bims-numges Biomed News
on Nucleotide metabolism and genome stability
Issue of 2022‒04‒24
thirty papers selected by
Sean Rudd
Karolinska Institutet
  1. Disruption of dNTPs Homeostasis by Ribonucleotide Reductase Hyperactivation Overcomes AML Differentiation Blockade.
  2. DNA replication is highly resilient and persistent under the challenge of mild replication stress.
  3. A POLD3/BLM dependent pathway handles DSBs in transcribed chromatin upon excessive RNA:DNA hybrid accumulation.
  4. MCMBP promotes the assembly of the MCM2-7 hetero-hexamer to ensure robust DNA replication in human cells.
  5. Insights into Mechanisms of Damage Recognition and Catalysis by APE1-like Enzymes.
  6. Break-induced replication: unraveling each step.
  7. Consequences of telomere replication failure: the other end-replication problem.
  8. Sources, resolution and physiological relevance of R-loops and RNA-DNA hybrids.
  9. hMSH5 Regulates NHEJ and Averts Excessive Nucleotide Alterations at Repair Joints.
  10. H3K4 methylation by SETD1A/BOD1L facilitates RIF1-dependent NHEJ.
  11. Nej1 interacts with Sae2 at DNA double-stranded breaks to inhibit DNA resection.
  12. Integrating Sister Chromatid Cohesion Establishment to DNA Replication.
  13. CDYL1-dependent decrease in lysine crotonylation at DNA double-strand break sites functionally uncouples transcriptional silencing and repair.
  14. p53 at the crossroad of DNA replication and ribosome biogenesis stress pathways.
  15. DNA damage response and repair in the development and treatment of brain tumors.
  16. Purine Synthesis Inhibitor L-Alanosine Impairs Mitochondrial Function and Stemness of Brain Tumor Initiating Cells.
  17. Heat-induced SIRT1-mediated H4K16ac deacetylation impairs resection and SMARCAD1 recruitment to double strand breaks.
  18. Development and Evolution of DNA-Dependent Protein Kinase Inhibitors toward Cancer Therapy.
  19. Stable G-quadruplex DNA structures promote replication-dependent genome instability.
  20. Kinetic Analysis of the Effect of N-Terminal Acetylation on Thymine DNA Glycosylase.
  21. Heterochromatic repeat clustering imposes a physical barrier on homologous recombination to prevent chromosomal translocations.
  22. Replicative Stress Coincides with Impaired Nuclear DNA Damage Response in COX4-1 Deficiency.
  23. Glucose Increases STAT3 Activation, Promoting Sustained XRCC1 Expression and Increasing DNA Repair.
  24. VGLL3 increases the dependency of cancer cells on de novo nucleotide synthesis through GART expression.
  25. CCNE1 amplification is synthetic lethal with PKMYT1 kinase inhibition.
  26. Artesunate improves venetoclax plus cytarabine AML cell targeting by regulating the Noxa/Bim/Mcl-1/p-Chk1 axis.
  27. TOP1-DNA trapping by exatecan and combination therapy with ATR inhibitor.
  28. Inosine triphosphate pyrophosphatase from Trypanosoma brucei cleanses cytosolic pools from deaminated nucleotides.
  29. Mismatch discrimination and sequence bias during end-joining by DNA ligases.
  30. Purine nucleoside analogs plus rituximab are an effective treatment choice for hairy cell leukemia-variant.
  1. Blood. 2022 Apr 19. pii: blood.2021015108. [Epub ahead of print]
    Disruption of dNTPs Homeostasis by Ribonucleotide Reductase Hyperactivation Overcomes AML Differentiation Blockade.
    Hanying Wang, Xin He, Lei Zhang, Haojie Dong, Feiteng Huang, Jie Xian, Min Li, Wei Chen, Xiyuan Lu, Khyatiben V Pathak, Wenfeng Huang, Zheng Li, Lianjun Zhang, Le Xuan Truong Nguyen, Lu Yang, Lifeng Feng, David Gordon, Jing Zhang, Patrick Pirrotte, Chun-Wei Chen, Amandeep Salhotra, Ya-Huei Kuo, David A Horne, Guido Marcucci, David B Sykes, Stefano Tiziani, Hongchuan Jin, Xian Wang, Ling Li.
      Differentiation blockade is a hallmark of AML. Strategy to overcome such blockade is a promising approach against the disease. Lack of understanding underlying mechanisms hampers development of such strategy. Dysregulated ribonucleotide reductase (RNR) is considered a druggable target in proliferative cancers susceptible to deoxynucleoside triphosphates (dNTPs) depletion. Herein, we report an unanticipated discovery that hyperactivating RNR enables differentiation and decreases leukemia cell growth. We integrate pharmacogenomics and metabolomics analyses to identify that pharmacologically (e.g., nelarabine) or genetically upregulating RNR subunit M2 (RRM2) level, creates a dNTP pool imbalance and overcomes differentiation arrest. Moreover, R-loop mediated DNA replication stress signaling is responsible for RRM2 activation by nelarabine treatment. Further aggravating dNTP imbalance by depleting the dNTP hydrolase SAM domain and HD domain-containing protein 1 (SAMHD1) enhances ablation of leukemia stem cells by RRM2 hyperactivation. Mechanistically, excessive activation of ERK signaling downstream of the imbalance contributes to cellular outcomes of RNR hyperactivation. A CRISPR screen identifies a synthetic lethal interaction between loss of DUSP6, an ERK negative regulator, and nelarabine treatment. These data demonstrate that dNTP homeostasis governs leukemia maintenance, and combination of DUSP inhibition and nelarabine represents a therapeutic strategy.
    DOI:  https://doi.org/10.1182/blood.2021015108
  2. Cell Rep. 2022 Apr 19. pii: S2211-1247(22)00459-4. [Epub ahead of print]39(3): 110701
    DNA replication is highly resilient and persistent under the challenge of mild replication stress.
    Camelia Mocanu, Eleftheria Karanika, María Fernández-Casañas, Alex Herbert, Tomisin Olukoga, Mete Emir Özgürses, Kok-Lung Chan.
      Mitotic DNA synthesis (MiDAS) has been proposed to restart DNA synthesis during mitosis because of replication fork stalling in late interphase caused by mild replication stress (RS). Contrary to this proposal, we find that cells exposed to mild RS in fact maintain continued DNA replication throughout G2 and during G2-M transition in RAD51- and RAD52-dependent manners. Persistent DNA synthesis is necessary to resolve replication intermediates accumulated in G2 and disengage an ATR-imposed block to mitotic entry. Because of its continual nature, DNA synthesis at very late replication sites can overlap with chromosome condensation, generating the phenomenon of mitotic DNA synthesis. Unexpectedly, we find that the commonly used CDK1 inhibitor RO3306 interferes with replication to preclude detection of G2 DNA synthesis, leading to the impression of a mitosis-driven response. Our study reveals the importance of persistent DNA replication and checkpoint control to lessen the risk for severe genome under-replication under mild RS.
