bims-numges 2022-03-06 papers

bims-numges

Biomed News

on Nucleotide metabolism and genome stability

Issue of 2022–03–06
thirty papers selected by
Sean Rudd, Karolinska Institutet

Pharmacological targeting of MTHFD2 suppresses acute myeloid leukemia by inducing thymidine depletion and replication stress.
CTP sensing and Mec1ATR-Rad53CHK1/CHK2 mediate a two-layered response to inhibition of glutamine metabolism.
Intra-S phase checkpoint kinase Chk1 dissociates replication proteins Treslin and TopBP1 through multiple mechanisms during replication stress.
Fanconi anemia proteins and genome fragility: unraveling replication defects for cancer therapy.
Chk1 dynamics in G2 phase upon replication stress predict daughter cell outcome.
Histone variants: The unsung guardians of the genome.
LAP2α preserves genome integrity through assisting RPA deposition on damaged chromatin.
Rapid recruitment of p53 to DNA damage sites directs DNA repair choice and integrity.
XPG: a multitasking genome caretaker.
Emvododstat, a Potent Dihydroorotate Dehydrogenase Inhibitor, Is Effective in Preclinical Models of Acute Myeloid Leukemia.
Interlocking activities of DNA polymerase β in the base excision repair pathway.
Micropeptide PACMP inhibition elicits synthetic lethal effects by decreasing CtIP and poly(ADP-ribosyl)ation.
CDC7 kinase (DDK) inhibition disrupts DNA replication leading to mitotic catastrophe in Ewing sarcoma.
Double-strand breaks: When DNA repair events accidentally meet.
Sex disparities in DNA damage response pathways: Novel determinants in cancer formation and therapy.
Single-molecule mapping of replisome progression.
Genomic Reporter Constructs to Monitor Pathway-Specific Repair of DNA Double-Strand Breaks.
Effects of N7-Alkylguanine Conformation and Metal Cofactors on the Translesion Synthesis by Human DNA Polymerase η.
Defective repair of topoisomerase I induced chromosomal damage in Huntington's disease.
Human topoisomerases and their roles in genome stability and organization.
The influence of cdG on 8-oxodG excision by OGG1 and FPG glycosylases.
NAMPT: a critical driver and therapeutic target for cancer.
Are we still on the right Path(way)?: the altered expression of the pentose phosphate pathway in solid tumors and the potential of its inhibition in combination therapy.
8-oxodG accumulation within super-enhancers marks fragile CTCF-mediated chromatin loops.
Identification of Nanog as a novel inhibitor of Rad51.
Different mechanisms of drug resistance to hypomethylating agents in the treatment of myelodysplastic syndromes and acute myeloid leukemia.
The role of p53 and p21 on 8-chloro-adenosine-induced cellular response.
Trapping of 5-Fluorodeoxyuridine Monophosphate by Thymidylate Synthase Confers Resistance to 5-Fluorouracil.
SERAC1 is a component of the mitochondrial serine transporter complex required for the maintenance of mitochondrial DNA.
Releasing the trap: How the segregase p97 extracts PARP1 from chromatin.

Nat Cancer. 2022 Feb;3(2): 156-172

Pharmacological targeting of MTHFD2 suppresses acute myeloid leukemia by inducing thymidine depletion and replication stress.

Nadilly Bonagas, Nina M S Gustafsson, Martin Henriksson, Petra Marttila, Robert Gustafsson, Elisée Wiita, Sanjay Borhade, Alanna C Green, Karl S A Vallin, Antonio Sarno, Richard Svensson, Camilla Göktürk, Therese Pham, Ann-Sofie Jemth, Olga Loseva, Victoria Cookson, Nicole Kiweler, Lars Sandberg, Azita Rasti, Judith E Unterlass, Martin Haraldsson, Yasmin Andersson, Emma R Scaletti, Christoffer Bengtsson, Cynthia B J Paulin, Kumar Sanjiv, Eldar Abdurakhmanov, Linda Pudelko, Ben Kunz, Matthieu Desroses, Petar Iliev, Katarina Färnegårdh, Andreas Krämer, Neeraj Garg, Maurice Michel, Sara Häggblad, Malin Jarvius, Christina Kalderén, Amanda Bögedahl Jensen, Ingrid Almlöf, Stella Karsten, Si Min Zhang, Maria Häggblad, Anders Eriksson, Jianping Liu, Björn Glinghammar, Natalia Nekhotiaeva, Fredrik Klingegård, Tobias Koolmeister, Ulf Martens, Sabin Llona-Minguez, Ruth Moulson, Helena Nordström, Vendela Parrow, Leif Dahllund, Birger Sjöberg, Irene L Vargas, Duy Duc Vo, Johan Wannberg, Stefan Knapp, Hans E Krokan, Per I Arvidsson, Martin Scobie, Johannes Meiser, Pål Stenmark, Ulrika Warpman Berglund, Evert J Homan, Thomas Helleday.

The folate metabolism enzyme MTHFD2 (methylenetetrahydrofolate dehydrogenase/cyclohydrolase) is consistently overexpressed in cancer but its roles are not fully characterized, and current candidate inhibitors have limited potency for clinical development. In the present study, we demonstrate a role for MTHFD2 in DNA replication and genomic stability in cancer cells, and perform a drug screen to identify potent and selective nanomolar MTHFD2 inhibitors; protein cocrystal structures demonstrated binding to the active site of MTHFD2 and target engagement. MTHFD2 inhibitors reduced replication fork speed and induced replication stress followed by S-phase arrest and apoptosis of acute myeloid leukemia cells in vitro and in vivo, with a therapeutic window spanning four orders of magnitude compared with nontumorigenic cells. Mechanistically, MTHFD2 inhibitors prevented thymidine production leading to misincorporation of uracil into DNA and replication stress. Overall, these results demonstrate a functional link between MTHFD2-dependent cancer metabolism and replication stress that can be exploited therapeutically with this new class of inhibitors.

DOI: https://doi.org/10.1038/s43018-022-00331-y
PLoS Genet. 2022 Mar 03. 18(3): e1010101

CTP sensing and Mec1ATR-Rad53CHK1/CHK2 mediate a two-layered response to inhibition of glutamine metabolism.

Arta Ajazi, Ramveer Choudhary, Laura Tronci, Angela Bachi, Christopher Bruhn.

