The metabolic growth limitations of petite cells lacking the mitochondrial genome

Vowinckel, Jakob; Hartl, Johannes; Marx, Hans; Kerick, Martin; Runggatscher, Kathrin; Keller, Markus A.; Mülleder, Michael; Day, Jason; Weber, Manuela; Rinnerthaler, Mark; Yu, Jigui; Aulakh, Simran Kaur; Lehmann, Andrea; Mattanovich, Diethard; Timmermann, Bernd; Zhang, Nianshu; Dunn, Cory D.; MacRae, James I.; Breitenbach, Michael; Ralser, Markus

doi:10.1038/s42255-021-00477-6

Cited by 31 publications

(38 citation statements)

References 116 publications

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“…Mitochondrial membrane depolarization by both EUG and BER, individually, has been reported previously ( da Silva et al, 2016 ; Alves et al, 2017 ). Furthermore, polarity is partly retained in the absence of mtDNA ( Appleby et al, 1999 ; Vowinckel et al, 2021 ), consistent with the partial-retention of synergy in the rho 0 mutant shown above. Therefore, we probed mitochondrial membrane depolarization (MMD), using flow cytometric analysis of decreased retention of rhodamine 123 fluorescence by cells, at doses of EUG + BER combinations giving approximately 10, 25, and 50% growth inhibition ( Figure 5A ).…”

Section: Resultssupporting

confidence: 82%

Discovery of Natural Products With Antifungal Potential Through Combinatorial Synergy

Augostine

Avery

2022

Front. Microbiol.

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The growing prevalence of antifungal drug resistance coupled with the slow development of new, acceptable drugs and fungicides has raised interest in natural products (NPs) for their therapeutic potential and level of acceptability. However, a number of well-studied NPs are considered promiscuous molecules. In this study, the advantages of drug–drug synergy were exploited for the discovery of pairwise NP combinations with potentiated antifungal activity and, potentially, increased target specificity. A rational approach informed by previously known mechanisms of action of selected NPs did not yield novel antifungal synergies. In contrast, a high-throughput screening approach with yeast revealed 34 potential synergies from 800 combinations of a diverse NP library with four selected NPs of interest (eugenol, EUG; β-escin, ESC; curcumin, CUR; berberine hydrochloride, BER). Dedicated assays validated the most promising synergies, namely, EUG + BER, CUR + sclareol, and BER + pterostilbene (PTE) [fractional inhibitory concentrations (FIC) indices ≤ 0.5 in all cases], reduced to as low as 35 (BER) and 7.9 mg L–1 (PTE). These three combinations synergistically inhibited a range of fungi, including human or crop pathogens Candida albicans, Aspergillus fumigatus, Zymoseptoria tritici, and Botrytis cinerea, with synergy also against azole-resistant isolates and biofilms. Further investigation indicated roles for mitochondrial membrane depolarization and reactive oxygen species (ROS) formation in the synergistic mechanism of EUG + BER action. This study establishes proof-of-principle for utilizing high-throughput screening of pairwise NP interactions as a tool to find novel antifungal synergies. Such NP synergies, with the potential also for improved specificity, may help in the management of fungal pathogens.

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Section: Resultssupporting

confidence: 82%

Discovery of Natural Products With Antifungal Potential Through Combinatorial Synergy

Augostine

Avery

2022

Front. Microbiol.

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“… C. albicans is a so-called “petite-negative” yeast, i.e., it does not tolerate loss of mitochondrial DNA and respiration. Candida glabrata and Saccharomyces cerevisiae are “petite-positive,” meaning they can grow in the absence of mitochondrial DNA and respiration ( 44 , 45 ). Due to poor growth of S. cerevisiae in RPMI and at 37°C, this experiment was carried out in yeast nitrogen base (YNB) and at 30°C.…”

Section: Resultsmentioning

confidence: 99%

Disruption of Iron Homeostasis and Mitochondrial Metabolism Are Promising Targets to Inhibit Candida auris

et al. 2022

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Over the last decade, Candida auris has emerged as a human pathogen around the world causing life-threatening infections with wide-spread antifungal drug resistance, including pandrug resistance in some cases. In this study, we addressed the mechanism of action of the antiparasitic drug pyrvinium pamoate against C. auris and show how metabolism could be inhibited to curb C. auris proliferation.

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“…However, the ability of the H 2 S chelator Fe-EDTA to restore the growth of petite FY4- met15Δ cells grown in liquid cultures ( Figure 4D ), as well as the ability of these cells to grow on solid SD-Met+Glu medium ( Figure 4E ), strongly suggests that H 2 S is limiting the growth of met15Δ cells in part through a pathway that does not involve the mitochondrial genome. The precise molecular mechanisms at play may be dissected in future genome-scale investigations of suppressor and synthetic lethal mutations, similar to how a recent study dissected the mechanisms underlying the growth defect associated with petites (Vowinckel et al, 2021).…”

Section: Discussionmentioning

confidence: 95%

On the illusion of auxotrophy:met15Δyeast cells can grow on inorganic sulfur thanks to the previously uncharacterized homocysteine synthase Yll058w

Oss

Parikh

Coelho

et al. 2022

Preprint

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In budding yeast, the Met15 enzyme has long been assumed to be the sole homocysteine synthase, facilitating de novo synthesis of sulfur-containing organic compounds (organosulfurs) from inorganic precursors. Here we show that an alternative homocysteine synthase encoded by the previously uncharacterized gene YLL058W supports growth of mutants lacking MET15 in the absence of exogenous organosulfurs. This growth is observed specifically when cells are deposited in an automated fashion to seed colonies, but not with traditional cell propagation techniques such as thick patches of cells or liquid cultures. We show that the lack of growth in these contexts, which has historically justified the status of MET15 as a classic auxotrophic marker, is largely due to toxic levels of hydrogen sulfide accumulation rather than an inability to perform de novo homocysteine biosynthesis. These data have broad implications for investigations of sulfur starvation/metabolism, including studies of aging and emerging cancer therapeutics.

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The metabolic growth limitations of petite cells lacking the mitochondrial genome

Cited by 31 publications

References 116 publications

Discovery of Natural Products With Antifungal Potential Through Combinatorial Synergy

Discovery of Natural Products With Antifungal Potential Through Combinatorial Synergy

Disruption of Iron Homeostasis and Mitochondrial Metabolism Are Promising Targets to Inhibit Candida auris

On the illusion of auxotrophy:met15Δyeast cells can grow on inorganic sulfur thanks to the previously uncharacterized homocysteine synthase Yll058w

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