    Keywords:  ATR-mediated replication checkpoint; CDK1 inhibition; CP: Molecular biology; G2 DNA synthesis; MiDAS; RAD51; RAD52; RO3306; S-to-M DNA synthesis runover; aphidicolin; mild replication stress
    DOI:  https://doi.org/10.1016/j.celrep.2022.110701
  3. Nat Commun. 2022 Apr 19. 13(1): 2012
    A POLD3/BLM dependent pathway handles DSBs in transcribed chromatin upon excessive RNA:DNA hybrid accumulation.
    S Cohen, A Guenolé, I Lazar, A Marnef, T Clouaire, D V Vernekar, N Puget, V Rocher, C Arnould, M Aguirrebengoa, M Genais, N Firmin, R A Shamanna, R Mourad, V A Bohr, V Borde, G Legube.
      Transcriptionally active loci are particularly prone to breakage and mounting evidence suggests that DNA Double-Strand Breaks arising in active genes are handled by a dedicated repair pathway, Transcription-Coupled DSB Repair (TC-DSBR), that entails R-loop accumulation and dissolution. Here, we uncover a function for the Bloom RecQ DNA helicase (BLM) in TC-DSBR in human cells. BLM is recruited in a transcription dependent-manner at DSBs where it fosters resection, RAD51 binding and accurate Homologous Recombination repair. However, in an R-loop dissolution-deficient background, we find that BLM promotes cell death. We report that upon excessive RNA:DNA hybrid accumulation, DNA synthesis is enhanced at DSBs, in a manner that depends on BLM and POLD3. Altogether our work unveils a role for BLM at DSBs in active chromatin, and highlights the toxic potential of RNA:DNA hybrids that accumulate at transcription-associated DSBs.
    DOI:  https://doi.org/10.1038/s41467-022-29629-2
  4. Elife. 2022 Apr 19. pii: e77393. [Epub ahead of print]11
    MCMBP promotes the assembly of the MCM2-7 hetero-hexamer to ensure robust DNA replication in human cells.
    Yuichiro Saito, Venny Santosa, Kei-Ichiro Ishiguro, Masato T Kanemaki.
      The MCM2-7 hetero-hexamer is the replicative DNA helicase that plays a central role in eukaryotic DNA replication. In proliferating cells, the expression level of the MCM2-7 hexamer is kept high, which safeguards the integrity of the genome. However, how the MCM2-7 hexamer is assembled in living cells remains unknown. Here, we revealed that the MCM-binding protein (MCMBP) plays a critical role in the assembly of this hexamer in human cells. MCMBP associates with MCM3 which is essential for maintaining the level of the MCM2-7 hexamer. Acute depletion of MCMBP demonstrated that it contributes to MCM2-7 assembly using nascent MCM3. Cells depleted of MCMBP gradually ceased to proliferate because of reduced replication licensing. Under this condition, p53-positive cells exhibited arrest in the G1 phase, whereas p53-null cells entered the S phase and lost their viability because of the accumulation of DNA damage, suggesting that MCMBP is a potential target for killing p53-deficient cancers.
    Keywords:  DNA replication; MCM2–7; MCMBP; cell biology; chromosomes; gene expression; genome instability; human; protein complex assembly
    DOI:  https://doi.org/10.7554/eLife.77393
  5. Int J Mol Sci. 2022 Apr 14. pii: 4361. [Epub ahead of print]23(8):
    Insights into Mechanisms of Damage Recognition and Catalysis by APE1-like Enzymes.
    Anatoly A Bulygin, Olga S Fedorova, Nikita A Kuznetsov.
      Apurinic/apyrimidinic (AP) endonucleases are the key DNA repair enzymes in the base excision repair (BER) pathway, and are responsible for hydrolyzing phosphodiester bonds on the 5' side of an AP site. The enzymes can recognize not only AP sites but also some types of damaged bases, such as 1,N6-ethenoadenosine, α-adenosine, and 5,6-dihydrouridine. Here, to elucidate the mechanism underlying such a broad substrate specificity as that of AP endonucleases, we performed a computational study of four homologous APE1-like endonucleases: insect (Drosophila melanogaster) Rrp1, amphibian (Xenopus laevis) APE1 (xAPE1), fish (Danio rerio) APE1 (zAPE1), and human APE1 (hAPE1). The contact between the amino acid residues of the active site of each homologous APE1-like enzyme and the set of damaged DNA substrates was analyzed. A comparison of molecular dynamic simulation data with the known catalytic efficiency of these enzymes allowed us to gain a deep insight into the differences in the efficiency of the cleavage of various damaged nucleotides. The obtained data support that the amino acid residues within the "damage recognition" loop containing residues Asn222-Ala230 significantly affect the catalytic-complex formation. Moreover, every damaged nucleotide has its unique position and a specific set of interactions with the amino acid residues of the active site.
    Keywords:  active-site plasticity; apurinic/apyrimidinic endonuclease; base excision repair; conformational dynamics; damaged nucleotide; nucleotide eversion; nucleotide incision activity
    DOI:  https://doi.org/10.3390/ijms23084361
  6. Trends Genet. 2022 Apr 19. pii: S0168-9525(22)00072-5. [Epub ahead of print]
    Break-induced replication: unraveling each step.
    Liping Liu, Anna Malkova.
      Break-induced replication (BIR) repairs one-ended double-strand DNA breaks through invasion into a homologous template followed by DNA synthesis. Different from S-phase replication, BIR copies the template DNA in a migrating displacement loop (D-loop) and results in conservative inheritance of newly synthesized DNA. This unusual mode of DNA synthesis makes BIR a source of various genetic instabilities like those associated with cancer in humans. This review focuses on recent progress in delineating the mechanism of Rad51-dependent BIR in budding yeast. In addition, we discuss new data that describe changes in BIR efficiency and fidelity on encountering replication obstacles as well as the implications of these findings for BIR-dependent processes such as telomere maintenance and the repair of collapsed replication forks.
    Keywords:  break-induced replication (BIR); kinetics and rate of BIR; lagging-strand BIR synthesis; replication obstacles; yeast
    DOI:  https://doi.org/10.1016/j.tig.2022.03.011
  7. Trends Biochem Sci. 2022 Apr 16. pii: S0968-0004(22)00072-X. [Epub ahead of print]
    Consequences of telomere replication failure: the other end-replication problem.
    Kirsten A Brenner, Jayakrishnan Nandakumar.
      Telomeres are chromosome-capping structures that protect ends of the linear genome from DNA damage sensors. However, these structures present obstacles during DNA replication. Incomplete telomere replication accelerates telomere shortening and limits replicative lifespan. Therefore, continued proliferation under conditions of replication stress requires a means of telomere repair, particularly in the absence of telomerase. It was recently revealed that replication stress triggers break-induced replication (BIR) and mitotic DNA synthesis (MiDAS) at mammalian telomeres; however, these mechanisms are error prone and primarily utilized in tumorigenic contexts. In this review article, we discuss the consequences of replication stress at telomeres and how use of available repair pathways contributes to genomic instability. Current research suggests that fragile telomeres are ultimately tumor-suppressive and thus may be better left unrepaired.
