bims-numges Biomed News
on Nucleotide metabolism and genome stability
Issue of 2021‒03‒21
twenty-two papers selected by
Sean Rudd
Karolinska Institutet
  1. Peroxisomal-derived ether phospholipids link nucleotides to respirasome assembly.
  2. Rewiring of purine metabolism in response to acidosis stress in glioma stem cells.
  3. Melanoma subpopulations that rapidly escape MAPK pathway inhibition incur DNA damage and rely on stress signalling.
  4. Chk1 inhibition induces a DNA damage bystander effect in cocultured tumour cells.
  5. ATM controls the extent of DNA end resection by eliciting sequential posttranslational modifications of CtIP.
  6. DSB repair pathway choice is regulated by recruitment of 53BP1 through cell cycle-dependent regulation of Sp1.
  7. Mitochondrial genome stability in human: understanding the role of DNA repair pathways.
  8. Excision of mutagenic replication-blocking lesions suppresses cancer but promotes cytotoxicity and lethality in nitrosamine-exposed mice.
  9. Beyond DNA repair and chromosome instability-Fanconi anaemia as a cellular senescence-associated syndrome.
  10. Bloom syndrome and the underlying causes of genetic instability.
  11. Histones on fire: the effect of Dun1 and Mrc1 on origin firing and replication of hyper-acetylated genomes.
  12. NEK1 deficiency affects mitochondrial functions and the transcriptome of key DNA repair pathways.
  13. Human RecQ Helicases in DNA Double-Strand Break Repair.
  14. Stabilisation of half MCM ring by Cdt1 during DNA insertion.
  15. Regulating Polθ in Breast Cancer.
  16. Pol α-primase dependent nuclear localization of the mammalian CST complex.
  17. Karonudib has potent anti-tumor effects in preclinical models of B-cell lymphoma.
  18. Targeting Cancer Metabolism and Current Anti-Cancer Drugs.
  19. Reversible chromatin condensation by the DNA repair and demethylation factor thymine DNA glycosylase.
  20. Terminal Peptide Extensions Augment the Retinal IMPDH1 Catalytic Activity and Attenuate the ATP-induced Fibrillation Events.
  21. Structural basis for the interaction modes of dihydroorotase with the anticancer drugs 5-fluorouracil and 5-aminouracil.
  22. Metabolic implications of non-Electrogenic ATP/ADP exchange in Cancer cells: A mechanistic basis for the Warburg effect.
  1. Nat Chem Biol. 2021 Mar 15.
    Peroxisomal-derived ether phospholipids link nucleotides to respirasome assembly.
    Christopher F Bennett, Katherine E O'Malley, Elizabeth A Perry, Eduardo Balsa, Pedro Latorre-Muro, Christopher L Riley, Chi Luo, Mark Jedrychowski, Steven P Gygi, Pere Puigserver.
      The protein complexes of the mitochondrial electron transport chain exist in isolation and in higher order assemblies termed supercomplexes (SCs) or respirasomes (SC I+III2+IV). The association of complexes I, III and IV into the respirasome is regulated by unknown mechanisms. Here, we designed a nanoluciferase complementation reporter for complex III and IV proximity to determine in vivo respirasome levels. In a chemical screen, we found that inhibitors of the de novo pyrimidine synthesis enzyme dihydroorotate dehydrogenase (DHODH) potently increased respirasome assembly and activity. By-passing DHODH inhibition via uridine supplementation decreases SC assembly by altering mitochondrial phospholipid composition, specifically elevated peroxisomal-derived ether phospholipids. Cell growth rates upon DHODH inhibition depend on ether lipid synthesis and SC assembly. These data reveal that nucleotide pools signal to peroxisomes to modulate synthesis and transport of ether phospholipids to mitochondria for SC assembly, which are necessary for optimal cell growth in conditions of nucleotide limitation.
    DOI:  https://doi.org/10.1038/s41589-021-00772-z
  2. Cell Death Dis. 2021 Mar 15. 12(3): 277
    Rewiring of purine metabolism in response to acidosis stress in glioma stem cells.
    Xiaoyu Xu, Liping Wang, Qingce Zang, Shanshan Li, Limei Li, Zhixing Wang, Jiuming He, Boqin Qiang, Wei Han, Ruiping Zhang, Xiaozhong Peng, Zeper Abliz.