Glutamine analogs are potent suppressors of general glutamine metabolism with anti-cancer activity. 6-diazo-5-oxo-L-norleucine (DON) is an orally available glutamine analog which has been recently improved by structural modification for cancer treatment. Here, we explored the chemogenomic landscape of DON sensitivity using budding yeast as model organism. We identify evolutionarily conserved proteins that mediate cell resistance to glutamine analogs, namely Ura8CTPS1/2, Hpt1HPRT1, Mec1ATR, Rad53CHK1/CHK2 and Rtg1. We describe a function of Ura8 as inducible CTP synthase responding to inhibition of glutamine metabolism and propose a model for its regulation by CTP levels and Nrd1-dependent transcription termination at a cryptic unstable transcript. Disruption of the inducible CTP synthase under DON exposure hyper-activates the Mec1-Rad53 DNA damage response (DDR) pathway, which prevents chromosome breakage. Simultaneous inhibition of CTP synthase and Mec1 kinase synergistically sensitizes cells to DON, whereas CTP synthase over-expression hampers DDR mutant sensitivity. Using genome-wide suppressor screening, we identify factors promoting DON-induced CTP depletion (TORC1, glutamine transporter) and DNA breakage in DDR mutants. Together, our results identify CTP regulation and the Mec1-Rad53 DDR axis as key glutamine analog response pathways, and provide a rationale for the combined targeting of glutamine and CTP metabolism in DDR-deficient cancers.

DOI: https://doi.org/10.1371/journal.pgen.1010101
J Biol Chem. 2022 Feb 26. pii: S0021-9258(22)00217-4. [Epub ahead of print] 101777

Intra-S phase checkpoint kinase Chk1 dissociates replication proteins Treslin and TopBP1 through multiple mechanisms during replication stress.

Rebecca L Kelly, Amelia M Huehls, Annapoorna Venkatachalam, Catherine J Huntoon, Yuichi J Machida, Larry M Karnitz.

  Replication stress impedes DNA polymerase progression causing activation of the ATR signaling pathway, which promotes the intra-S phase checkpoint activity through phosphorylation of Chk1. Chk1 suppresses replication origin firing, in part, by disrupting the interaction between the pre-initiation complex components Treslin and TopBP1, an interaction that is mediated by TopBP1 BRCT domain-binding to two cyclin-dependent kinase (CDK) phosphorylation sites, T968 and S1000, in Treslin. Two non-exclusive models for how Chk1 regulates the Treslin-TopBP1 interaction have been proposed in the literature: in one model, these proteins dissociate due to a Chk1-induced decrease in CDK activity that reduces phosphorylation of the Treslin sites required for TopBP1 binding; and in the second model, Chk1 directly phosphorylates Treslin, resulting in dissociation from TopBP1. However, these models have not been formally examined. We show here that Treslin T968 phosphorylation was decreased in a Chk1-dependent manner, while Treslin S1000 phosphorylation was unchanged, demonstrating that T968 and S1000 are differentially regulated. However, CDK2-mediated phosphorylation alone did not fully account for Chk1 regulation of the Treslin-TopBP1 interaction. We also identified additional Chk1 phosphorylation sites on Treslin that contributed to disruption of the Treslin-TopBP1 interaction, including S1114. Finally, we showed that both of the proposed mechanisms regulate origin firing in cancer cell line models undergoing replication stress, with the prevalence of each mechanism varying among cell lines. This study demonstrates that Chk1 regulates Treslin through multiple mechanisms to promote efficient dissociation of Treslin and TopBP1, and furthers our understanding of Treslin regulation during the intra-S phase checkpoint.

Keywords:  Chk1; Chk1 inhibitors; TopBP1; Treslin; cytarabine; origin firing; replication stress

DOI:  https://doi.org/10.1016/j.jbc.2022.101777
Trends Cancer. 2022 Feb 26. pii: S2405-8033(22)00022-X. [Epub ahead of print]

Fanconi anemia proteins and genome fragility: unraveling replication defects for cancer therapy.

Nibal Badra Fajardo, Stavros Taraviras, Zoi Lygerou.

  Accurate and complete genome duplication is crucial to maintain cell survival and prevent malignant transformation. The Fanconi anemia (FA) pathway has traditionally been associated with the repair of DNA interstrand crosslinks that impede the progression of the replication machinery. Recent studies demonstrate that FA proteins also regulate cell-cycle checkpoints and/or promote replication fork remodeling in response to multiple DNA impediments, and redefine the FA pathway as a fundamental mechanism to preserve genome integrity upon different insults. Alterations in FA genes fuel genomic fragility and constitute a driving force of tumorigenesis. We highlight current understanding of FA signaling in safeguarding genome stability during replication, and discuss the identification of novel determinants of cancer cell survival in FA-deficient tumors.

Keywords:  DNA repair; Fanconi anemia; checkpoint regulation; fork stability; replication stress; synthetic lethality

DOI:  https://doi.org/10.1016/j.trecan.2022.01.015
Dev Cell. 2022 Feb 23. pii: S1534-5807(22)00079-X. [Epub ahead of print]

Chk1 dynamics in G2 phase upon replication stress predict daughter cell outcome.

Vicente Lebrec, Marion Poteau, Jean-Philippe Morretton, Olivier Gavet.

  In human cells, ATR/Chk1 signaling couples S phase exit with the expression of mitotic inducers and prevents premature mitosis upon replication stress (RS). Nonetheless, under-replicated DNA can persist at mitosis, prompting chromosomal instability. To decipher how the DNA replication checkpoint (DRC) allows cells to enter mitosis over time upon RS, we developed a FRET-based Chk1 activity sensor. During unperturbed growth, a basal Chk1 activity level is sustained throughout S phase and relies on replication origin firing. Incremental RS triggers stepwise Chk1 over-activation that delays S-phase, suggesting a rheostat-like role for DRC coupled with the replication machinery. Upon RS, Chk1 is inactivated as DNA replication terminates but surprisingly is reactivated in a subset of G2 cells, which relies on Cdk1/2 and Plk1 and prevents mitotic entry. Cells can override active Chk1 signaling and reach mitosis onset, revealing checkpoint adaptation. Cell division following Chk1 reactivation in G2 results in a p53/p21-dependent G1 arrest, eliminating the daughter cells from proliferation.

Keywords:  DNA replication checkpoint; FRET biosensor; G2 phase; cell cycle recovery; checkpoint adaptation; checkpoint kinase 1; replication stress

DOI:  https://doi.org/10.1016/j.devcel.2022.02.013
DNA Repair (Amst). 2022 Apr;pii: S1568-7864(22)00030-1. [Epub ahead of print]112 103301

Histone variants: The unsung guardians of the genome.

Ernest O N Phillips, Akash Gunjan.

