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Current Genetics Apr 2019The molecular chaperones Hsp70 and Hsp90 bind and fold a significant proportion of the proteome. They are responsible for the activity and stability of many... (Review)
Review
The molecular chaperones Hsp70 and Hsp90 bind and fold a significant proportion of the proteome. They are responsible for the activity and stability of many disease-related proteins including those in cancer. Substantial effort has been devoted to developing a range of chaperone inhibitors for clinical use. Recent studies have identified the oncogenic ribonucleotide reductase (RNR) complex as an interactor of chaperones. While several generations of RNR inhibitor have been developed for use in cancer patients, many of these produce severe side effects such as nausea, vomiting and hair loss. Development of more potent, less patient-toxic anti-RNR strategies would be highly desirable. Inhibition of chaperones and associated co-chaperone molecules in both cancer and model organisms such as budding yeast result in the destabilization of RNR subunits and a corresponding sensitization to RNR inhibitors. Going forward, this may form part of a novel strategy to target cancer cells that are resistant to standard RNR inhibitors.
Topics: Animals; Antineoplastic Agents; DNA Damage; DNA Replication; Enzyme Activation; Gene Expression Regulation; HSP70 Heat-Shock Proteins; HSP90 Heat-Shock Proteins; Humans; Molecular Chaperones; Neoplasms; Protein Binding; Proteomics; Ribonucleotide Reductases
PubMed: 30519713
DOI: 10.1007/s00294-018-0916-7 -
Journal of Biomedical Science May 2022Aberrant DNA repair pathways contribute to malignant transformation or disease progression and the acquisition of drug resistance in multiple myeloma (MM); therefore,...
BACKGROUND
Aberrant DNA repair pathways contribute to malignant transformation or disease progression and the acquisition of drug resistance in multiple myeloma (MM); therefore, these pathways could be therapeutically exploited. Ribonucleotide reductase (RNR) is the rate-limiting enzyme for the biosynthesis of deoxyribonucleotides (dNTPs), which are essential for DNA replication and DNA damage repair. In this study, we explored the efficacy of the novel RNR inhibitor, 4-hydroxysalicylanilide (HDS), in myeloma cells and xenograft model. In addition, we assessed the clinical activity and safety of HDS in patients with MM.
METHODS
We applied bioinformatic, genetic, and pharmacological approaches to demonstrate that HDS was an RNR inhibitor that directly bound to RNR subunit M2 (RRM2). The activity of HDS alone or in synergy with standard treatments was evaluated in vitro and in vivo. We also initiated a phase I clinical trial of single-agent HDS in MM patients (ClinicalTrials.gov: NCT03670173) to assess safety and efficacy.
RESULTS
HDS inhibited the activity of RNR by directly targeting RRM2. HDS decreased the RNR-mediated dNTP synthesis and concomitantly inhibited DNA damage repair, resulting in the accumulation of endogenous unrepaired DNA double-strand breaks (DSBs), thus inhibiting MM cell proliferation and inducing apoptosis. Moreover, HDS overcame the protective effects of IL-6, IGF-1 and bone marrow stromal cells (BMSCs) on MM cells. HDS prolonged survival in a MM xenograft model and induced synergistic anti-myeloma activity in combination with melphalan and bortezomib. HDS also showed a favorable safety profile and demonstrated clinical activity against MM.
CONCLUSIONS
Our study provides a rationale for the clinical evaluation of HDS as an anti-myeloma agent, either alone or in combination with standard treatments for MM.
TRIAL REGISTRATION
ClinicalTrials.gov, NCT03670173, Registered 12 September 2018.
Topics: DNA Breaks, Double-Stranded; DNA Damage; DNA Repair; DNA Replication; Humans; Multiple Myeloma; Ribonucleotide Reductases
PubMed: 35546402
DOI: 10.1186/s12929-022-00813-2 -
Annals of Botany Sep 2022Ribonucleotide reductase (RNR), functioning in the de novo synthesis of deoxyribonucleoside triphosphates (dNTPs), is crucial for DNA replication and cell cycle...
BACKGROUND AND AIMS
Ribonucleotide reductase (RNR), functioning in the de novo synthesis of deoxyribonucleoside triphosphates (dNTPs), is crucial for DNA replication and cell cycle progression. In most plants, the large subunits of RNR have more than one homologous gene. However, the different functions of these homologous genes in plant development remain unknown. In this study, we obtained the mutants of two large subunits of RNR in tomato and studied their functions.