      Glioma stem cells (GSCs) contribute to therapy resistance and poor outcomes for glioma patients. A significant feature of GSCs is their ability to grow in an acidic microenvironment. However, the mechanism underlying the rewiring of their metabolism in low pH remains elusive. Here, using metabolomics and metabolic flux approaches, we cultured GSCs at pH 6.8 and pH 7.4 and found that cells cultured in low pH exhibited increased de novo purine nucleotide biosynthesis activity. The overexpression of glucose-6-phosphate dehydrogenase, encoded by G6PD or H6PD, supports the metabolic dependency of GSCs on nucleotides when cultured under acidic conditions, by enhancing the pentose phosphate pathway (PPP). The high level of reduced glutathione (GSH) under acidic conditions also causes demand for the PPP to provide NADPH. Taken together, upregulation of G6PD/H6PD in the PPP plays an important role in acidic-driven purine metabolic reprogramming and confers a predilection toward glioma progression. Our findings indicate that targeting G6PD/H6PD, which are closely related to glioma patient survival, may serve as a promising therapeutic target for improved glioblastoma therapeutics. An integrated metabolomics and metabolic flux analysis, as well as considering microenvironment and cancer stem cells, provide a precise insight into understanding cancer metabolic reprogramming.
    Keywords:  Acidic microenvironment; Glioma stem cells; Glucose-6-phosphate dehydrogenase; Metabolomics; Purine metabolism
    DOI:  https://doi.org/10.1038/s41419-021-03543-9
  3. Nat Commun. 2021 Mar 19. 12(1): 1747
    Melanoma subpopulations that rapidly escape MAPK pathway inhibition incur DNA damage and rely on stress signalling.
    Chen Yang, Chengzhe Tian, Timothy E Hoffman, Nicole K Jacobsen, Sabrina L Spencer.
      Despite the increasing number of effective anti-cancer therapies, successful treatment is limited by the development of drug resistance. While the contribution of genetic factors to drug resistance is undeniable, little is known about how drug-sensitive cells first evade drug action to proliferate in drug. Here we track the responses of thousands of single melanoma cells to BRAF inhibitors and show that a subset of cells escapes drug via non-genetic mechanisms within the first three days of treatment. Cells that escape drug rely on ATF4 stress signalling to cycle periodically in drug, experience DNA replication defects leading to DNA damage, and yet out-proliferate other cells over extended treatment. Together, our work reveals just how rapidly melanoma cells can adapt to drug treatment, generating a mutagenesis-prone subpopulation that expands over time.
    DOI:  https://doi.org/10.1038/s41467-021-21549-x
  4. DNA Repair (Amst). 2021 Mar 13. pii: S1568-7864(21)00055-0. [Epub ahead of print]101 103099
    Chk1 inhibition induces a DNA damage bystander effect in cocultured tumour cells.
    Teresa Brooks, Joanne Wayne, Andrew J Massey.
      Inhibitors of Chk1 kinase, a key effector of the DNA damage response pathway, are currently undergoing Phase 1 and 2 clinical trials as single agents and in combination with cytotoxic chemotherapy. Understanding the biological effects of Chk1 inhibitors on cancer cells is critical for their continued clinical development. Treatment of adherent HT29 or HCC1937 cancer cells or suspension Jurkat or THP1 cells with a Chk1 inhibitor increased γH2AX in these cells. Chk1i pre-treated HCC1937 or HT29 cells resulted in γH2AX induction in cocultured Jurkat or THP1 cells despite these cells never being treated with a Chk1i. Pre-treatment of HT29 cells with camptothecin or gemcitabine followed by a Chk1i increased the DNA damage bystander effect in naïve cocultured THP1 cells compared to camptothecin or gemcitabine alone. This bystander effect appeared to occur through soluble factors via ATR, ATM, and DNA-PKcs activation in the bystander cells. Chk1 silencing by siRNA in HCC1937 or HT29 cells induced a DNA damage bystander effect in cocultured THP1 cells. However, this bystander effect induced by siRNA appeared mechanistically different to that induced by the Chk1 inhibitor. This work suggests that a Chk1 inhibitor-induced bystander effect may increase the clinical effectiveness of Chk1 inhibitors by inducing additional DNA damage or replication stress in cancer cells not directly exposed to the inhibitor. Conversely, it may also contribute to Chk1 inhibitor toxicity by increasing DNA damage in non-tumour cells.
    Keywords:  Bystander effect; Chk1; DNA damage; Kinase inhibitor
    DOI:  https://doi.org/10.1016/j.dnarep.2021.103099
  5. Proc Natl Acad Sci U S A. 2021 Mar 23. pii: e2022600118. [Epub ahead of print]118(12):
    ATM controls the extent of DNA end resection by eliciting sequential posttranslational modifications of CtIP.
    Jinhua Han, Li Wan, Guixing Jiang, Liping Cao, Feiyu Xia, Tian Tian, Xiaomei Zhu, Mingjie Wu, Michael S Y Huen, Yi Wang, Ting Liu, Jun Huang.