  Histones H2A, H2B, H3, H4 and H1 are highly conserved, positively charged proteins which form a disc-shaped protein core around which genomic DNA is wrapped to form a nucleosome. Immediately following DNA synthesis, replication-dependent canonical histones help package the DNA into nucleosomes to form compact chromatin fibers that can fit within the confines of the cell nucleus. Histone variants, which vary from the canonical histones in their primary amino acid sequence and expression patterns, replace their canonical counterparts throughout the cell cycle in important biological processes such as transcription, replication, DNA repair and heterochromatin formation. DNA damage is a continual threat to genomic stability and cell survival. Unrepaired DNA lesions are either lethal or can promote mutations if the damaged cells escape programmed cell death due to apoptosis. In order to repair DNA damage, cells use multiple DNA repair pathways, all of which require the recruitment of a multiple DNA damage signaling and repair factors. In order for these repair factors to be recruited efficiently and function properly at sites of DNA damage, the local chromatin environment surrounding the DNA lesion is often altered. Cells are able to regulate chromatin structure in the vicinity of DNA lesions through the addition of posttranslational modifications on histones and DNA, as well as through histone variant incorporation or removal. Recruitment or removal of histone variants at sites of DNA damage can alter the local chromatin structure by destabilizing it and making it more accessible to repair factors. Alternatively, some histone variants and their modifications may also provide specific binding sites for the recruitment of various DNA repair factors, thereby influencing repair pathway choice or repair efficiency, or both. This review seeks to provide an overview of our current understanding of the roles played by histone variants in DNA repair, especially in mammalian cells.

Keywords:  Cancer; Chromatin; DNA damage; DNA repair; Genome stability; Histone variant

DOI:  https://doi.org/10.1016/j.dnarep.2022.103301
Genome Biol. 2022 Feb 28. 23(1): 64

LAP2α preserves genome integrity through assisting RPA deposition on damaged chromatin.

Kaiwen Bao, Qi Zhang, Shuai Liu, Nan Song, Qiushi Guo, Ling Liu, Shanshan Tian, Jihui Hao, Yi Zhu, Kai Zhang, Ding Ai, Jie Yang, Zhi Yao, Roland Foisner, Lei Shi.

   BACKGROUND: Single-stranded DNA (ssDNA) coated with replication protein A (RPA) acts as a key platform for the recruitment and exchange of genome maintenance factors in DNA damage response. Yet, how the formation of the ssDNA-RPA intermediate is regulated remains elusive.
RESULTS: Here, we report that the lamin-associated protein LAP2α is physically associated with RPA, and LAP2α preferentially facilitates RPA deposition on damaged chromatin via physical contacts between LAP2α and RPA1. Importantly, LAP2α-promoted RPA binding to ssDNA plays a critical role in protection of replication forks, activation of ATR, and repair of damaged DNA. We further demonstrate that the preference of LAP2α-promoted RPA loading on damaged chromatin depends on poly ADP-ribose polymerase PARP1, but not poly(ADP-ribosyl)ation.
CONCLUSIONS: Our study provides mechanistic insight into RPA deposition in response to DNA damage and reveals a genome protection role of LAP2α.

Keywords:  ATR activation; Genome stability; Homologous recombination; LAP2α; PARP1; RPA loading

DOI:  https://doi.org/10.1186/s13059-022-02638-6
Proc Natl Acad Sci U S A. 2022 Mar 08. 119(10): e2113233119

Rapid recruitment of p53 to DNA damage sites directs DNA repair choice and integrity.

Yu-Hsiu Wang, Teresa L F Ho, Anushya Hariharan, Hui Chin Goh, Yao Liang Wong, Nicole S Verkaik, May Yin Lee, Wai Leong Tam, Dik C van Gent, Ashok R Venkitaraman, Michael P Sheetz, David P Lane.

  SignificanceOur work focuses on the critical longstanding question of the nontranscriptional role of p53 in tumor suppression. We demonstrate here that poly(ADP-ribose) polymerase (PARP)-dependent modification of p53 enables rapid recruitment of p53 to damage sites, where it in turn directs early repair pathway selection. Specifically, p53-mediated recruitment of 53BP1 at early time points promotes nonhomologous end joining over the more error-prone microhomology end-joining. Similarly, p53 directs nucleotide excision repair by mediating DDB1 recruitment. This property of p53 also correlates with tumor suppression in vivo. Our study provides mechanistic insight into how certain transcriptionally deficient p53 mutants may retain tumor-suppressive functions through regulating the DNA damage response.

Keywords:  DNA repair; laser microirradiation; p53; recruitment kinetics; tumor suppression

DOI:  https://doi.org/10.1073/pnas.2113233119
Cell Mol Life Sci. 2022 Mar 01. 79(3): 166

XPG: a multitasking genome caretaker.

Alba Muniesa-Vargas, Arjan F Theil, Cristina Ribeiro-Silva, Wim Vermeulen, Hannes Lans.

  The XPG/ERCC5 endonuclease was originally identified as the causative gene for Xeroderma Pigmentosum complementation group G. Ever since its discovery, in depth biochemical, structural and cell biological studies have provided detailed mechanistic insight into its function in excising DNA damage in nucleotide excision repair, together with the ERCC1-XPF endonuclease. In recent years, it has become evident that XPG has additional important roles in genome maintenance that are independent of its function in NER, as XPG has been implicated in protecting replication forks by promoting homologous recombination as well as in resolving R-loops. Here, we provide an overview of the multitasking of XPG in genome maintenance, by describing in detail how its activity in NER is regulated and the evidence that points to important functions outside of NER. Furthermore, we present the various disease phenotypes associated with inherited XPG deficiency and discuss current ideas on how XPG deficiency leads to these different types of disease.

Keywords:  DNA damage response; NER; Structure; XPG/ERCC5; Xeroderma pigmentosum–Cockayne syndrome

DOI:  https://doi.org/10.1007/s00018-022-04194-5
Front Oncol. 2022 ;12 832816

Emvododstat, a Potent Dihydroorotate Dehydrogenase Inhibitor, Is Effective in Preclinical Models of Acute Myeloid Leukemia.

Arthur Branstrom, Liangxian Cao, Bansri Furia, Christopher Trotta, Marianne Santaguida, Jason D Graci, Joseph M Colacino, Balmiki Ray, Wencheng Li, Josephine Sheedy, Anna Mollin, Shirley Yeh, Ronald Kong, Richard Sheridan, John D Baird, Kylie O'Keefe, Robert Spiegel, Elizabeth Goodwin, Suzanne Keating, Marla Weetall.