    Keywords:  alternative lengthening of telomeres; break-induced replication; mitotic DNA synthesis; replication stress; telomeres
    DOI:  https://doi.org/10.1016/j.tibs.2022.03.013
  8. Nat Rev Mol Cell Biol. 2022 Apr 22.
    Sources, resolution and physiological relevance of R-loops and RNA-DNA hybrids.
    Eva Petermann, Li Lan, Lee Zou.
      RNA-DNA hybrids are generated during transcription, DNA replication and DNA repair and are crucial intermediates in these processes. When RNA-DNA hybrids are stably formed in double-stranded DNA, they displace one of the DNA strands and give rise to a three-stranded structure called an R-loop. R-loops are widespread in the genome and are enriched at active genes. R-loops have important roles in regulating gene expression and chromatin structure, but they also pose a threat to genomic stability, especially during DNA replication. To keep the genome stable, cells have evolved a slew of mechanisms to prevent aberrant R-loop accumulation. Although R-loops can cause DNA damage, they are also induced by DNA damage and act as key intermediates in DNA repair such as in transcription-coupled repair and RNA-templated DNA break repair. When the regulation of R-loops goes awry, pathological R-loops accumulate, which contributes to diseases such as neurodegeneration and cancer. In this Review, we discuss the current understanding of the sources of R-loops and RNA-DNA hybrids, mechanisms that suppress and resolve these structures, the impact of these structures on DNA repair and genome stability, and opportunities to therapeutically target pathological R-loops.
    DOI:  https://doi.org/10.1038/s41580-022-00474-x
  9. Genes (Basel). 2022 Apr 11. pii: 673. [Epub ahead of print]13(4):
    hMSH5 Regulates NHEJ and Averts Excessive Nucleotide Alterations at Repair Joints.
    Aneesa T Al-Soodani, Xiling Wu, Nicole C Kelp, Alexander J Brown, Steven A Roberts, Chengtao Her.
      Inappropriate repair of DNA double-strand breaks (DSBs) leads to genomic instability, cell death, or malignant transformation. Cells minimize these detrimental effects by selectively activating suitable DSB repair pathways in accordance with their underlying cellular context. Here, we report that hMSH5 down-regulates NHEJ and restricts the extent of DSB end processing before rejoining, thereby reducing "excessive" deletions and insertions at repair joints. RNAi-mediated knockdown of hMSH5 led to large nucleotide deletions and longer insertions at the repair joints, while at the same time reducing the average length of microhomology (MH) at repair joints. Conversely, hMSH5 overexpression reduced end-joining activity and increased RPA foci formation (i.e., more stable ssDNA at DSB ends). Furthermore, silencing of hMSH5 delayed 53BP1 chromatin spreading, leading to increased end resection at DSB ends.
    Keywords:  53BP1; DSB repair; MSH5; NHEJ; end resection; genome instability
    DOI:  https://doi.org/10.3390/genes13040673
  10. Mol Cell. 2022 Apr 07. pii: S1097-2765(22)00268-4. [Epub ahead of print]
    H3K4 methylation by SETD1A/BOD1L facilitates RIF1-dependent NHEJ.
    Rachel Bayley, Valerie Borel, Rhiannon J Moss, Ellie Sweatman, Philip Ruis, Alice Ormrod, Amalia Goula, Rachel M A Mottram, Tyler Stanage, Graeme Hewitt, Marco Saponaro, Grant S Stewart, Simon J Boulton, Martin R Higgs.
      The 53BP1-RIF1-shieldin pathway maintains genome stability by suppressing nucleolytic degradation of DNA ends at double-strand breaks (DSBs). Although RIF1 interacts with damaged chromatin via phospho-53BP1 and facilitates recruitment of the shieldin complex to DSBs, it is unclear whether other regulatory cues contribute to this response. Here, we implicate methylation of histone H3 at lysine 4 by SETD1A-BOD1L in the recruitment of RIF1 to DSBs. Compromising SETD1A or BOD1L expression or deregulating H3K4 methylation allows uncontrolled resection of DNA ends, impairs end-joining of dysfunctional telomeres, and abrogates class switch recombination. Moreover, defects in RIF1 localization to DSBs are evident in patient cells bearing loss-of-function mutations in SETD1A. Loss of SETD1A-dependent RIF1 recruitment in BRCA1-deficient cells restores homologous recombination and leads to resistance to poly(ADP-ribose)polymerase inhibition, reinforcing the clinical relevance of these observations. Mechanistically, RIF1 binds directly to methylated H3K4, facilitating its recruitment to, or stabilization at, DSBs.
    Keywords:  53BP1; BOD1L; H3K4 methylation; PARP inhibitors; RIF1; SETD1A; class switch recombination; double-strand break repair; shieldin
    DOI:  https://doi.org/10.1016/j.molcel.2022.03.030
  11. J Biol Chem. 2022 Apr 13. pii: S0021-9258(22)00377-5. [Epub ahead of print] 101937
    Nej1 interacts with Sae2 at DNA double-stranded breaks to inhibit DNA resection.
    Aditya Mojumdar, Nancy Adam, Jennifer A Cobb.
      The two major pathways of DNA double strand break (DSB) repair, non-homologous end-joining (NHEJ) and homologous recombination (HR), are highly conserved from yeast to mammals. The regulation of 5'-DNA resection controls repair pathway choice and influences repair outcomes. Nej1 was first identified as a canonical NHEJ factor involved in stimulating the ligation of broken DNA ends, and more recently it was shown to participate in DNA end-bridging and in the inhibition of 5'-resection mediated by the nuclease/helicase complex Dna2-Sgs1. Here we show that Nej1 interacts with Sae2 to impact DSB repair in three ways. First, we show that Nej1 inhibits interaction of Sae2 with the Mre11/Rad50/Xrs2 (MRX) complex and Sae2 localization to DSBs. Second, we found that Nej1 inhibits Sae2-dependent recruitment of Dna2 independently of Sgs1. Third, we determined that NEJ1 and SAE2 showed an epistatic relationship for end-bridging, an event that restrains broken DNA ends and reduces the frequency of genomic deletions from developing at the break site. Finally, we demonstrate that deletion of NEJ1 suppressed the synthetic lethality of sae2Δ sgs1Δ mutants, and that triple mutant viability was dependent on Dna2 nuclease activity. Taken together, these findings provide mechanistic insight to how Nej1 functionality inhibits the initiation of DNA resection, a role that is distinct from its involvement in end-joining repair at DSBs.