      DNA end resection is a critical step in the repair of DNA double-strand breaks (DSBs) via homologous recombination (HR). However, the mechanisms governing the extent of resection at DSB sites undergoing homology-directed repair remain unclear. Here, we show that, upon DSB induction, the key resection factor CtIP is modified by the ubiquitin-like protein SUMO at lysine 578 in a PIAS4-dependent manner. CtIP SUMOylation occurs on damaged chromatin and requires prior hyperphosphorylation by the ATM protein kinase. SUMO-modified hyperphosphorylated CtIP is targeted by the SUMO-dependent E3 ubiquitin ligase RNF4 for polyubiquitination and subsequent degradation. Consequently, disruption of CtIP SUMOylation results in aberrant accumulation of CtIP at DSBs, which, in turn, causes uncontrolled excessive resection, defective HR, and increased cellular sensitivity to DSB-inducing agents. These findings reveal a previously unidentified regulatory mechanism that regulates CtIP activity at DSBs and thus the extent of end resection via ATM-dependent sequential posttranslational modification of CtIP.
    Keywords:  ATM; CtIP; DNA end resection; homologous recombination; hyperphosphorylation
    DOI:  https://doi.org/10.1073/pnas.2022600118
  6. Cell Rep. 2021 Mar 16. pii: S2211-1247(21)00154-6. [Epub ahead of print]34(11): 108840
    DSB repair pathway choice is regulated by recruitment of 53BP1 through cell cycle-dependent regulation of Sp1.
    Michelle L Swift, Kate Beishline, Samuel Flashner, Jane Azizkhan-Clifford.
      Although many of the factors, epigenetic changes, and cell cycle stages that distinguish repair of double-strand breaks (DSBs) by homologous recombination (HR) from non-homologous end joining (NHEJ) are known, the underlying mechanisms that determine pathway choice are incompletely understood. Previously, we found that the transcription factor Sp1 is recruited to DSBs and is necessary for repair. Here, we demonstrate that Sp1 localizes to DSBs in G1 and is necessary for recruitment of the NHEJ repair factor, 53BP1. Phosphorylation of Sp1-S59 in early S phase evicts Sp1 and 53BP1 from the break site; inhibition of that phosphorylation results in 53BP1 and Sp1 remaining at DSBs in S phase cells, precluding BRCA1 binding and suppressing HR. Expression of Sp1-S59A increases sensitivity of BRCA1+/+ cells to poly (ADP-ribose) polymerase (PARP) inhibition similar to BRCA1 deficiency. These data demonstrate how Sp1 integrates the cell cycle and DSB repair pathway choice to favor NHEJ.
    Keywords:  53BP1; BRCA1; DSB repair; HR; NHEJ; Sp1; cell cycle; synthetic lethality
    DOI:  https://doi.org/10.1016/j.celrep.2021.108840
  7. Biochem J. 2021 Mar 26. 478(6): 1179-1197
    Mitochondrial genome stability in human: understanding the role of DNA repair pathways.
    Sumedha Dahal, Sathees C Raghavan.
      Mitochondria are semiautonomous organelles in eukaryotic cells and possess their own genome that replicates independently. Mitochondria play a major role in oxidative phosphorylation due to which its genome is frequently exposed to oxidative stress. Factors including ionizing radiation, radiomimetic drugs and replication fork stalling can also result in different types of mutations in mitochondrial DNA (mtDNA) leading to genome fragility. Mitochondria from myopathies, dystonia, cancer patient samples show frequent mtDNA mutations such as point mutations, insertions and large-scale deletions that could account for mitochondria-associated disease pathogenesis. The mechanism by which such mutations arise following exposure to various DNA-damaging agents is not well understood. One of the well-studied repair pathways in mitochondria is base excision repair. Other repair pathways such as mismatch repair, homologous recombination and microhomology-mediated end joining have also been reported. Interestingly, nucleotide excision repair and classical nonhomologous DNA end joining are not detected in mitochondria. In this review, we summarize the potential causes of mitochondrial genome fragility, their implications as well as various DNA repair pathways that operate in mitochondria.
    Keywords:  base excision repair; double-strand breaks; homologous recombination; microhomology-mediated end joining; mitochondrial diseases; nonhomologous end joining
    DOI:  https://doi.org/10.1042/BCJ20200920
  8. Cell Rep. 2021 Mar 16. pii: S2211-1247(21)00178-9. [Epub ahead of print]34(11): 108864
    Excision of mutagenic replication-blocking lesions suppresses cancer but promotes cytotoxicity and lethality in nitrosamine-exposed mice.