  Blocking the pyrimidine nucleotide de novo synthesis pathway by inhibiting dihydroorotate dehydrogenase (DHODH) results in the cell cycle arrest and/or differentiation of rapidly proliferating cells including activated lymphocytes, cancer cells, or virally infected cells. Emvododstat (PTC299) is an orally bioavailable small molecule that inhibits DHODH. We evaluated the potential for emvododstat to inhibit the progression of acute myeloid leukemia (AML) using several in vitro and in vivo models of the disease. Broad potent activity was demonstrated against multiple AML cell lines, AML blasts cultured ex vivo from patient blood samples, and AML tumor models including patient-derived xenograft models. Emvododstat induced differentiation, cytotoxicity, or both in primary AML patient blasts cultured ex vivo with 8 of 10 samples showing sensitivity. AML cells with diverse driver mutations were sensitive, suggesting the potential of emvododstat for broad therapeutic application. AML cell lines that are not sensitive to emvododstat are likely to be more reliant on the salvage pathway than on de novo synthesis of pyrimidine nucleotides. Pharmacokinetic experiments in rhesus monkeys demonstrated that emvododstat levels rose rapidly after oral administration, peaking about 2 hours post-dosing. This was associated with an increase in the levels of dihydroorotate (DHO), the substrate for DHODH, within 2 hours of dosing indicating that DHODH inhibition is rapid. DHO levels declined as drug levels declined, consistent with the reversibility of DHODH inhibition by emvododstat. These preclinical findings provide a rationale for clinical evaluation of emvododstat in an ongoing Phase 1 study of patients with relapsed/refractory acute leukemias.

Keywords:  AML; DHODH; PTC299; differentiation; dihydroorotate dehydrogenase; emvododstat; pyrimidine nucleotide de novo synthesis

DOI:  https://doi.org/10.3389/fonc.2022.832816
Proc Natl Acad Sci U S A. 2022 Mar 08. 119(10): e2118940119

Interlocking activities of DNA polymerase β in the base excision repair pathway.

Adarsh Kumar, Andrew J Reed, Walter J Zahurancik, Sasha M Daskalova, Sidney M Hecht, Zucai Suo.

  SignificanceBase excision repair (BER) is one of the major DNA repair pathways used to fix a myriad of cellular DNA lesions. The enzymes involved in BER, including DNA polymerase β (Polβ), have been identified and characterized, but how they act together to efficiently perform BER has not been fully understood. Through gel electrophoresis, mass spectrometry, and kinetic analysis, we discovered that the two enzymatic activities of Polβ can be interlocked, rather than functioning independently from each other, when processing DNA intermediates formed in BER. The finding prompted us to hypothesize a modified BER pathway. Through conventional and time-resolved X-ray crystallography, we solved 11 high-resolution crystal structures of cross-linked Polβ complexes and proposed a detailed chemical mechanism for Polβ's 5'-deoxyribose-5-phosphate lyase activity.

Keywords:  DNA base excision repair pathway; DNA polymerase β; Schiff base formation; dRP lyase chemical mechanism; β-elimination

DOI:  https://doi.org/10.1073/pnas.2118940119
Mol Cell. 2022 Feb 17. pii: S1097-2765(22)00085-5. [Epub ahead of print]

Micropeptide PACMP inhibition elicits synthetic lethal effects by decreasing CtIP and poly(ADP-ribosyl)ation.

Chuanchao Zhang, Bo Zhou, Feng Gu, Hongmei Liu, Honglin Wu, Fuwen Yao, Hui Zheng, Hui Fu, Wei Chong, Shurui Cai, Min Huang, Xiaolu Ma, Zhifang Guo, Tingting Li, Wenyuan Deng, Meiwen Zheng, Qiao Ji, Yongliang Zhao, Yongjie Ma, Qi-En Wang, Tie-Shan Tang, Caixia Guo.

  Synthetic lethality through combinatorial targeting DNA damage response (DDR) pathways provides exciting anticancer therapeutic benefit. Currently, the long noncoding RNAs (lncRNAs) have been implicated in tumor drug resistance; however, their potential significance in DDR is still largely unknown. Here, we report that a human lncRNA, CTD-2256P15.2, encodes a micropeptide, named PAR-amplifying and CtIP-maintaining micropeptide (PACMP), with a dual function to maintain CtIP abundance and promote poly(ADP-ribosyl)ation. PACMP not only prevents CtIP from ubiquitination through inhibiting the CtIP-KLHL15 association but also directly binds DNA damage-induced poly(ADP-ribose) chains to enhance PARP1-dependent poly(ADP-ribosyl)ation. Targeting PACMP alone inhibits tumor growth by causing a synthetic lethal interaction between CtIP and PARP inhibitions and confers sensitivity to PARP/ATR/CDK4/6 inhibitors, ionizing radiation, epirubicin, and camptothecin. Our findings reveal that a lncRNA-derived micropeptide regulates cancer progression and drug resistance by modulating DDR, whose inhibition could be employed to augment the existing anticancer therapeutic strategies.

Keywords:  CtIP; DNA damage response; DNA double-strand break repair; PARP inhibitor; drug resistance; long noncoding RNA; micropeptide; poly(ADP-ribosyl)ation; synthetic lethal

DOI:  https://doi.org/10.1016/j.molcel.2022.01.020
Cell Death Discov. 2022 Feb 26. 8(1): 85

CDC7 kinase (DDK) inhibition disrupts DNA replication leading to mitotic catastrophe in Ewing sarcoma.

Jeffrey C Martin, Jennie R Sims, Ajay Gupta, Tamara J Hagoel, Lingqiu Gao, Miranda L Lynch, Anna Woloszynska, Thomas Melendy, Jeremy F Kane, Joseph Kuechle, Joyce E Ohm.

Ewing sarcoma is the second most common bone malignancy in children and adolescents. In recent years, a large body of evidence has emerged that suggests Ewing tumors harbor large amounts of replication stress (RS). CDC7, also known as DDK (DBF4-dependent kinase), is a serine/threonine kinase that is involved in a diverse array of cellular functions including the regulation of DNA replication initiation and activation of the RS response. Due to DDK's diverse roles during replication, coupled with the fact that there is an increased level of RS within Ewing tumors, we hypothesized that Ewing sarcoma cells would be particularly vulnerable to DDK inhibition. Here, we report that DDK inhibition resulted a significant reduction in cell viability and the induction of apoptosis, specifically in Ewing sarcoma cells. Treatment with DDK inhibitors dramatically reduced the rate of replication, prolonged S-phase, and led to a pronounced increase in phospho-CDC2 (Y15), indicating delay of mitotic entry. The induction of cell death corresponded to mitotic exit and G1 entry, suggesting improper mitotic progression. In accordance with this, we find that DDK inhibition caused premature mitotic entry resulting in mitotic abnormalities such as anaphase bridges, lagging chromosomes, and cells with >2 poles in Ewing sarcoma cells. This abnormal progression through mitosis resulted in mitotic catastrophe as evidenced by the formation of micronuclei and induction of DNA damage. Together, these findings suggest that DDK activity is required for the faithful and timely completion of DNA replication in Ewing cells and that DDK inhibition may present a viable therapeutic strategy for the treatment of Ewing sarcoma.