    Keywords:  5’ resection; DNA damage; DSB repair; Dna2; Nej1; Repair pathway choice; Saccharomyces cerevisiae; Sae2; end-bridging; homologous recombination; molecular biology; protein‐protein interaction; yeast genetics
    DOI:  https://doi.org/10.1016/j.jbc.2022.101937
  12. Genes (Basel). 2022 Mar 31. pii: 625. [Epub ahead of print]13(4):
    Integrating Sister Chromatid Cohesion Establishment to DNA Replication.
    Caitlin M Zuilkoski, Robert V Skibbens.
      The intersection through which two fundamental processes meet provides a unique vantage point from which to view cellular regulation. On the one hand, DNA replication is at the heart of cell division, generating duplicate chromosomes that allow each daughter cell to inherit a complete copy of the parental genome. Among other factors, the PCNA (proliferating cell nuclear antigen) sliding clamp ensures processive DNA replication during S phase and is essential for cell viability. On the other hand, the process of chromosome segregation during M phase-an act that occurs long after DNA replication-is equally fundamental to a successful cell division. Eco1/Ctf7 ensures that chromosomes faithfully segregate during mitosis, but functions during DNA replication to activate cohesins and thereby establish cohesion between sister chromatids. To achieve this, Eco1 binds PCNA and numerous other DNA replication fork factors that include MCM helicase, Chl1 helicase, and the Rtt101-Mms1-Mms22 E3 ubiquitin ligase. Here, we review the multi-faceted coordination between cohesion establishment and DNA replication. SUMMARY STATEMENT: New findings provide important insights into the mechanisms through which DNA replication and the establishment of sister chromatid cohesion are coupled.
    Keywords:  DNA replication; Eco1/Ctf7/ESCO2; PCNA; RFC complexes; sister chromatid cohesion
    DOI:  https://doi.org/10.3390/genes13040625
  13. Mol Cell. 2022 Apr 12. pii: S1097-2765(22)00269-6. [Epub ahead of print]
    CDYL1-dependent decrease in lysine crotonylation at DNA double-strand break sites functionally uncouples transcriptional silencing and repair.
    Enas R Abu-Zhayia, Laila A Bishara, Feras E Machour, Alma Sophia Barisaac, Bella M Ben-Oz, Nabieh Ayoub.
      Previously, we showed that CDYL1 is recruited to DNA double-strand breaks (DSBs) to promote homologous recombination (HR) repair and foster transcriptional silencing. However, how CDYL1 elicits DSB-induced silencing is not fully understood. Here, we identify a CDYL1-dependent local decrease in the transcriptionally active marks histone lysine crotonylation (Kcr) and crotonylated lysine 9 of H3 (H3K9cr) at AsiSI-induced DSBs, which correlates with transcriptional silencing. Mechanistically, we reveal that CDYL1 crotonyl-CoA hydratase activity counteracts Kcr and H3K9cr at DSB sites, which triggers the eviction of the transcription elongation factor ENL and fosters transcriptional silencing. Furthermore, genetic inhibition of CDYL1 hydratase activity blocks the reduction in H3K9cr and alleviates DSB-induced silencing, whereas HR efficiency unexpectedly remains intact. Therefore, our results functionally uncouple the repair and silencing activity of CDYL1 at DSBs. In a broader context, we address a long-standing question concerning the functional relationship between HR repair and DSB-induced silencing, suggesting that they may occur independently.
    Keywords:  CDYL1; ENL; H3K9cr; NHEJ; double-strand break; homologous recombination; lysine acetylation; lysine crotonylation; transcriptional repression
    DOI:  https://doi.org/10.1016/j.molcel.2022.03.031
  14. Cell Death Differ. 2022 Apr 20.
    p53 at the crossroad of DNA replication and ribosome biogenesis stress pathways.
    Mikael S Lindström, Jiri Bartek, Apolinar Maya-Mendoza.
      Despite several decades of intense research focused on understanding function(s) and disease-associated malfunction of p53, there is no sign of any "mid-life crisis" in this rapidly advancing area of biomedicine. Firmly established as the hub of cellular stress responses and tumor suppressor targeted in most malignancies, p53's many talents continue to surprise us, providing not only fresh insights into cell and organismal biology, but also new avenues to cancer treatment. Among the most fruitful lines of p53 research in recent years have been the discoveries revealing the multifaceted roles of p53-centered pathways in the fundamental processes of DNA replication and ribosome biogenesis (RiBi), along with cellular responses to replication and RiBi stresses, two intertwined areas of cell (patho)physiology that we discuss in this review. Here, we first provide concise introductory notes on the canonical roles of p53, the key interacting proteins, downstream targets and post-translational modifications involved in p53 regulation. We then highlight the emerging involvement of p53 as a key component of the DNA replication Fork Speed Regulatory Network and the mechanistic links of p53 with cellular checkpoint responses to replication stress (RS), the driving force of cancer-associated genomic instability. Next, the tantalizing, yet still rather foggy functional crosstalk between replication and RiBi (nucleolar) stresses is considered, followed by the more defined involvement of p53-mediated monitoring of the multistep process of RiBi, including the latest updates on the RPL5/RPL11/5 S rRNA-MDM2-p53-mediated Impaired Ribosome Biogenesis Checkpoint (IRBC) pathway and its involvement in tumorigenesis. The diverse defects of RiBi and IRBC that predispose and/or contribute to severe human pathologies including developmental syndromes and cancer are then outlined, along with examples of promising small-molecule-based strategies to therapeutically target the RS- and particularly RiBi- stress-tolerance mechanisms to which cancer cells are addicted due to their aberrant DNA replication, repair, and proteo-synthesis demands.
    DOI:  https://doi.org/10.1038/s41418-022-00999-w
  15. Eur J Pharmacol. 2022 Apr 14. pii: S0014-2999(22)00218-7. [Epub ahead of print]924 174957
    DNA damage response and repair in the development and treatment of brain tumors.
    Parisa Maleki Dana, Fatemeh Sadoughi, Hamed Mirzaei, Zatollah Asemi, Bahman Yousefi.
      DNA damage response (DDR) comprising DNA repair and cell-cycle checkpoint pathways, is considered as a protective process that maintains the integrity of the genome. However, this mechanism may not be favorable in the context of cancer. Indeed, studies have shown that DDR and repair mechanisms can be involved in the development of different cancers. Furthermore, they may lead to the failure of therapeutic approaches. Thus, studying these mechanisms can be beneficial in a better understanding of cancer development and developing more efficient treatments. Scopus, Google Scholar, and PubMed databases were used for searching articles published on "DNA damage response and DNA repair in the development and treatment of brain tumors". Herein, we review the literature on DNA damage response and DNA repair mechanisms in the development of brain tumors, such as glioma, glioblastoma, and medulloblastoma. Moreover, we summarize the studies that conducted on the role of targeting components of DNA damage response and DNA repair in treating different types of brain cancers, enhancing the currently available therapeutic approaches, and solving the problems in the field of brain cancer therapy.