    Jennifer E Kay, Joshua J Corrigan, Amanda L Armijo, Ilana S Nazari, Ishwar N Kohale, Dorothea K Torous, Svetlana L Avlasevich, Robert G Croy, Dushan N Wadduwage, Sebastian E Carrasco, Stephen D Dertinger, Forest M White, John M Essigmann, Leona D Samson, Bevin P Engelward.
      N-Nitrosodimethylamine (NDMA) is a DNA-methylating agent that has been discovered to contaminate water, food, and drugs. The alkyladenine DNA glycosylase (AAG) removes methylated bases to initiate the base excision repair (BER) pathway. To understand how gene-environment interactions impact disease susceptibility, we study Aag-knockout (Aag-/-) and Aag-overexpressing mice that harbor increased levels of either replication-blocking lesions (3-methyladenine [3MeA]) or strand breaks (BER intermediates), respectively. Remarkably, the disease outcome switches from cancer to lethality simply by changing AAG levels. To understand the underlying basis for this observation, we integrate a suite of molecular, cellular, and physiological analyses. We find that unrepaired 3MeA is somewhat toxic, but highly mutagenic (promoting cancer), whereas excess strand breaks are poorly mutagenic and highly toxic (suppressing cancer and promoting lethality). We demonstrate that the levels of a single DNA repair protein tip the balance between blocks and breaks and thus dictate the disease consequences of DNA damage.
    Keywords:  DNA damage; DNA strand break; alkyladenine DNA glycosylase; cancer; liver; mutation; nitrosamine; replication-blocking lesion
    DOI:  https://doi.org/10.1016/j.celrep.2021.108864
  9. Cell Death Differ. 2021 Mar 15.
    Beyond DNA repair and chromosome instability-Fanconi anaemia as a cellular senescence-associated syndrome.
    Anne Helbling-Leclerc, Cécile Garcin, Filippo Rosselli.
      Fanconi anaemia (FA) is the most frequent inherited bone marrow failure syndrome, due to mutations in genes encoding proteins involved in replication fork protection, DNA interstrand crosslink repair and replication rescue through inducing double-strand break repair and homologous recombination. Clinically, FA is characterised by aplastic anaemia, congenital defects and cancer predisposition. In in vitro studies, FA cells presented hallmarks defining senescent cells, including p53-p21 axis activation, altered telomere length, mitochondrial dysfunction, chromatin alterations, and a pro-inflammatory status. Senescence is a programme leading to proliferation arrest that is involved in different physiological contexts, such as embryogenesis, tissue remodelling and repair and guarantees tumour suppression activity. However, senescence can become a driving force for developmental abnormalities, aging and cancer. Herein, we summarise the current knowledge in the field to highlight the mutual relationships between FA and senescence that lead us to consider FA not only as a DNA repair and chromosome fragility syndrome but also as a "senescence syndrome".
    DOI:  https://doi.org/10.1038/s41418-021-00764-5
  10. Mol Genet Metab. 2021 Mar 10. pii: S1096-7192(21)00059-7. [Epub ahead of print]
    Bloom syndrome and the underlying causes of genetic instability.
    Mouna Ababou.
      Autosomal hereditary recessive diseases characterized by genetic instability are often associated with cancer predisposition. Bloom syndrome (BS), a rare genetic disorder, with <300 cases reported worldwide, combines both. Indeed, patients with Bloom's syndrome are 150 to 300 times more likely to develop cancers than normal individuals. The wide spectrum of cancers developed by BS patients suggests that early initial events occur in BS cells which may also be involved in the initiation of carcinogenesis in the general population and these may be common to several cancers. BS is caused by mutations of both copies of the BLM gene, encoding the RecQ BLM helicase. This review discusses the different aspects of BS and the different cellular functions of BLM in genome surveillance and maintenance through its major roles during DNA replication, repair, and transcription. BLM's activities are essential for the stabilization of centromeric, telomeric and ribosomal DNA sequences, and the regulation of innate immunity. One of the key objectives of this work is to establish a link between BLM functions and the main clinical phenotypes observed in BS patients, as well as to shed new light on the correlation between the genetic instability and diseases such as immunodeficiency and cancer. The different potential implications of the BLM helicase in the tumorigenic process and the use of BLM as new potential target in the field of cancer treatment are also debated.
    Keywords:  Bloom syndrome; Cancer predisposition; DNA replication and repair; Genetic instability; RecQ BLM helicase; Transcription
    DOI:  https://doi.org/10.1016/j.ymgme.2021.03.003
  11. Curr Genet. 2021 Mar 14.
    Histones on fire: the effect of Dun1 and Mrc1 on origin firing and replication of hyper-acetylated genomes.