METHODS
The mutant ylc1 was obtained by ethyl methyl sulfonate (EMS) treatment. Through map-based cloning, complementation and knock-out experiments, it was confirmed that YLC1 encodes a large subunit of RNR (SlRNRL1). The expression level of the genes related to cell cycle progression, chloroplast biogenesis and photosynthesis was assessed by RNA-sequencing. In addition, we knocked out SlRNRL2 (a SlRNRL1 homologue) using CRISPR-Cas9 technology in the tomato genome, and we down-regulated SlRNRL2 expression in the genetic background of slrnrl1-1 using a tobacco rattle virus-induced gene silencing (VIGS) system.
KEY RESULTS
The mutant slrnrl1 exhibited dwarf stature, chlorotic young leaves and smaller fruits. Physiological and transcriptomic analyses indicated that SlRNRL1 plays a crucial role in the regulation of cell cycle progression, chloroplast biogenesis and photosynthesis in tomato. The slrnrl2 mutant did not exhibit any visible phenotype. SlRNRL2 has a redundant function with SlRNRL1, and the double mutant slrnrl1slrnrl2 is lethal.
CONCLUSIONS
SlRNRL1 is essential for cell cycle progression, chloroplast biogenesis and photosynthesis. In addition, SlRNRL1 and SlRNRL2 possess redundant functions and at least one of these RNRLs is required for tomato survival, growth and development.
Topics: Cell Cycle; Chloroplasts; Gene Expression Regulation, Plant; Solanum lycopersicum; Photosynthesis; Ribonucleotide Reductases
PubMed: 35700127
DOI: 10.1093/aob/mcac078 -
Nature Communications Jul 2019Ribonucleotide reductase (RNR) catalyzes the de novo synthesis of deoxyribonucleoside diphosphates (dNDPs) to provide dNTP precursors for DNA synthesis. Here, we report...
Ribonucleotide reductase (RNR) catalyzes the de novo synthesis of deoxyribonucleoside diphosphates (dNDPs) to provide dNTP precursors for DNA synthesis. Here, we report that acetylation and deacetylation of the RRM2 subunit of RNR acts as a molecular switch that impacts RNR activity, dNTP synthesis, and DNA replication fork progression. Acetylation of RRM2 at K95 abrogates RNR activity by disrupting its homodimer assembly. RRM2 is directly acetylated by KAT7, and deacetylated by Sirt2, respectively. Sirt2, which level peak in S phase, sustains RNR activity at or above a threshold level required for dNTPs synthesis. We also find that radiation or camptothecin-induced DNA damage promotes RRM2 deacetylation by enhancing Sirt2-RRM2 interaction. Acetylation of RRM2 at K95 results in the reduction of the dNTP pool, DNA replication fork stalling, and the suppression of tumor cell growth in vitro and in vivo. This study therefore identifies acetylation as a regulatory mechanism governing RNR activity.
Topics: Acetylation; Camptothecin; Cell Cycle; Cell Line, Tumor; Cell Transformation, Neoplastic; DNA Damage; DNA Replication; Gene Expression Regulation, Neoplastic; Histone Acetyltransferases; Humans; Ribonucleoside Diphosphate Reductase; Ribonucleotide Reductases; S Phase; Sirtuin 2
PubMed: 31324785
DOI: 10.1038/s41467-019-11214-9 -
Current Opinion in Chemical Biology Feb 2020Stereotyped as a nexus of dNTP synthesis, the dual-subunit enzyme - ribonucleotide reductase (RNR) - is coming into view as a paradigm of oligomerization and... (Review)
Review
Stereotyped as a nexus of dNTP synthesis, the dual-subunit enzyme - ribonucleotide reductase (RNR) - is coming into view as a paradigm of oligomerization and moonlighting behavior. In the present issue of 'omics', we discuss what makes the larger subunit of this enzyme (RNR-α) so interesting, highlighting its emerging cellular interactome based on its unique oligomeric dynamism that dictates its compartment-specific occupations. Linking the history of the field with the multivariable nature of this exceedingly sophisticated enzyme, we further discuss implications of new data pertaining to DNA-damage response, S-phase checkpoints, and ultimately tumor suppression. We hereby hope to provide ideas for those interested in these fields and exemplify conceptual frameworks and tools that are useful to study RNR's broader roles in biology.
Topics: Animals; DNA Damage; DNA Helicases; Deoxyadenosines; Humans; Models, Molecular; Protein Interaction Maps; Protein Structure, Quaternary; Ribonucleotide Reductases
PubMed: 31734537
DOI: 10.1016/j.cbpa.2019.09.003 -
Journal of the American Chemical Society Apr 2021Ribonucleotide reductase (RNR) is an essential enzyme in DNA synthesis for all living organisms. It reduces ribonucleotides to the corresponding deoxyribonucleotides by...