    Keywords:  DNA damage response; Drug resistance; Glioblastoma; Glioma; Medulloblastoma
    DOI:  https://doi.org/10.1016/j.ejphar.2022.174957
  16. Biomedicines. 2022 Mar 23. pii: 751. [Epub ahead of print]10(4):
    Purine Synthesis Inhibitor L-Alanosine Impairs Mitochondrial Function and Stemness of Brain Tumor Initiating Cells.
    Simranjit X Singh, Rui Yang, Kristen Roso, Landon J Hansen, Changzheng Du, Lee H Chen, Paula K Greer, Christopher J Pirozzi, Yiping He.
      Glioblastoma (GBM) is a lethal brain cancer exhibiting high levels of drug resistance, a feature partially imparted by tumor cell stemness. Recent work shows that homozygous MTAP deletion, a genetic alteration occurring in about half of all GBMs, promotes stemness in GBM cells. Exploiting MTAP loss-conferred deficiency in purine salvage, we demonstrate that purine blockade via treatment with L-Alanosine (ALA), an inhibitor of de novo purine synthesis, attenuates stemness of MTAP-deficient GBM cells. This ALA-induced reduction in stemness is mediated in part by compromised mitochondrial function, highlighted by ALA-induced elimination of mitochondrial spare respiratory capacity. Notably, these effects of ALA are apparent even when the treatment was transient and with a low dose. Finally, in agreement with diminished stemness and compromised mitochondrial function, we show that ALA sensitizes GBM cells to temozolomide (TMZ) in vitro and in an orthotopic GBM model. Collectively, these results identify purine supply as an essential component in maintaining mitochondrial function in GBM cells and highlight a critical role of mitochondrial function in sustaining GBM stemness. We propose that purine synthesis inhibition can be beneficial in combination with the standard of care for MTAP-deficient GBMs, and that it may be feasible to achieve this benefit without inflicting major toxicity.
    Keywords:  MTAP; adenine; alanosine; drug resistance; glioma stem cells; mitochondria; purine
    DOI:  https://doi.org/10.3390/biomedicines10040751
  17. iScience. 2022 Apr 15. 25(4): 104142
    Heat-induced SIRT1-mediated H4K16ac deacetylation impairs resection and SMARCAD1 recruitment to double strand breaks.
    Sharmistha Chakraborty, Mayank Singh, Raj K Pandita, Vipin Singh, Calvin S C Lo, Fransisca Leonard, Nobuo Horikoshi, Eduardo G Moros, Deblina Guha, Clayton R Hunt, Eric Chau, Kazi M Ahmed, Prayas Sethi, Vijaya Charaka, Biana Godin, Kalpana Makhijani, Harry Scherthan, Jeanette Deck, Michael Hausmann, Arjamand Mushtaq, Mohammad Altaf, Kenneth S Ramos, Krishna M Bhat, Nitika Taneja, Chandrima Das, Tej K Pandita.
      Hyperthermia inhibits DNA double-strand break (DSB) repair that utilizes homologous recombination (HR) pathway by a poorly defined mechanism(s); however, the mechanisms for this inhibition remain unclear. Here we report that hyperthermia decreases H4K16 acetylation (H4K16ac), an epigenetic modification essential for genome stability and transcription. Heat-induced reduction in H4K16ac was detected in humans, Drosophila, and yeast, indicating that this is a highly conserved response. The examination of histone deacetylase recruitment to chromatin after heat-shock identified SIRT1 as the major deacetylase subsequently enriched at gene-rich regions. Heat-induced SIRT1 recruitment was antagonized by chromatin remodeler SMARCAD1 depletion and, like hyperthermia, the depletion of the SMARCAD1 or combination of the two impaired DNA end resection and increased replication stress. Altered repair protein recruitment was associated with heat-shock-induced γ-H2AX chromatin changes and DSB repair processing. These results support a novel mechanism whereby hyperthermia impacts chromatin organization owing to H4K16ac deacetylation, negatively affecting the HR-dependent DSB repair.
    Keywords:  Biological sciences; Epigenetics; Molecular biology
    DOI:  https://doi.org/10.1016/j.isci.2022.104142
  18. Int J Mol Sci. 2022 Apr 12. pii: 4264. [Epub ahead of print]23(8):
    Development and Evolution of DNA-Dependent Protein Kinase Inhibitors toward Cancer Therapy.
    Yoshihisa Matsumoto.
      DNA double-strand break (DSB) is considered the most deleterious type of DNA damage, which is generated by ionizing radiation (IR) and a subset of anticancer drugs. DNA-dependent protein kinase (DNA-PK), which is composed of a DNA-PK catalytic subunit (DNA-PKcs) and Ku80-Ku70 heterodimer, acts as the molecular sensor for DSB and plays a pivotal role in DSB repair through non-homologous end joining (NHEJ). Cells deficient for DNA-PKcs show hypersensitivity to IR and several DNA-damaging agents. Cellular sensitivity to IR and DNA-damaging agents can be augmented by the inhibition of DNA-PK. A number of small molecules that inhibit DNA-PK have been developed. Here, the development and evolution of inhibitors targeting DNA-PK for cancer therapy is reviewed. Significant parts of the inhibitors were developed based on the structural similarity of DNA-PK to phosphatidylinositol 3-kinases (PI3Ks) and PI3K-related kinases (PIKKs), including Ataxia-telangiectasia mutated (ATM). Some of DNA-PK inhibitors, e.g., NU7026 and NU7441, have been used extensively in the studies for cellular function of DNA-PK. Recently developed inhibitors, e.g., M3814 and AZD7648, are in clinical trials and on the way to be utilized in cancer therapy in combination with radiotherapy and chemotherapy.
    Keywords:  DNA double-strand break (DSB); DNA-dependent protein kinase (DNA-PK); chemosensitization; inhibitor; non-homologous end joining (NHEJ); phosphatidylinositol 3-kinase; radiosensitization
    DOI:  https://doi.org/10.3390/ijms23084264
  19. J Biol Chem. 2022 Apr 18. pii: S0021-9258(22)00387-8. [Epub ahead of print] 101947
    Stable G-quadruplex DNA structures promote replication-dependent genome instability.
    S Dean Rider, Rujuta Yashodhan Gadgil, David C Hitch, French J Damewood, Nathen Zavada, Matilyn Shanahan, Venicia Alhawach, Resha Shrestha, Kazuo Shin-Ya, Michael Leffak.