    Lihi Gershon, Martin Kupiec.
      As cells replicate their DNA, there is a need to synthesize new histones with which to wrap it. Newly synthesized H3 histones that are incorporated into the assembling chromatin behind the replication fork are acetylated at lysine 56. The acetylation is removed by two deacetylases, Hst3 and Hst4. This process is tightly regulated and any perturbation leads to genomic instability and replicative stress. We recently showed that Dun1, a kinase implicated mainly in the regulation of dNTPs, is vital in cells with hyper-acetylation, to counteract Rad53's inhibition on late-firing origins of replication. Our work showed that ∆hst3 ∆hst4 cells depend on late origin firing for survival, and are unable to prevent Rad53's inhibition when Dun1 is inactive. Thus, our work describes a role for Dun1 that is independent on its known function as a regulator of dNTP levels. Here we show that Mrc1 (Claspin in mammals), a protein that moves with the replicating fork and participates in both replication and checkpoint functions, plays also an essential role in the absence of H3K56Ac deacetylation. The sum of the results shown here and in our recent publication suggests that dormant origins are also utilized in these cells, making Mrc1, which regulates firing from these origins, also essential when histone H3 is hyper-acetylated. Thus, cells suffering from hyper-acetylation of H3K56 experience replication stress caused by a combination of prone-to-collapse forks and limited replication tracts. This combination makes both Dun1 and Mrc1, each acting on different targets, essential for viability.
    Keywords:  CTF18; DBF4; H3K56 acetylation; RTT101; SLD3; Saccharomyces cerevisiae; Yeast
    DOI:  https://doi.org/10.1007/s00294-021-01175-2
  12. Mutagenesis. 2021 Mar 19. pii: geab011. [Epub ahead of print]
    NEK1 deficiency affects mitochondrial functions and the transcriptome of key DNA repair pathways.
    Mariana Bonjiorno Martins, Arina Marina Perez, Vilhelm A Bohr, David M Wilson Iii, Jörg Kobarg.
      Previous studies have indicated important roles for NIMA-related kinase 1 (NEK1) in modulating DNA damage checkpoints and DNA repair capacity. To broadly assess the contributions of NEK1 to genotoxic stress and mitochondrial functions, we characterized several relevant phenotypes of NEK1 CRISPR knockout (KO) and WT HAP1 cells. Our studies revealed that NEK1 KO cells resulted in increased apoptosis and hypersensitivity to the alkylator methyl methanesulfonate, the radiomimetic bleomycin, and UVC light, yet increased resistance to the crosslinker cisplatin. Mitochondrial functionalities were also altered in NEK1 KO cells, with phenotypes of reduced mitophagy, increased total mitochondria, elevated levels of reactive oxygen species, impaired complex I activity, and higher amounts of mitochondrial DNA damage. RNA-seq transcriptome analysis coupled with qRT-PCR studies comparing NEK1 KO cells with NEK1 overexpressing cells revealed that the expression of genes involved in DNA repair pathways, such as base excision repair, nucleotide excision repair, and double-strand break repair, are altered in a way that might influence genotoxin resistance. Together, our studies underline and further support that NEK1 serves as a hub signaling kinase in response to DNA damage, modulating DNA repair capacity, mitochondrial activity and cell fate determination.
    Keywords:  DNA Damage; DNA Repair; Mitochondria; NEK1
    DOI:  https://doi.org/10.1093/mutage/geab011
  13. Front Cell Dev Biol. 2021 ;9 640755
    Human RecQ Helicases in DNA Double-Strand Break Repair.
    Huiming Lu, Anthony J Davis.
      RecQ DNA helicases are a conserved protein family found in bacteria, fungus, plants, and animals. These helicases play important roles in multiple cellular functions, including DNA replication, transcription, DNA repair, and telomere maintenance. Humans have five RecQ helicases: RECQL1, Bloom syndrome protein (BLM), Werner syndrome helicase (WRN), RECQL4, and RECQL5. Defects in BLM and WRN cause autosomal disorders: Bloom syndrome (BS) and Werner syndrome (WS), respectively. Mutations in RECQL4 are associated with three genetic disorders, Rothmund-Thomson syndrome (RTS), Baller-Gerold syndrome (BGS), and RAPADILINO syndrome. Although no genetic disorders have been reported due to loss of RECQL1 or RECQL5, dysfunction of either gene is associated with tumorigenesis. Multiple genetically independent pathways have evolved that mediate the repair of DNA double-strand break (DSB), and RecQ helicases play pivotal roles in each of them. The importance of DSB repair is supported by the observations that defective DSB repair can cause chromosomal aberrations, genomic instability, senescence, or cell death, which ultimately can lead to premature aging, neurodegeneration, or tumorigenesis. In this review, we will introduce the human RecQ helicase family, describe in detail their roles in DSB repair, and provide relevance between the dysfunction of RecQ helicases and human diseases.