Ribonucleotide reductase (RNR) is an essential enzyme in DNA synthesis for all living organisms. It reduces ribonucleotides to the corresponding deoxyribonucleotides by a reversible radical transfer mechanism. The active form of Ia RNR is composed of two subunits, α and β, which form an active asymmetric αβ complex. The radical transfer pathway involves a series of proton-coupled electron transfer (PCET) reactions spanning α and β over ∼32 Å. Herein, quantum mechanical/molecular mechanical free energy simulations of PCET between tyrosine residues Y730 and Y731 are performed on the recently solved cryo-EM structure of the active αβ complex, which includes a pre-turnover α/β pair with an ordered PCET pathway and a post-turnover α'/β' pair. The free energy surfaces in both the pre- and post-turnover states are computed. According to the simulations, forward radical transfer from Y731 to Y730 is thermodynamically favored in the pre-turnover state, and backward radical transfer is favored in the post-turnover state, consistent with the reversible mechanism. E623, a glutamate residue that is near these tyrosines only in the pre-turnover state, is discovered to play a key role in facilitating forward radical transfer by thermodynamically stabilizing the radical on Y730 through hydrogen-bonding and electrostatic interactions and lowering the free energy barrier via a proton relay mechanism. Introduction of fluorinated Y731 exhibits expected thermodynamic trends without altering the basic mechanism. These simulations suggest that E623 influences the directionality of PCET between Y731 and Y730 and predict that mutation of E623 will impact catalysis.
Topics: Electron Transport; Escherichia coli; Glutamic Acid; Hydrogen Bonding; Molecular Dynamics Simulation; Protein Structure, Quaternary; Protons; Quantum Theory; Ribonucleotide Reductases; Static Electricity; Thermodynamics; Tyrosine
PubMed: 33856807
DOI: 10.1021/jacs.1c02152 -
Current Issues in Molecular Biology 2012Malaria is caused by species in the apicomplexan genus Plasmodium, which infect hundreds of millions of people each year and kill close to one million. While malaria is... (Review)
Review
Malaria is caused by species in the apicomplexan genus Plasmodium, which infect hundreds of millions of people each year and kill close to one million. While malaria is the most notorious of the apicomplexan-caused diseases, other members of eukaryotic phylum Apicomplexa are responsible for additional, albeit less well-known, diseases in humans, economically important livestock, and a variety of other vertebrates. Diseases such as babesiosis (hemolytic anemia), theileriosis and East Coast Fever, cryptosporidiosis, and toxoplasmosis are caused by the apicomplexans Babesia, Theileria, Cryptosporidium and Toxoplasma, respectively. In addition to the loss of human life, these diseases are responsible for losses of billions of dollars annually. Hence, the research into new drug targets remains a high priority. Ribonucleotide reductase (RNR) is an essential enzyme found in all domains of life. It is the only means by which de novo synthesis of deoxyribonucleotides occurs, without which DNA replication and repair cannot proceed. RNR has long been the target of antiviral, antibacterial and anti-cancer therapeutics. Herein, we review the chemotherapeutic methods used to inhibit RNR, with particular emphasis on the role of RNR inhibition in Apicomplexa, and in light of the novel RNR R2_e2 subunit recently identified in apicomplexan parasites.
Topics: Amino Acid Sequence; Animals; Antiprotozoal Agents; Apicomplexa; Humans; Molecular Sequence Data; Molecular Targeted Therapy; Parasites; Protozoan Infections; Ribonucleotide Reductases
PubMed: 21791713
DOI: No ID Found -
European Journal of Medicinal Chemistry Apr 2024The development of cancer is influenced by several variables, including altered protein expression, and signaling pathways. Cancers are inherently heterogeneous and... (Review)
Review
The development of cancer is influenced by several variables, including altered protein expression, and signaling pathways. Cancers are inherently heterogeneous and exhibit genetic and epigenetic aberrations; therefore, developing therapies that act on numerous biological targets is encouraged. To achieve this, two approaches are employed: combination therapy and dual/multiple targeting chemotherapeutics. Two enzymes, histone deacetylases (HDACs) and ribonucleotide reductase (RR), are crucial for several biological functions, including replication and repair of DNA, division of cells, transcription of genes, etc. However, it has been noted that different cancers exhibit abnormal functions of these enzymes. Potent inhibitors for each of these proteins have been extensively researched. Many medications based on these inhibitors have been successfully food and drug administration (FDA) approved, and the majority are undergoing various stages of clinical testing. This review discusses various studies of HDAC and RR inhibitors in combination therapy and dual-targeting chemotherapeutics.
Topics: Humans; Ribonucleotide Reductases; Histone Deacetylases; Histone Deacetylase Inhibitors; Neoplasms
PubMed: 38520762
DOI: 10.1016/j.ejmech.2024.116324 -
Microbiology Spectrum Aug 2022A balance in the deoxyribonucleotide (dNTPs) intracellular concentration is critical for the DNA replication and repair processes. In the model yeast Saccharomyces...