      G quadruplex-prone structures are abundant in mammalian genomes, where they have been shown to influence DNA replication, transcription, and genome stability. In this study we constructed cells with a single ectopic homopurine-homopyrimidine (Pu/Py) repeat tract derived from the PKD1 locus, which is capable of forming triplex (H3) and G-quadruplex (G4) DNA structures. We show that ligand stabilization of these G4 structures results in deletions of the G4 consensus sequence, as well as kilobase deletions spanning the quadruplex and ectopic sites. Furthermore, we show DNA double strand breaks (DSBs) at the ectopic site are dependent on the nuclease Mus81. Hypermutagenesis during chromatid repair extends several kilobases from the G4 site, and breaks at the G4 site result in microhomology-mediated translocations. To determine whether triplex or quadruplex structures are responsible for Pu/Py tract instability, we derived constructs and cell lines from the PKD1 repeat which can only form H3 or G4 structures. Under normal growth conditions, we found G4 cell lines lost the G4 consensus sequence early during clonal outgrowth, while H3 cells showed DNA instability early during outgrowth but only lost reporter gene expression after prolonged growth. Thus, both the H3 and G4 non-B conformation DNAs exhibit genomic instability, but they respond differently to endogenous replication stress. Our results show the outcomes of replication-dependent DSBs at non-B DNAs model the instability observed in microhomology-mediated break-induced replication (MMBIR). Marked variability in the frequency of mutagenesis during BIR suggests possible dynamic heterogeneity in the BIR replisome.
    DOI:  https://doi.org/10.1016/j.jbc.2022.101947
  20. Biochemistry. 2022 Apr 18.
    Kinetic Analysis of the Effect of N-Terminal Acetylation on Thymine DNA Glycosylase.
    Mary E Tarantino, Sarah Delaney.
      Thymine DNA glycosylase (TDG) is tasked with initiating DNA base excision repair by recognizing and removing T, U, the chemotherapeutic 5-fluorouracil (5-FU), and many other oxidized and halogenated pyrimidine bases. TDG contains a long, unstructured N-terminus that contains four known sites of acetylation: lysine (K) residues 59, 83, 84, and 87. Here, K to glutamine (Q) mutants are used as acetyl-lysine (AcK) analogues to probe the effect of N-terminal acetylation on the kinetics of TDG. We find that mimicking acetylation affects neither the maximal single-turnover rate kmax nor the turnover rate kTO, indicating that the steps after initial binding, through chemistry and product release, are not affected. Under subsaturating conditions, however, acetylation changes the processing of U substrates. Subtle differences among AcK analogues are revealed with 5-FU in single-stranded DNA. We propose that the subtleties observed among the AcK analogues may be amplified on the genomic scale, leading to regulation of TDG activity. N-terminal acetylation, though, may also play a structural, rather than kinetic role in vivo.
    DOI:  https://doi.org/10.1021/acs.biochem.1c00823
  21. Mol Cell. 2022 Apr 13. pii: S1097-2765(22)00289-1. [Epub ahead of print]
    Heterochromatic repeat clustering imposes a physical barrier on homologous recombination to prevent chromosomal translocations.
    Ioanna Mitrentsi, Jieqiong Lou, Adèle Kerjouan, John Verigos, Bernardo Reina-San-Martin, Elizabeth Hinde, Evi Soutoglou.
      Mouse pericentromeric DNA is composed of tandem major satellite repeats, which are heterochromatinized and cluster together to form chromocenters. These clusters are refractory to DNA repair through homologous recombination (HR). The mechanisms by which pericentromeric heterochromatin imposes a barrier on HR and the implications of repeat clustering are unknown. Here, we compare the spatial recruitment of HR factors upon double-stranded DNA breaks (DSBs) induced in human and mouse pericentromeric heterochromatin, which differ in their capacity to form clusters. We show that while DSBs increase the accessibility of human pericentromeric heterochromatin by disrupting HP1α dimerization, mouse pericentromeric heterochromatin repeat clustering imposes a physical barrier that requires many layers of de-compaction to be accessed. Our results support a model in which the 3D organization of heterochromatin dictates the spatial activation of DNA repair pathways and is key to preventing the activation of HR within clustered repeats and the onset of chromosomal translocations.
    Keywords:  DNA repair; HP1α; HR; Rad51; pericentromeric heterochromatin
    DOI:  https://doi.org/10.1016/j.molcel.2022.03.033
  22. Int J Mol Sci. 2022 Apr 08. pii: 4149. [Epub ahead of print]23(8):
    Replicative Stress Coincides with Impaired Nuclear DNA Damage Response in COX4-1 Deficiency.
    Liza Douiev, Chaya Miller, Guy Keller, Hadar Benyamini, Bassam Abu-Libdeh, Ann Saada.
      Cytochrome c oxidase (COX), a multimeric protein complex, is the final electron acceptor in the mitochondrial electron transfer chain. Primary COX deficiency, caused by mutations in either mitochondrial DNA or nuclear-encoded genes, is a heterogenous group of mitochondrial diseases with a wide range of presentations, ranging from fatal infantile to subtler. We previously reported a patient with primary COX deficiency due to a pathogenic variant in COX4I1 (encoding the common isoform of COX subunit 4, COX4-1), who presented with bone marrow failure, genomic instability, and short stature, mimicking Fanconi anemia (FA). In the present study, we demonstrated that accumulative DNA damage coincided primarily with proliferative cells in the patient's fibroblasts and in COX4i1 knockdown cells. Expression analysis implicated a reduction in DNA damage response pathways, which was verified by demonstrating impaired recovery from genotoxic insult and decreased DNA repair. The premature senescence of the COX4-1-deficient cells prevented us from undertaking additional studies; nevertheless, taken together, our results indicate replicative stress and impaired nuclear DNA damage response in COX4-1 deficiency. Interestingly, our in vitro findings recapitulated the patient's presentation and present status.
    Keywords:  COX4i1; DNA damage; cytochrome c oxidase; mitochondria; mitochondrial respiratory chain; replicative stress
    DOI:  https://doi.org/10.3390/ijms23084149
  23. Int J Mol Sci. 2022 Apr 13. pii: 4314. [Epub ahead of print]23(8):
    Glucose Increases STAT3 Activation, Promoting Sustained XRCC1 Expression and Increasing DNA Repair.
    Griffin M Wright, Natalie R Gassman.
      Dysregulation of DNA repair is a hallmark of cancer, though few cancer-specific mechanisms that drive the overexpression of DNA repair proteins are known. We previously identified STAT3 as a novel transcriptional regulator of X-ray cross-complementing group 1 (XRCC1), an essential scaffold protein in base excision repair in triple-negative breast cancers. We also identified an inducible response to IL-6 and epidermal growth factor stimulation in the non-tumorigenic embryonic kidney cell line HEK293T. As IL-6 and EGF signaling are growth and inflammatory-inducible responses, we examined if glucose challenge can increase STAT3 activation, promoting adaptive changes in XRCC1 expression in different cell types. Acute high glucose exposure promoted XRCC1 expression through STAT3 activation, increasing the repair of methyl methanesulfonate-induced DNA damage in HEK293T cells and the osteosarcoma cell line U2OS. Sustained exposure to high glucose promoted the overexpression of XRCC1, which can be reversed upon glucose restriction and down-regulation of STAT3 activation. Thus, we have identified a novel link between XRCC1 expression and STAT3 activation following exogenous exposures, which could play a critical role in dictating a cancer cell's response to DNA-damaging agents.
