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1.
FEBS Lett ; 596(23): 3087-3102, 2022 Dec.
Article in English | MEDLINE | ID: mdl-36053795

ABSTRACT

Elucidating the mechanism of action of an antifungal or cytotoxic compound is a time-consuming process. Yeast chemogenomic profiling provides a compelling solution to the problem but is experimentally complex. Here, we demonstrate the use of a highly simplified yeast chemical genetic assay comprising just 89 yeast deletion strains, each diagnostic for a specific mechanism of action. We use the assay to investigate the mechanism of action of two antifungal chalcone compounds, trans-chalcone and 4'-hydroxychalcone, and narrow down the mechanism to transcriptional stress. Crucially, the assay eliminates mechanisms of action such as topoisomerase I inhibition and membrane disruption that have been suggested for related chalcone compounds. We propose this simplified assay as a useful tool to rapidly identify common off-target mechanisms.


Subject(s)
Chalcone , Chalcones , Chalcone/pharmacology , Saccharomyces cerevisiae/genetics , Antifungal Agents/pharmacology , Chalcones/pharmacology
2.
Aging (Albany NY) ; 11(23): 11686-11721, 2019 12 08.
Article in English | MEDLINE | ID: mdl-31812950

ABSTRACT

Topoisomerase II (Top2) removes topological linkages between replicated chromosomes. Top2 inhibition leads to mitotic catastrophe (MC) when cells unsuccessfully try to split their genetic material between the two daughter cells. Herein, we have characterized the fate of these daughter cells in the budding yeast. Clonogenic and microcolony experiments, in combination with vital and apoptotic stains, showed that 75% of daughter cells become senescent in the short term; they are unable to divide but remain alive. Decline in cell vitality then occurred, yet slowly, uncoordinatedly when comparing pairs of daughters, and independently of the cell death mediator Mca1/Yca1. Furthermore, we showed that senescence can be modulated by ploidy, suggesting that gross chromosome imbalances during segregation may account for this phenotype. Indeed, we found that diploid long-term survivors of the MC are prone to genomic imbalances such as trisomies, uniparental disomies and terminal loss of heterozygosity (LOH), the latter affecting the longest chromosome arms.


Subject(s)
DNA Topoisomerases, Type II/metabolism , Gene Expression Regulation, Enzymologic/physiology , Gene Expression Regulation, Fungal/physiology , Saccharomyces cerevisiae/enzymology , Cell Survival , DNA Topoisomerases, Type II/genetics , Mitosis , Mutation , Saccharomyces cerevisiae/genetics , Single-Cell Analysis
3.
ACS Chem Biol ; 13(8): 1950-1957, 2018 08 17.
Article in English | MEDLINE | ID: mdl-29878754

ABSTRACT

Naphthoquinones are among the most active natural products obtained from plants and microorganisms. Naphthoquinones exert their biological activities through pleiotropic mechanisms that include reactivity against cell nucleophiles, generation of reactive oxygen species (ROS), and inhibition of proteins. Here, we report a mechanistic antiproliferative study performed in the yeast Saccharomyces cerevisiae for several derivatives of three important natural naphthoquinones: lawsone, juglone, and ß-lapachone. We have found that (i) the free hydroxyl group of lawsone and juglone modulates toxicity; (ii) lawsone and juglone derivatives differ in their mechanisms of action, with ROS generation being more important for the former; and (iii) a subset of derivatives possess the capability to disrupt mitochondrial function, with ß-lapachones being the most potent compounds in this respect. In addition, we have cross-compared yeast results with antibacterial and antitumor activities. We discuss the relationship between the mechanistic findings, the antiproliferative activities, and the physicochemical properties of the naphthoquinones.


Subject(s)
Cell Proliferation/drug effects , Mitochondria/drug effects , Naphthoquinones/pharmacology , Anti-Bacterial Agents/chemistry , Anti-Bacterial Agents/pharmacology , Antifungal Agents/chemistry , Antifungal Agents/pharmacology , Antineoplastic Agents/chemistry , Antineoplastic Agents/pharmacology , Cell Line, Tumor , Humans , Molecular Structure , Naphthoquinones/chemistry , Oxidative Stress/drug effects , Saccharomyces cerevisiae/drug effects , Staphylococcus aureus/drug effects , Structure-Activity Relationship
4.
Bioorg Med Chem Lett ; 27(3): 484-489, 2017 02 01.
Article in English | MEDLINE | ID: mdl-28040393

ABSTRACT

Based on previous Topoisomerase II docking studies of naphthoquinone derivatives, a series of naphthoquinone-coumarin conjugates was synthesized through a multicomponent reaction from aromatic aldehydes, 4-hydroxycoumarin and 2-hydroxynaphthoquinone. The hybrid structures were evaluated against the α isoform of human topoisomerase II (hTopoIIα), Escherichia coli DNA Gyrase and E. coli Topoisomerase I. All tested compounds inhibited the hTopoIIα-mediated relaxation of negatively supercoiled circular DNA in the low micromolar range. This inhibition was specific since neither DNA Gyrase nor Topoisomerase I were affected. Cleavage assays pointed out that naphthoquinone-coumarins act by catalytically inhibiting hTopoIIα. ATPase assays and molecular docking studies further pointed out that the mode of action is related to the hTopoIIα ATP-binding site.


Subject(s)
Coumarins/pharmacology , DNA Topoisomerases, Type II/metabolism , Naphthoquinones/pharmacology , Topoisomerase II Inhibitors/pharmacology , Coumarins/chemistry , Dose-Response Relationship, Drug , Humans , Molecular Structure , Naphthoquinones/chemistry , Structure-Activity Relationship , Topoisomerase II Inhibitors/chemical synthesis , Topoisomerase II Inhibitors/chemistry
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