DOI: https://doi.org/10.1038/s41420-022-00877-x
DNA Repair (Amst). 2022 Apr;pii: S1568-7864(22)00032-5. [Epub ahead of print]112 103303

Double-strand breaks: When DNA repair events accidentally meet.

Shingo Fujii, Robert W Sobol, Robert P Fuchs.

  The cellular response to alkylation damage is complex, involving multiple DNA repair pathways and checkpoint proteins, depending on the DNA lesion, the cell type, and the cellular proliferation state. The repair of and response to O-alkylation damage, primarily O6-methylguaine DNA adducts (O6-mG), is the purview of O6-methylguanine-DNA methyltransferase (MGMT). Alternatively, this lesion, if left un-repaired, induces replication-dependent formation of the O6-mG:T mis-pair and recognition of this mis-pair by the post-replication mismatch DNA repair pathway (MMR). Two models have been suggested to account for MMR and O6-mG DNA lesion dependent formation of DNA double-strand breaks (DSBs) and the resulting cytotoxicity - futile cycling and direct DNA damage signaling. While there have been hints at crosstalk between the MMR and base excision repair (BER) pathways, clear mechanistic evidence for such pathway coordination in the formation of DSBs has remained elusive. However, using a novel protein capture approach, Fuchs and colleagues have demonstrated that DSBs result from an encounter between MMR-induced gaps initiated at alkylation induced O6-mG:C sites and BER-induced nicks at nearby N-alkylation adducts in the opposite strand. The accidental encounter between these two repair events is causal in the formation of DSBs and the resulting cellular response, documenting a third model to account for O6-mG induced cell death in non-replicating cells. This graphical review highlights the details of this Repair Accident model, as compared to current models, and we discuss potential strategies to improve clinical use of alkylating agents such as temozolomide, that can be inferred from the Repair Accident model.

Keywords:  Alkylation; Base excision repair; Cell death; Futile cycle; MGMT; Mismatch repair

DOI:  https://doi.org/10.1016/j.dnarep.2022.103303
iScience. 2022 Mar 18. 25(3): 103875

Sex disparities in DNA damage response pathways: Novel determinants in cancer formation and therapy.

Miriana Cardano, Giacomo Buscemi, Laura Zannini.

  Cancer incidence and survival are different between men and women. Indeed, females have a lesser risk and a better prognosis than males in many tumors unrelated to reproductive functions. Although the reasons for these disparities are still unknown, they constitute an important starting point for the development of personalized cancer therapies. One of the mechanisms that fuels carcinogenesis is the accumulation of defects in DNA damage response (DDR) pathways, a complex signaling cascade that senses DNA lesions and, depending on the severity, coordinates transient cell-cycle arrest, DNA replication, repair, apoptosis, and senescence, preventing genomic instability and cancer. Recently, evidence of sexual dimorphisms is emerging in these pathways, therefore providing new opportunities for precision medicine. Here, we will discuss current knowledge about sexual disparities in the DDR, their role in tumorigenesis and cancer progression, and the importance of considering sex contribution in both research and cancer therapies.

Keywords:  Cancer; Cell biology; Cellular physiology

DOI:  https://doi.org/10.1016/j.isci.2022.103875
Mol Cell. 2022 Feb 24. pii: S1097-2765(22)00115-0. [Epub ahead of print]

Single-molecule mapping of replisome progression.

Clémence Claussin, Jacob Vazquez, Iestyn Whitehouse.

  Fundamental aspects of DNA replication, such as the anatomy of replication stall sites, how replisomes are influenced by gene transcription, and whether the progression of sister replisomes is coordinated, are poorly understood. Available techniques do not allow the precise mapping of the positions of individual replisomes on chromatin. We have developed a method called Replicon-seq that entails the excision of full-length replicons by controlled nuclease cleavage at replication forks. Replicons are sequenced using Nanopore, which provides a single-molecule readout of long DNA. Using Replicon-seq, we found that sister replisomes function autonomously and yet progress through chromatin with remarkable consistency. Replication forks that encounter obstacles pause for a short duration but rapidly resume synthesis. The helicase Rrm3 plays a critical role both in mitigating the effect of protein barriers and with facilitating efficient termination. Replicon-seq provides a high-resolution means of defining how individual replisomes move across the genome.

Keywords:  DNA replication; Nanopore sequencing; Replicon-seq; Rrm3; rDNA; replication termination; replisome pausing; sister replisome

DOI:  https://doi.org/10.1016/j.molcel.2022.02.010
Front Genet. 2021 ;12 809832

Genomic Reporter Constructs to Monitor Pathway-Specific Repair of DNA Double-Strand Breaks.

Bert van de Kooij, Haico van Attikum.

  Repair of DNA Double-Strand Breaks (DSBs) can be error-free or highly mutagenic, depending on which of multiple mechanistically distinct pathways repairs the break. Hence, DSB-repair pathway choice directly affects genome integrity, and it is therefore of interest to understand the parameters that direct repair towards a specific pathway. This has been intensively studied using genomic reporter constructs, in which repair of a site-specific DSB by the pathway of interest generates a quantifiable phenotype, generally the expression of a fluorescent protein. The current developments in genome editing with targetable nucleases like Cas9 have increased reporter usage and accelerated the generation of novel reporter constructs. Considering these recent advances, this review will discuss and compare the available DSB-repair pathway reporters, provide essential considerations to guide reporter choice, and give an outlook on potential future developments.

Keywords:  double-strand break repair pathway choice; end-joining; genomic reporter constructs; homologous recombination; single-strand annealing

DOI:  https://doi.org/10.3389/fgene.2021.809832
Chem Res Toxicol. 2022 Mar 03.

Effects of N7-Alkylguanine Conformation and Metal Cofactors on the Translesion Synthesis by Human DNA Polymerase η.

Hunmin Jung, Naveen Kumar Rayala, Seongmin Lee.