    Keywords:  DNA damage; DNA repair; STAT3; XRCC1; glucose; stress
    DOI:  https://doi.org/10.3390/ijms23084314
  24. J Cell Biochem. 2022 Apr 17.
    VGLL3 increases the dependency of cancer cells on de novo nucleotide synthesis through GART expression.
    Tomohiro Kawamura, Yuki Takehora, Naoto Hori, Yuki Takakura, Naoto Yamaguchi, Hiroyuki Takano, Noritaka Yamaguchi.
      Vestigial-like family member 3 (VGLL3) is a member of the VGLL family that serves as cofactors for TEA-domain transcription factors. Although VGLL3 is involved in the proliferation of cancer cells, the molecular mechanisms underlying VGLL3-mediated cell proliferation remain largely unknown. In this study, we found that stable expression of VGLL3 in human lung cancer A549 cells affects glutamine metabolism and increases their dependency on de novo nucleotide synthesis for proliferation. Mechanistically, VGLL3 was found to induce the expression of GART, which encodes a trifunctional enzyme that catalyzes de novo purine synthesis from glutamine. GART knockdown and the glycinamide ribonucleotide synthase, aminoimidazole ribonucleotide synthase, and glycinamide ribonucleotide formyltransferase trifunctional protein (GART) inhibitor lometrexol repressed the proliferation and survival of A549 cells stably expressing VGLL3. Mesenchymal breast cancer BT549 cells and MDA-MB-231 cells showed high expression of VGLL3, and VGLL3 knockdown was found to reduce GART expression. Lometrexol also repressed the proliferation of these breast cancer cells, whereas addition of inosine monophosphate, an important metabolite downstream of GART, rescued this repression. Taken together, these results suggest that VGLL3 induces GART expression and thereby confers de novo nucleotide-dependent cell proliferation in cancer cells.
    Keywords:  GART protein; cell proliferation; glutamine; lometrexol; nucleotides; phosphoribosylglycinamide formyltransferase; transcription factors
    DOI:  https://doi.org/10.1002/jcb.30251
  25. Nature. 2022 Apr 20.
    CCNE1 amplification is synthetic lethal with PKMYT1 kinase inhibition.
    David Gallo, Jordan T F Young, Jimmy Fourtounis, Giovanni Martino, Alejandro Álvarez-Quilón, Cynthia Bernier, Nicole M Duffy, Robert Papp, Anne Roulston, Rino Stocco, Janek Szychowski, Artur Veloso, Hunain Alam, Prasamit S Baruah, Alexanne Bonneau Fortin, Julian Bowlan, Natasha Chaudhary, Jessica Desjardins, Evelyne Dietrich, Sara Fournier, Chloe Fugère-Desjardins, Theo Goullet de Rugy, Marie-Eve Leclaire, Bingcan Liu, Vivek Bhaskaran, Yael Mamane, Henrique Melo, Olivier Nicolas, Akul Singhania, Rachel K Szilard, Ján Tkáč, Shou Yun Yin, Stephen J Morris, Michael Zinda, C Gary Marshall, Daniel Durocher.
      Amplification of the CCNE1 locus on chromosome 19q12 is prevalent in multiple tumour types, particularly in high-grade serous ovarian cancer, uterine tumours and gastro-oesophageal cancers, where high cyclin E levels are associated with genome instability, whole-genome doubling and resistance to cytotoxic and targeted therapies1-4. To uncover therapeutic targets for tumours with CCNE1 amplification, we undertook genome-scale CRISPR-Cas9-based synthetic lethality screens in cellular models of CCNE1 amplification. Here we report that increasing CCNE1 dosage engenders a vulnerability to the inhibition of the PKMYT1 kinase, a negative regulator of CDK1. To inhibit PKMYT1, we developed RP-6306, an orally bioavailable and selective inhibitor that shows single-agent activity and durable tumour regressions when combined with gemcitabine in models of CCNE1 amplification. RP-6306 treatment causes unscheduled activation of CDK1 selectively in CCNE1-overexpressing cells, promoting early mitosis in cells undergoing DNA synthesis. CCNE1 overexpression disrupts CDK1 homeostasis at least in part through an early activation of the MMB-FOXM1 mitotic transcriptional program. We conclude that PKMYT1 inhibition is a promising therapeutic strategy for CCNE1-amplified cancers.
    DOI:  https://doi.org/10.1038/s41586-022-04638-9
  26. Cell Death Dis. 2022 Apr 20. 13(4): 379
    Artesunate improves venetoclax plus cytarabine AML cell targeting by regulating the Noxa/Bim/Mcl-1/p-Chk1 axis.
    Jingyi Zhang, Yuetong Wang, Chujie Yin, Ping Gong, Zhenwei Zhang, Linxiang Zhao, Samuel Waxman, Yongkui Jing.
      Venetoclax plus cytarabine therapy is approved for elderly acute myeloid leukemia (AML) patients and needs further improvement. We studied the mechanisms of venetoclax plus cytarabine treatment and searched for a third agent to enhance their effects. Cytarabine induces S phase arrest-mediated DNA damage with activation of DNA replication checkpoint kinase 1 (Chk1) through phosphorylation, while venetoclax induces B cell lymphoma 2 (Bcl-2)-interacting mediator of cell death (Bim)-mediated apoptotic DNA damage. Myeloid cell leukemia-1 (Mcl-1) plays negative roles in both events by sequestering Bim and accelerating Chk1 phosphorylation. Venetoclax releases Bim from Bcl-2 with increased Bim binding to Mcl-1. Artesunate, an antimalaria drug, induces Noxa to replace Bim from Mcl-1 and induces synergistic apoptosis with venetoclax accompanied with Mcl-1 reduction. Silencing Mcl-1 or adding venetoclax/artesunate diminishes the cytarabine resistance pathway p-Chk1. The triple combination exhibits S phase arrest with enhanced DNA damage, improves AML colony formation inhibition, and prolongs survival of two mice xenograft models compared to the venetoclax/cytarabine dual combination. Artesunate serves as a bridge for venetoclax and cytarabine combination by Noxa and Bim-mediated apoptosis and Mcl-1 reduction. We provide a new triple combination for AML treatment by targeting the Noxa/Mcl-1/Bim axis to reverse Mcl-1/p-Chk1 resistance of cytarabine therapy.
    DOI:  https://doi.org/10.1038/s41419-022-04810-z
  27. Mol Cancer Ther. 2022 Apr 19. pii: molcanther.1000.2021. [Epub ahead of print]
    TOP1-DNA trapping by exatecan and combination therapy with ATR inhibitor.
    Ukhyun Jo, Yasuhisa Murai, Keli K Agama, Yilun Sun, Liton Kumar Saha, Xi Yang, Yasuhiro Arakawa, Sophia Gayle, Kelli Jones, Vishwas Paralkar, Ranjini K Sundaram, Jinny Van Doorn, Juan C Vasquez, Ranjit S Bindra, Woo Suk Choi, Yves Pommier.