    Keywords:  BLM; DNA double-strand break repair; RECQL1; RECQL4; RECQL5; RecQ helicase; WRN; genome stability
    DOI:  https://doi.org/10.3389/fcell.2021.640755
  14. Nat Commun. 2021 Mar 19. 12(1): 1746
    Stabilisation of half MCM ring by Cdt1 during DNA insertion.
    Marina Guerrero-Puigdevall, Narcis Fernandez-Fuentes, Jordi Frigola.
      Origin licensing ensures precise once per cell cycle replication in eukaryotic cells. The Origin Recognition Complex, Cdc6 and Cdt1 load Mcm2-7 helicase (MCM) into a double hexamer, bound around duplex DNA. The complex formed by ORC-Cdc6 bound to duplex DNA (OC) recruits the MCM-Cdt1 complex into the replication origins. Through the stacking of both complexes, the duplex DNA is inserted inside the helicase by an unknown mechanism. In this paper we show that the DNA insertion comes with a topological problem in the stacking of OC with MCM-Cdt1. Unless an essential, conserved C terminal winged helix domain (C-WHD) of Cdt1 is present, the MCM splits into two halves. The binding of this domain with the essential C-WHD of Mcm6, allows the latching between the MCM-Cdt1 and OC, through a conserved Orc5 AAA-lid interaction. Our work provides new insights into how DNA is inserted into the eukaryotic replicative helicase, through a series of synchronized events.
    DOI:  https://doi.org/10.1038/s41467-021-21932-8
  15. Cancer Res. 2021 Mar 15. 81(6): 1441-1442
    Regulating Polθ in Breast Cancer.
    Denisse Carvajal-Maldonado, Richard D Wood.
      DNA polymerase θ, a protein encoded by the POLQ gene, is the defining factor for the DNA double-strand break repair pathway known as theta-mediated end-joining (TMEJ). Some cancers depend on TMEJ for survival and tumor growth. TMEJ might be useful as a biomarker to guide patient treatment and is now an active target for drug development, making it critical to understand how it is regulated in cells. In a recent article, Prodhomme and colleagues provide the first identification of a transcription regulator of POLQ expression and TMEJ activity: the transcription factor, ZEB1.See related article by Prodhomme et al., p. 1595.
    DOI:  https://doi.org/10.1158/0008-5472.CAN-20-4253
  16. Commun Biol. 2021 Mar 17. 4(1): 349
    Pol α-primase dependent nuclear localization of the mammalian CST complex.
    Joseph M Kelich, Harry Papaioannou, Emmanuel Skordalakes.
      The human CST complex composed of CTC1, STN1, and TEN1 is critically involved in telomere maintenance and homeostasis. Specifically, CST terminates telomere extension by inhibiting telomerase access to the telomeric overhang and facilitates lagging strand fill in by recruiting DNA Polymerase alpha primase (Pol α-primase) to the telomeric C-strand. Here we reveal that CST has a dynamic intracellular localization that is cell cycle dependent. We report an increase in nuclear CST several hours after the initiation of DNA replication, followed by exit from the nucleus prior to mitosis. We identify amino acids of CTC1 involved in Pol α-primase binding and nuclear localization. We conclude, the CST complex does not contain a nuclear localization signal (NLS) and suggest that its nuclear localization is reliant on Pol α-primase. Hypomorphic mutations affecting CST nuclear import are associated with telomere syndromes and cancer, emphasizing the important role of this process in health.
    DOI:  https://doi.org/10.1038/s42003-021-01845-4
  17. Sci Rep. 2021 Mar 18. 11(1): 6317
    Karonudib has potent anti-tumor effects in preclinical models of B-cell lymphoma.
    Morten P Oksvold, Ulrika Warpman Berglund, Helge Gad, Baoyan Bai, Trond Stokke, Idun Dale Rein, Therese Pham, Kumar Sanjiv, Geir Frode Øy, Jens Henrik Norum, Erlend B Smeland, June H Myklebust, Thomas Helleday, Thea Kristin Våtsveen.