Non-enzymatic alkylation on DNA often generates N7-alkyl-2'-deoxyguanosine (N7alkylG) adducts as major lesions. N7alkylG adducts significantly block replicative DNA polymerases and can be bypassed by translesion synthesis (TLS) polymerases such as polymerase η (polη). To gain insights into the bypass of N7alkylG by TLS polymerases, we conducted kinetic and structural studies of polη catalyzing across N7BnG, a genotoxic lesion generated by the carcinogenic N-nitrosobenzylmethylamine. The presence of templating N7BnG in the polη catalytic site decreased the replication fidelity by ∼9-fold, highlighting the promutagenicity of N7BnG. The catalytic efficiency for dCTP incorporation opposite N7BnG decreased ∼22-fold and ∼7-fold compared to the incorporation opposite undamaged guanine in the presence of Mg2+ and Mn2+, respectively. A crystal structure of the complexes grown with polη, templating N7BnG, incoming dCTP, and Mg2+ ions showed the lack of the incoming nucleotide and metal cofactors in the polη catalytic site. Interestingly, the templating N7BnG adopted a syn conformation, which has not been observed in the published N7alkylG structures. The preferential formation of syn-N7BnG conformation at the templating site may deter the binding of an incoming dCTP, causing the inefficient bypass by polη. In contrast, the use of Mn2+ in place of Mg2+ in co-crystallization yielded a ternary complex displaying an anti-N7BnG:dCTP base pair and catalytic metal ions, which would be a close mimic of a catalytically competent state. We conclude that certain bulky N7-alkylG lesions can slow TLS polymerase-mediated bypass by adopting a catalytically unfavorable syn conformation in the replicating base pair site.

DOI: https://doi.org/10.1021/acs.chemrestox.1c00416
Cell Mol Life Sci. 2022 Feb 28. 79(3): 160

Defective repair of topoisomerase I induced chromosomal damage in Huntington's disease.

Nelma M Palminha, Cleide Dos Santos Souza, Jon Griffin, Chunyan Liao, Laura Ferraiuolo, Sherif F El-Khamisy.

  Topoisomerase1 (TOP1)-mediated chromosomal breaks are endogenous sources of DNA damage that affect neuronal genome stability. Whether TOP1 DNA breaks are sources of genomic instability in Huntington's disease (HD) is unknown. Here, we report defective 53BP1 recruitment in multiple HD cell models, including striatal neurons derived from HD patients. Defective 53BP1 recruitment is due to reduced H2A ubiquitination caused by the limited RNF168 activity. The reduced availability of RNF168 is caused by an increased interaction with p62, a protein involved in selective autophagy. Depletion of p62 or disruption of the interaction between RNAF168 and p62 was sufficient to restore 53BP1 enrichment and subsequent DNA repair in HD models, providing new opportunities for therapeutic interventions. These findings are reminiscent to what was described for p62 accumulation caused by C9orf72 expansion in ALS/FTD and suggest a common mechanism by which protein aggregation perturb DNA repair signaling.

Keywords:  Chromatin ubiquitination; DNA repair; Huntington’s disease; RNF168; TOP1cc; p62/SQSTM1

DOI:  https://doi.org/10.1007/s00018-022-04204-6
Nat Rev Mol Cell Biol. 2022 Feb 28.

Human topoisomerases and their roles in genome stability and organization.

Yves Pommier, André Nussenzweig, Shunichi Takeda, Caroline Austin.

Human topoisomerases comprise a family of six enzymes: two type IB (TOP1 and mitochondrial TOP1 (TOP1MT), two type IIA (TOP2A and TOP2B) and two type IA (TOP3A and TOP3B) topoisomerases. In this Review, we discuss their biochemistry and their roles in transcription, DNA replication and chromatin remodelling, and highlight the recent progress made in understanding TOP3A and TOP3B. Because of recent advances in elucidating the high-order organization of the genome through chromatin loops and topologically associating domains (TADs), we integrate the functions of topoisomerases with genome organization. We also discuss the physiological and pathological formation of irreversible topoisomerase cleavage complexes (TOPccs) as they generate topoisomerase DNA-protein crosslinks (TOP-DPCs) coupled with DNA breaks. We discuss the expanding number of redundant pathways that repair TOP-DPCs, and the defects in those pathways, which are increasingly recognized as source of genomic damage leading to neurological diseases and cancer.

DOI: https://doi.org/10.1038/s41580-022-00452-3
Acta Biochim Pol. 2022 Mar 02. 69(1): 227-232

The influence of cdG on 8-oxodG excision by OGG1 and FPG glycosylases.

Michał Szewczuk, Bolesław Karwowski.

Human genome is exposed to the variety of damaging factors, such as ionizing radiation. 5',8-cyclo-2'-deoxypurines (cdPus) are well described unfavorable outcomes of DNA damage, especially devastating as a part of clustered DNA lesions (CDL). Since cdPus are not repaired by base excision repair (BER) and poorly repaired by nucleotide excision repair (NER), it is important to unveil the mechanisms of cdPus action within the genome. In this study the influence of both 5'S and 5'R diastereomers of 5',8-cyclo-2'-deoxyguanosine (cdG) on the activity of OGG1 and FPG was examined. Synthetic oligonucleotides containing cdG and two molecules of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG) were designed as model of single-stranded CDL. The activity of both enzymes increased in the presence of cdG, compared to the control DNA strands, and the increase was greater in the case of 5'R diastereomer. These results are supported by previous studies concerning cdPus and confirm the impact of lesions proximity on the DNA repair efficiency. Due to the biological importance of cdPus, it is necessary to understand the mechanisms of lesions recognition by repair proteins in further studies.

DOI: https://doi.org/10.18388/abp.2020_5966
Int J Biochem Cell Biol. 2022 Feb 24. pii: S1357-2725(22)00034-6. [Epub ahead of print] 106189

NAMPT: a critical driver and therapeutic target for cancer.

Massimiliano Gasparrini, Valentina Audrito.

  Nicotinamide phosphoribosyltransferase (NAMPT) possesses a vital role in mammalian cells due to its activity as a rate-limiting enzyme in the biosynthesis of nicotinamide adenine dinucleotide (NAD) from nicotinamide. NAD is an essential redox cofactor, but it also functions as a substrate for NAD-consuming enzymes, regulating multiple cellular processes such as DNA repair and gene expression, fundamental to sustain tumor growth and survival and energetic needs. A common strategy that several tumor types adopt to sustain NAD synthesis is to over-express NAMPT. However, beside its intracellular functions, this enzyme has a second life outside of cells exerting cytokine-like functions and mediating pro-inflammatory conditions activating signaling pathways. While the effects of NAMPT/NAD axis on energetic metabolism in tumors has been well-established, increasing evidence demonstrated the impact of NAMPT over-expression (intra-/extra-cellular) on several tumor cellular processes, including DNA repair, gene expression, signaling pathways, proliferation, invasion, stemness, phenotype plasticity, metastatization, angiogenesis, immune regulation, and drug resistance. For all these reasons, NAMPT targeting has emerged as promising anti-cancer strategy to deplete NAD and impair cellular metabolism, but also to counteract the other NAMPT-related functions. In this review, we summarize the key role of NAMPT in multiple biological processes implicated in cancer biology and the impact of NAMPT inhibition as therapeutic strategy for cancer treatment.