A balance in the deoxyribonucleotide (dNTPs) intracellular concentration is critical for the DNA replication and repair processes. In the model yeast Saccharomyces cerevisiae, the Mec1-Rad53-Dun1 kinase cascade mainly regulates the ribonucleotide reductase (RNR) gene expression during DNA replication and DNA damage stress. However, the RNR regulatory mechanisms in basidiomycete fungi during DNA replication and damage stress remain elusive. Here, we observed that in C. neoformans (large RNR subunit) and (one small RNR subunit) were required for cell viability, but not (another small RNR subunit). overexpression compensated for the lethality of suppression. In contrast to the regulatory mechanisms of RNRs in S. cerevisiae, Rad53 and Chk1 kinases cooperatively or divergently controlled and expression under DNA damage and DNA replication stress. In particular, this study revealed that Chk1 mainly regulated expression during DNA replication stress, whereas Rad53, rather than Chk1, played a significant role in controlling the expression of during DNA damage stress. Furthermore, the expression of , not but and , was suppressed by the Ssn6-Tup1 complex during DNA replication stress. Notably, we observed that expression was mainly regulated by Mbs1, whereas expression was cooperatively controlled by Mbs1 and Bdr1 as downstream factors of Rad53 and Chk1 during DNA replication and damage stress. Collectively, the regulation of RNRs in C. neoformans has both evolutionarily conserved and divergent features in DNA replication and DNA damage stress, compared with other yeasts. Upon DNA replication or damage stresses, it is critical to provide proper levels of deoxynucleotide triphosphates (dNTPs) and activate DNA repair machinery. Ribonucleotide reductases (RNRs), which are composed of large and small subunits, are required for synthesizing dNTP. An imbalance in the intracellular concentration of dNTPs caused by the perturbation of RNR results in a reduction in DNA repair fidelity. Despite the importance of their roles, functions and regulations of RNR have not been elucidated in the basidiomycete fungi. In this study, we found that the roles of , , and genes encoding RNR subunits in the viability of C. neoformans. Furthermore, their expression levels are divergently regulated by the Rad53-Chk1 pathway and the Ssn6-Tup1 complex in response to DNA replication and damage stresses. Therefore, this study provides insight into the regulatory mechanisms of RNR genes to DNA replication and damage stresses in basidiomycete fungi.
Topics: Checkpoint Kinase 2; Cryptococcus neoformans; DNA Damage; DNA Replication; Ribonucleotide Reductases; Saccharomyces cerevisiae
PubMed: 35736239
DOI: 10.1128/spectrum.01044-22 -
Neoplasia (New York, N.Y.) Jul 2021Ribonucleotide reductase (RNR) is the key enzyme that catalyzes the production of deoxyribonucleotides (dNTPs) for DNA replication and it is also essential for cancer...
Ribonucleotide reductase (RNR) is the key enzyme that catalyzes the production of deoxyribonucleotides (dNTPs) for DNA replication and it is also essential for cancer cell proliferation. As the RNR inhibitor, Gemcitabine is widely used in cancer therapies, however, resistance limits its therapeutic efficacy and curative potential. Here, we identified that mTORC2 is a main driver of gemcitabine resistance in non-small cell lung cancers (NSCLC). Pharmacological or genetic inhibition of mTORC2 greatly enhanced gemcitabine induced cytotoxicity and DNA damage. Mechanistically, mTORC2 directly interacted and phosphorylated RNR large subunit RRM1 at Ser 631. Ser631 phosphorylation of RRM1 enhanced its interaction with small subunit RRM2 to maintain sufficient RNR enzymatic activity for efficient DNA replication. Targeting mTORC2 retarded DNA replication fork progression and improved therapeutic efficacy of gemcitabine in NSCLC xenograft model in vivo. Thus, these results identified a mechanism through mTORC2 regulating RNR activity and DNA replication, conferring gemcitabine resistance to cancer cells.
Topics: Antimetabolites, Antineoplastic; Carcinoma, Non-Small-Cell Lung; Cell Line, Tumor; DNA Damage; DNA Replication; Deoxycytidine; Drug Resistance, Neoplasm; Gene Expression Regulation, Neoplastic; Gene Knockdown Techniques; Histones; Humans; Mechanistic Target of Rapamycin Complex 2; Phosphorylation; Protein Binding; Ribonucleoside Diphosphate Reductase; Ribonucleotide Reductases; Signal Transduction; Gemcitabine
PubMed: 34126361
DOI: 10.1016/j.neo.2021.05.007