      Chemo-immunotherapy has improved survival in B-cell lymphoma patients, but refractory/relapsed diseases still represent a major challenge, urging for development of new therapeutics. Karonudib (TH1579) was developed to inhibit MTH1, an enzyme preventing oxidized dNTP-incorporation in DNA. MTH1 is highly upregulated in tumor biopsies from patients with diffuse large B-cell lymphoma (DLBCL) and Burkitt lymphoma, hence confirming a rationale for targeting MTH1. Here, we tested the efficacy of karonudib in vitro and in preclinical B-cell lymphoma models. Using a range of B-cell lymphoma cell lines, karonudib strongly reduced viability at concentrations well tolerated by activated normal B cells. In B-cell lymphoma cells, karonudib increased incorporation of 8-oxo-dGTP into DNA, and prominently induced prometaphase arrest and apoptosis due to failure in spindle assembly. MTH1 knockout cell lines were less sensitive to karonudib-induced apoptosis, but were displaying cell cycle arrest phenotype similar to the wild type cells, indicating a dual inhibitory role of the drug. Karonudib was highly potent as single agent in two different lymphoma xenograft models, including an ABC DLBCL patient derived xenograft, leading to prolonged survival and fully controlled tumor growth. Together, our preclinical findings provide a rationale for further clinical testing of karonudib in B-cell lymphoma.
    DOI:  https://doi.org/10.1038/s41598-021-85613-8
  18. Adv Exp Med Biol. 2021 ;1286 15-48
    Targeting Cancer Metabolism and Current Anti-Cancer Drugs.
    Witchuda Sukjoi, Jarunya Ngamkham, Paul V Attwood, Sarawut Jitrapakdee.
      Several studies have exploited the metabolic hallmarks that distinguish between normal and cancer cells, aiming at identifying specific targets of anti-cancer drugs. It has become apparent that metabolic flexibility allows cancer cells to survive during high anabolic demand or the depletion of nutrients and oxygen. Cancers can reprogram their metabolism to the microenvironments by increasing aerobic glycolysis to maximize ATP production, increasing glutaminolysis and anabolic pathways to support bioenergetic and biosynthetic demand during rapid proliferation. The increased key regulatory enzymes that support the relevant pathways allow us to design small molecules which can specifically block activities of these enzymes, preventing growth and metastasis of tumors. In this review, we discuss metabolic adaptation in cancers and highlight the crucial metabolic enzymes involved, specifically those involved in aerobic glycolysis, glutaminolysis, de novo fatty acid synthesis, and bioenergetic pathways. Furthermore, we also review the success and the pitfalls of the current anti-cancer drugs which have been applied in pre-clinical and clinical studies.
    Keywords:  Bioenergetic; Cancer; Drug target; Enzymes; Metabolism
    DOI:  https://doi.org/10.1007/978-3-030-55035-6_2
  19. Nucleic Acids Res. 2021 Mar 18. 49(5): 2450-2459
    Reversible chromatin condensation by the DNA repair and demethylation factor thymine DNA glycosylase.
    Charles E Deckard, Jonathan T Sczepanski.
      Chromatin structures (and modulators thereof) play a central role in genome organization and function. Herein, we report that thymine DNA glycosylase (TDG), an essential enzyme involved in DNA repair and demethylation, has the capacity to alter chromatin structure directly through its physical interactions with DNA. Using chemically defined nucleosome arrays, we demonstrate that TDG induces decompaction of individual chromatin fibers upon binding and promotes self-association of nucleosome arrays into higher-order oligomeric structures (i.e. condensation). Chromatin condensation is mediated by TDG's disordered polycationic N-terminal domain, whereas its C-terminal domain antagonizes this process. Furthermore, we demonstrate that TDG-mediated chromatin condensation is reversible by growth arrest and DNA damage 45 alpha (GADD45a), implying that TDG cooperates with its binding partners to dynamically control chromatin architecture. Finally, we show that chromatin condensation by TDG is sensitive to the methylation status of the underlying DNA. This new paradigm for TDG has specific implications for associated processes, such as DNA repair, DNA demethylation, and transcription, and general implications for the role of DNA modification 'readers' in controlling chromatin organization.
    DOI:  https://doi.org/10.1093/nar/gkab040
  20. Cell Biochem Biophys. 2021 Mar 17.
    Terminal Peptide Extensions Augment the Retinal IMPDH1 Catalytic Activity and Attenuate the ATP-induced Fibrillation Events.
    Behnaz Andashti, Razieh Yazdanparast, Maede Motahar, Ebrahim Barzegari, Hamid Galehdari.