Keywords:  NAD; NAMPT; cancer biology; cancer therapy; immune cell regulation; metabolism; tumor microenvironment

DOI:  https://doi.org/10.1016/j.biocel.2022.106189
Expert Opin Drug Metab Toxicol. 2022 Mar 03.

Are we still on the right Path(way)?: the altered expression of the pentose phosphate pathway in solid tumors and the potential of its inhibition in combination therapy.

Caroline J W Meskers, Marika Franczak, Ryszard T Smolenski, Elisa Giovannetti, Godefridus J Peters.

   INTRODUCTION: The pentose phosphate pathway (PPP) branches from glycolysis and is crucial for cell growth, since it provides necessary compounds for anabolic reactions, nucleotide synthesis and detoxification of reactive-oxygen-species (ROS). Overexpression of PPP enzymes has been reported in multiple cancer types and linked to therapy resistance, making their inhibition interesting targets for anti-cancer therapies.
AREAS COVERED: This review summarizes the extent of PPP upregulation across different cancer types, and the non-metabolic functions that PPP-enzymes might contribute to cancer initiation and maintenance. The effects of PPP-inhibition and their combinations with chemotherapeutics are summarized. We searched the databases provided by the University of Amsterdam to characterize the altered expression of the PPP across different cancer types, and to identify the effects of PPP-inhibition.
EXPERT OPINION: It can be concluded that there are synergistic and additive effects of PPP-inhibition and various classes of chemotherapeutics. These effects may be attributed to the increased susceptibility to ROS. However the toxicity, low efficacy and off-target effects of PPP-inhibitors make application in clinical practice challenging. Novel inhibitors are currently being developed, which could make PPP-inhibition a potential therapeutic strategy in the future, especially in combination with conventional chemotherapeutics and the inhibition of other metabolic pathways.

Keywords:  NAD/NADH; cancer; chemotherapy; glycolysis; pentose phosphate pathway

DOI:  https://doi.org/10.1080/17425255.2022.2049234
Nucleic Acids Res. 2022 Mar 02. pii: gkac143. [Epub ahead of print]

8-oxodG accumulation within super-enhancers marks fragile CTCF-mediated chromatin loops.

Giovanni Scala, Francesca Gorini, Susanna Ambrosio, Andrea M Chiariello, Mario Nicodemi, Luigi Lania, Barbara Majello, Stefano Amente.

8-Oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG), a major product of the DNA oxidization process, has been proposed to have an epigenetic function in gene regulation and has been associated with genome instability. NGS-based methodologies are contributing to the characterization of the 8-oxodG function in the genome. However, the 8-oxodG epigenetic role at a genomic level and the mechanisms controlling the genomic 8-oxodG accumulation/maintenance have not yet been fully characterized. In this study, we report the identification and characterization of a set of enhancer regions accumulating 8-oxodG in human epithelial cells. We found that these oxidized enhancers are mainly super-enhancers and are associated with bidirectional-transcribed enhancer RNAs and DNA Damage Response activation. Moreover, using ChIA-PET and HiC data, we identified specific CTCF-mediated chromatin loops in which the oxidized enhancer and promoter regions physically associate. Oxidized enhancers and their associated chromatin loops accumulate endogenous double-strand breaks which are in turn repaired by NHEJ pathway through a transcription-dependent mechanism. Our work suggests that 8-oxodG accumulation in enhancers-promoters pairs occurs in a transcription-dependent manner and provides novel mechanistic insights on the intrinsic fragility of chromatin loops containing oxidized enhancers-promoters interactions.

DOI: https://doi.org/10.1093/nar/gkac143
Cell Death Dis. 2022 02 26. 13(2): 193

Identification of Nanog as a novel inhibitor of Rad51.

Ying Xin, Juanjuan Wang, Yahong Wu, Qianqian Li, Mingyang Dong, Chang Liu, Qijia He, Ruifeng Wang, Dian Wang, Sen Jiang, Wei Xiao, Yang Tian, Weiwei Zhang.

To develop inhibitors targeting DNA damage repair pathways is important to improve the effectiveness of chemo- and radiotherapy for cancer patients. Rad51 mediates homologous recombination (HR) repair of DNA damages. It is widely overexpressed in human cancers and overwhelms chemo- and radiotherapy-generated DNA damages through enhancing HR repair signaling, preventing damage-caused cancer cell death. Therefore, to identify inhibitors of Rad51 is important to achieve effective treatment of cancers. Transcription factor Nanog is a core regulator of embryonic stem (ES) cells for its indispensable role in stemness maintenance. In this study, we identified Nanog as a novel inhibitor of Rad51. It interacts with Rad51 and inhibits Rad51-mediated HR repair of DNA damage through its C/CD2 domain. Moreover, Rad51 inhibition can be achieved by nanoscale material- or cell-penetrating peptide (CPP)-mediated direct delivery of Nanog-C/CD2 peptides into somatic cancer cells. Furthermore, we revealed that Nanog suppresses the binding of Rad51 to single-stranded DNAs to stall the HR repair signaling. This study provides explanation for the high γH2AX level in unperturbed ES cells and early embryos, and suggests Nanog-C/CD2 as a promising drug candidate applied to Rad51-related basic research and therapeutic application studies.

DOI: https://doi.org/10.1038/s41419-022-04644-9
Drug Resist Updat. 2022 Jan 21. pii: S1368-7646(22)00008-5. [Epub ahead of print]61 100805

Different mechanisms of drug resistance to hypomethylating agents in the treatment of myelodysplastic syndromes and acute myeloid leukemia.

Kristína Šimoničová, Ľuboš Janotka, Helena Kavcová, Zdena Sulová, Albert Breier, Lucia Messingerova.

