors for this article] (2017). Repair of double methylated bases in DNA. Full text in Research Archive
• Tesfahun, Almaz Nigatu; Guragain, Prashanna; Alexeeva, Marina; Arshad, Aysha; Tomkuviene, Migle & Laerdahl, Jon K. [Show all 9 contributors for this article] (2017). Repair of dimethylated bases in DNA. Full text in Research Archive
• Tesfahun, Almaz Nigatu; Klungland, Arne; Klimasauskas, Saulius & Bjelland, Svein (2017). Repair of double methylated bases in DNA. Full text in Research Archive
• Alexeeva, Marina; Moen, Marivi Nabong; Xu, Xiang Ming; Rasmussen, Anette; Kirpekar, Finn & Klungland, Arne [Show all 7 contributors for this article] (2017). hSMUG1 is a bi-functional DNA glycosylase. Full text in Research Archive
• Tesfahun, Almaz Nigatu; Klungland, Arne; Klimasauskas, Saulius & Bjelland, Svein (2016). Repair of double methylated bases in DNA. Full text in Research Archive
• Alexeeva, Marina; Moen, Marivi Nabong; Xu, Xiang Ming; Rasmussen, Anette; Kirpekar, Finn & Klungland, Arne [Show all 7 contributors for this article] (2016). hSMUG1-catalysed uracil-DNA strand incision. Full text in Research Archive
• Tesfahun, Almaz Nigatu; Klungland, Arne; Klimašauskas, Saulius & Bjelland, Svein (2016). Repair of double methylated bases in DNA. Full text in Research Archive
• Alexeeva, Marina; Moen, Marivi Nabong; Xu, Xiang Ming; Rasmussen, Anette; Kirpekar, Finn & Klungland, Arne [Show all 7 contributors for this article] (2016). hSMUG1-catalysed uracil-DNA strand incision. Full text in Research Archive
• Catterall, Tara Cherie; Johnson, Inna; Khan, Shaista; Jermstad, Ingunn; Nesse, Gaute & Klungland, Arne [Show all 7 contributors for this article] (2015). The Norwegian Transgenic Centre. Full text in Research Archive
• Khan, Shaista; Jermstad, Ingunn; Catterall, Tara Cherie; Nesse, Gaute; Klungland, Arne & Dalen, Knut Tomas (2014). The Norwegian Transgenic Centre. Full text in Research Archive
• Rasmussen, Erik; Vågbø, Cathrine Broberg; Dahl, John Arne; Münch, Daniel; Krokan, Hans Einar & Klungland, Arne [Show all 7 contributors for this article] (2014). Epigenetically modified cytosines in the honey bee genome. Full text in Research Archive
• Zheng, Guanqun; Dahl, John Arne; Niu, Yamei; Fu, Ye & Klungland, Arne (2013). Sprouts of RNA epigenetics The discovery of mammalian RNA demethylases. RNA Biology. ISSN 1547-6286. 10(6), p. 915–918. doi: 10.4161/rna.24711. Full text in Research Archive
• Rasmussen, Erik; Vågbø, Cathrine Broberg; Dahl, John Arne; Münch, Daniel; Krokan, Hans Einar & Klungland, Arne [Show all 7 contributors for this article] (2013). Presence of modified cytosines in the honey bee genome. Full text in Research Archive

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