      Exatecan and deruxtecan are antineoplastic camptothecin derivatives in development as tumor-targeted-delivery warheads in various formulations including peptides, liposomes, polyethylene glycol (PEG) nanoparticles, and antibody-drug conjugates (ADCs). Here, we report the molecular pharmacology of exatecan compared to the clinically approved topoisomerase I (TOP1) inhibitors and preclinical models for validating biomarkers and the combination of exatecan with ATR inhibitors. Modeling exatecan binding at the interface of a TOP1 cleavage complex suggests two novel molecular interactions with the flanking DNA base and the TOP1 residue N352, in addition to the three known interactions of camptothecins with the TOP1 residues R364, D533 and N722. Accordingly, exatecan showed much stronger TOP1 trapping, higher DNA damage and apoptotic cell death than the classical TOP1 inhibitors used clinically. We demonstrate the value of SLFN11 expression and homologous recombination (HR)-deficiency (HRD) as predictive biomarkers of response to exatecan. We also show that exatecan kills cancer cells synergistically with the clinical ATR inhibitor ceralasertib (AZD6738). To establish the translational potential of this combination, we tested CBX-12, a clinically developed pH-sensitive peptide-exatecan conjugate that selectively targets cancer cells and is currently in clinical trials. The combination of CBX-12 with ceralasertib significantly suppressed tumor growth in mouse xenografts. Collectively, our results demonstrate the potency of exatecan as a TOP1 inhibitor and its clinical potential in combination with ATR inhibitors, using SLFN11 and HRD as predictive biomarkers.
    DOI:  https://doi.org/10.1158/1535-7163.MCT-21-1000
  28. Sci Rep. 2022 Apr 18. 12(1): 6408
    Inosine triphosphate pyrophosphatase from Trypanosoma brucei cleanses cytosolic pools from deaminated nucleotides.
    Antonio E Vidal, Miriam Yagüe-Capilla, Blanca Martínez-Arribas, Daniel García-Caballero, Luis M Ruiz-Pérez, Dolores González-Pacanowska.
      Inosine triphosphate pyrophosphatases (ITPases) are ubiquitous house-cleaning enzymes that specifically recognize deaminated purine nucleotides and catalyze their hydrolytic cleavage. In this work, we have characterized the Trypanosoma brucei ITPase ortholog (TbITPA). Recombinant TbITPA efficiently hydrolyzes (deoxy)ITP and XTP nucleotides into their respective monophosphate form. Immunolocalization analysis performed in bloodstream forms suggests that the primary role of TbITPA is the exclusion of deaminated purines from the cytosolic nucleoside triphosphate pools. Even though ITPA-knockout bloodstream parasites are viable, they are more sensitive to inhibition of IMP dehydrogenase with mycophenolic acid, likely due to an expansion of IMP, the ITP precursor. On the other hand, TbITPA can also hydrolyze the activated form of the antiviral ribavirin although in this case, the absence of ITPase activity in the cell confers protection against this nucleoside analog. This unexpected phenotype is dependant on purine availability and can be explained by the fact that ribavirin monophosphate, the reaction product generated by TbITPA, is a potent inhibitor of trypanosomal IMP dehydrogenase and GMP reductase. In summary, the present study constitutes the first report on a protozoan inosine triphosphate pyrophosphatase involved in the removal of harmful deaminated nucleotides from the cytosolic pool.
    DOI:  https://doi.org/10.1038/s41598-022-10149-4
  29. Nucleic Acids Res. 2022 Apr 19. pii: gkac241. [Epub ahead of print]
    Mismatch discrimination and sequence bias during end-joining by DNA ligases.
    Katharina Bilotti, Vladimir Potapov, John M Pryor, Alexander T Duckworth, James L Keck, Gregory J S Lohman.
      DNA ligases, critical enzymes for in vivo genome maintenance and modern molecular biology, catalyze the joining of adjacent 3'-OH and 5'-phosphorylated ends in DNA. To determine whether DNA annealing equilibria or properties intrinsic to the DNA ligase enzyme impact end-joining ligation outcomes, we used a highly multiplexed, sequencing-based assay to profile mismatch discrimination and sequence bias for several ligases capable of efficient end-joining. Our data reveal a spectrum of fidelity and bias, influenced by both the strength of overhang annealing as well as sequence preferences and mismatch tolerances that vary both in degree and kind between ligases. For example, while T7 DNA ligase shows a strong preference for ligating high GC sequences, other ligases show little GC-dependent bias, with human DNA Ligase 3 showing almost none. Similarly, mismatch tolerance varies widely among ligases, and while all ligases tested were most permissive of G:T mismatches, some ligases also tolerated bulkier purine:purine mismatches. These comprehensive fidelity and bias profiles provide insight into the biology of end-joining reactions and highlight the importance of ligase choice in application design.
    DOI:  https://doi.org/10.1093/nar/gkac241
  30. Ann Hematol. 2022 Apr 18.
    Purine nucleoside analogs plus rituximab are an effective treatment choice for hairy cell leukemia-variant.
    Yi Wang, Tingyu Wang, Ying Yu, Qi Wang, Yuting Yan, Ru Li, Qi Sun, Wenjie Xiong, Rui Lyu, Zhen Yu, Wei Liu, Weiwei Sui, Wenyang Huang, Huijun Wang, Chengwen Li, Jun Wang, Dehui Zou, Gang An, Jianxiang Wang, Lugui Qiu, Shuhua Yi.
      Both characteristics and optimal treatment strategy for hairy cell leukemia-variant (HCL-v) remain elusive due to its rarity. We retrospectively analyzed the clinical features of HCL-v and the efficacy of first-line treatment options in a large Chinese cohort. In this study, we recruited 33 HCL-v patients (23 males and 10 females) with a median age of 59 years (range, 34-79 years). The chief complaints included abdominal mass and relative signs (67%) and abnormal complete blood count (27%). Immunophenotyping showed monoclonal B-cells positive for pan B-cell antigens and CD11c, weakly positive for CD103 and CD200, while negative for CD5, CD10, CD25, CD123, and annexin A1. No BRAF V600E mutation was detected, but TP53 abnormality was recurrent. Treatment choices included interferon-α (IFN-α) in 11 patients, chlorambucil (CLB) in 5 patients, single purine nucleoside analogs (PNA) in 3 patients, PNA plus rituximab (PNA + R) in 9 patients, and others in 3 patients. Four patients who received IFN-α or CLB treatment also underwent splenectomy. Patients who received PNA + R had a higher complete response rate (88% versus 5%, P < 0.001) and longer progression-free survival (PFS, 3-year PFS rate 42% [95% CI 1-84] vs. 16% [95% CI 3-40], P = 0.042) than those who received other regimens. Overall, HCL-v is an indolent lymphoma with unique characteristics. The PNA + R regimen is the preferred choice in the first-line treatment for HCL-v.
    Keywords:  Characteristics; Clinical outcome; First-line treatment; Hairy cell leukemia-variant
    DOI:  https://doi.org/10.1007/s00277-022-04795-x
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