  Resistance to the hypomethylating agents (HMAs) 5-azacytidine (AZA) and 5-aza-2'-deoxycytidine (DAC) represents a major obstacle in the treatment of elderly patients with myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML) which are not suitable for hematopoietic stem cells transplantation. Approximately 50 % of patients do not respond to HMA treatment because of intrinsic (primary) resistance, while others could acquire drug resistance during the repeated cycles of the treatment. To prevent, delay or surmount resistance development, the molecular mechanisms underlying drug resistance must be first identified. This is crucial as no further standard therapeutic opportunities are available for these patients who failed hypomethylating agents-based treatment. The current review provides an updated information about the different mechanisms that may contribute to the development of resistance to HMAs. Despite the similar structure and mechanism of action of HMA, several studies did not report the expected development of cross-resistance. It is clear that in addition to the common modalities of chemoresistance, there must be some specific mechanisms of drug resistance. Changes in transport and metabolism of HMAs are among the most studied mechanisms of resistance. Drug uptake provided by two solute carrier (SLC) families: SLC28 and SLC29 (also known as the concentrative and equilibrative nucleoside transporter families, respectively), could represent one of the mechanisms of cross-resistance. Changes in the metabolism of these drugs are the most likely mechanism responsible for the unique mode of resistance to AZA and DAC. Deoxycytidine kinase and uridine-cytidine kinase due to their necessity for drug activation, each could represent one of the response markers to treatment with DAC and AZA, respectively. Other mechanisms involved in the development of resistance common for both drugs involved: i. increased DNA repair (caused for example by constitutive activation of the ATM/BRCA1 pathway and inhibition of p53-dependent apoptosis); ii. changes in the regulation of apoptosis/disrupted apoptotic pathways (specifically increased levels of the anti-apoptotic protein BCL2) and iii. increased resilience of leukemic stem cells to multiple drugs including HMAs. Despite intense research on the resistance of MDS and AML patients to HMAs, the mechanisms that may reduce the response of these cells to HMAs are not known in detail. We herein highlight the most important directions that future research should take.

Keywords:  5-aza-2'-deoxycytidine; 5-azacytidine; Acute myeloid leukemia; Hypomethylating agents; Myelodysplastic syndromes; Resistance

DOI:  https://doi.org/10.1016/j.drup.2022.100805
Nucleosides Nucleotides Nucleic Acids. 2022 Feb 28. 1-16

The role of p53 and p21 on 8-chloro-adenosine-induced cellular response.

Scott Teakell, Lisa S Chen, Christine M Stellrecht, Varsha Gandhi.

  8-Chloro-adenosine (8-Cl-Ado) is currently in phase I clinical trial. Activation of p53 and transactivation of p21 regulate cell fate after genotoxic insult. Using HCT-116-isogenic-cell-lines, we evaluated the role of p53/p21 after 8-Cl-Ado-mediated response. Following 30 µM 8-Cl-Ado treatment, RNA synthesis was inhibited, p53 protein was stabilized, and p21 expression was activated. None of the cell types were arrested in G1/S phase, however, cells lacking p53 were blocked in G2/M. These cells had the least increase in apoptotic cells, although clonogenic survival demonstrated equal inhibition in all 4 cell types. Collectively, irrespective of p53 and p21 status, 8-Cl-Ado-induced cytotoxicity was similar.

Keywords:  8-Chloro-adenosine; Cell survival; colon cancer; nucleoside analogs; p21; p53; transcription inhibitors

DOI:  https://doi.org/10.1080/15257770.2022.2038200
ACS Omega. 2022 Feb 22. 7(7): 6046-6052

Trapping of 5-Fluorodeoxyuridine Monophosphate by Thymidylate Synthase Confers Resistance to 5-Fluorouracil.

Chinatsu Kurasaka, Nana Nishizawa, Yoko Ogino, Akira Sato.

The major metabolite of the anticancer agent 5-fluorouracil (5-FU) is 5-fluorodeoxyuridine monophosphate (FdUMP), which is a potent inhibitor of thymidylate synthase (TS). Recently, we hypothesized that 5-FU-resistant colorectal cancer (CRC) cells have increased levels of TS protein relative to 5-FU-sensitive CRC cells and use a fraction of their TS to trap FdUMP, which results in resistance to 5-FU. In this study, we analyzed the difference between the regulation of the balance of the free, active form of TS and the inactive FdUMP-TS form in 5-FU-resistant HCT116 cells and parental HCT116 cells. Silencing of TYMS, the gene that encodes TS, resulted in greater enhancement of the anticancer effect of 5-FU in the 5-FU-resistant HCT116RF10 cells than in the parental HCT116 cells. In addition, the trapping of FdUMP by TS was more effective in the 5-FU-resistant HCT116RF10 cells than in the parental HCT116 cells. Our observations suggest that the regulation of the balance between the storage of the active TS form and the accumulation of FdUMP-TS is responsible for direct resistance to 5-FU. The findings provide a better understanding of 5-FU resistance mechanisms and may enable the development of anticancer strategies that reverse the sensitivity of 5-FU resistance in CRC cells.

DOI: https://doi.org/10.1021/acsomega.1c06394
Sci Transl Med. 2022 Mar 02. 14(634): eabl6992

SERAC1 is a component of the mitochondrial serine transporter complex required for the maintenance of mitochondrial DNA.

Hezhi Fang, Anran Xie, Miaomiao Du, Xueyun Li, Kaiqiang Yang, Yinxu Fu, Xiangshu Yuan, Runxiao Fan, Weidong Yu, Zhuohua Zhou, Tiantian Sang, Ke Nie, Jin Li, Qiongya Zhao, Zhehui Chen, Yanling Yang, Chaoyang Hong, Jianxin Lyu.

SERAC1 deficiency is associated with the mitochondrial 3-methylglutaconic aciduria with deafness, (hepatopathy), encephalopathy, and Leigh-like disease [MEGD(H)EL] syndrome, but the role of SERAC1 in mitochondrial physiology remains unknown. Here, we generated Serac1-/- mice that mimic the major diagnostic clinical and biochemical phenotypes of the MEGD(H)EL syndrome. We found that SERAC1 localizes to the outer mitochondrial membrane and is a protein component of the one-carbon cycle. By interacting with the mitochondrial serine transporter protein SFXN1, SERAC1 facilitated and was required for SFXN1-mediated serine transport from the cytosol to the mitochondria. Loss of SERAC1 impaired the one-carbon cycle and disrupted the balance of the nucleotide pool, which led to primary mitochondrial DNA (mtDNA) depletion in mice, HEK293T cells, and patient-derived immortalized lymphocyte cells due to insufficient supply of nucleotides. Moreover, both in vitro and in vivo supplementation of nucleosides/nucleotides restored mtDNA content and mitochondrial function. Collectively, our findings suggest that MEGD(H)EL syndrome shares both clinical and molecular features with the mtDNA depletion syndrome, and nucleotide supplementation may be an effective therapeutic strategy for MEGD(H)EL syndrome.

DOI: https://doi.org/10.1126/scitranslmed.abl6992
Mol Cell. 2022 Mar 03. pii: S1097-2765(22)00117-4. [Epub ahead of print]82(5): 889-890

Releasing the trap: How the segregase p97 extracts PARP1 from chromatin.

Maximilian J Götz, Julian Stingele.

Krastev et al. (2022) identify a cellular mechanism that counteracts cytotoxic trapping of PARP1 induced by clinical PARP inhibitors. SUMO-targeted ubiquitylation of trapped PARP1 is shown to trigger the enzymes' extraction from chromatin by the p97 ATPase.

DOI: https://doi.org/10.1016/j.molcel.2022.02.012