      Defects in inosine monophosphate dehydrogenase-1 (IMPDH1) lead to insufficient biosyntheses of purine nucleotides. In eyes, these defects are believed to cause retinitis pigmentosa (RP). Major retinal isoforms of IMPDH1 are structurally distinct from those in other tissues, by bearing terminal extensions. Using recombinant mouse IMPDH1 (mH1), we evaluated the kinetics and oligomerization states of the retinal isoforms. Moreover, we adopted molecular simulation tools to study the possible effect of terminal tails on the function of major enzyme isoforms with the aim to find structural evidence in favor of contradictory observations on retinal IMPDH1 function. Our findings indicated higher catalytic activity for the major mouse retinal isoform (mH1603) along with lower fibrillation capacity under the influence of ATP. However, higher mass oligomerization products were formed by the mH1 (603) isoform in the presence of the enzyme inhibitors such as GTP and/or MPA. Collectively, our findings demonstrate that the structural differences between the retinal isoforms have led to functional variations possibly to justify the retinal cells' requirements.
    Keywords:  Catalytic activity; IMPDH1; Retinal isoform; Retinitis Pigmentosa; Terminal tail
    DOI:  https://doi.org/10.1007/s12013-021-00973-2
  21. Biochem Biophys Res Commun. 2021 Mar 11. pii: S0006-291X(21)00375-2. [Epub ahead of print]551 33-37
    Structural basis for the interaction modes of dihydroorotase with the anticancer drugs 5-fluorouracil and 5-aminouracil.
    Hong-Hsiang Guan, Yen-Hua Huang, En-Shyh Lin, Chun-Jung Chen, Cheng-Yang Huang.
      Dihydroorotase (DHOase) is the third enzyme in the de novo biosynthesis pathway of pyrimidine nucleotides and considered an attractive target for potential antimalarial, anticancer, and antipathogen chemotherapy. Whether the FDA-approved clinical drug 5-fluorouracil (5-FU) that is used to target the enzyme thymidylate synthase for anticancer therapy can also bind to DHOase remains unknown. Here, we report the crystal structures of DHOase from Saccharomyces cerevisiae (ScDHOase) complexed with malate, 5-FU, and 5-aminouracil (5-AU). ScDHOase shares structural similarity with Escherichia coli DHOase. We also characterized the binding of 5-FU and 5-AU to ScDHOase by using the fluorescence quenching method. These complexed structures revealed that residues Arg18, Asn43, Thr106, and Ala275 of ScDHOase were involved in the 5-FU (PDB entry 6L0B) and 5-AU binding (PDB entry 6L0F). Overall, these results provide structural insights that may facilitate the development of new inhibitors targeting DHOase and constitute the 5-FU and 5-AU interactomes for further clinical chemotherapies.
    Keywords:  5-Aminouracil; 5-Fluorouracil; Anticancer; Dihydroorotase; Dihydropyrimidinase; Pyrimidine biosynthesis
    DOI:  https://doi.org/10.1016/j.bbrc.2021.03.001
  22. Biochim Biophys Acta Bioenerg. 2021 Mar 12. pii: S0005-2728(21)00043-8. [Epub ahead of print] 148410
    Metabolic implications of non-Electrogenic ATP/ADP exchange in Cancer cells: A mechanistic basis for the Warburg effect.
    John J Lemasters.
      In post-mitotic cells, mitochondrial ATP/ADP exchange occurs by the adenine nucleotide translocator (ANT). Driven by membrane potential (ΔΨ), ANT catalyzes electrogenic exchange of ATP4- for ADP3-, leading to higher ATP/ADP ratios in the cytosol than mitochondria. In cancer cells, ATP/ADP exchange occurs not by ANT but likely via the non-electrogenic ATP-Mg/phosphate carrier. Consequences of non-electrogenic exchange are: 1) Cytosolic ATP/ADP decreases to stimulate aerobic glycolysis. 2) Without proton utilization for exchange, ATP/O increases by 35% for complete glucose oxidation. 3) Decreased cytosolic ATP/ADP•Pi increases NAD(P)H/NAD(P)+. Increased NADH increases lactate/pyruvate, and increased NADPH promotes anabolic metabolism. Fourth, increased mitochondrial NADH/NAD+ magnifies the redox span across Complexes I and III, which increases ΔΨ, reactive oxygen species generation, and susceptibility to ferroptosis. 5) Increased mitochondrial NADPH/NADP+ favors a reverse isocitrate dehydrogenase-2 reaction with citrate accumulation and export for biomass formation. Consequently, 2-oxoglutarate formation occurs largely via oxidation of glutamine, the preferred respiratory substrate of cancer cells. Overall, non-electrogenic ATP/ADP exchange promotes aerobic glycolysis (Warburg effect) and confers growth advantages to cancer cells.
    Keywords:  ATP/ADP exchange; Aerobic glycolysis; Cancer; Glutamine; Mitochondria; Warburg effect
    DOI:  https://doi.org/10.1016/j.bbabio.2021.148410
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