Periodic Reporting for period 1 - FungiFeS (Iron-sulphur cluster biogenesis: deciphering new molecular mechanisms and exploiting novel antifungal and fungicide targets)
Reporting period: 2021-09-01 to 2023-08-31
Fungal infections have major socioeconomic impacts. Recent estimates suggest that invasive human fungal infections kill more than 1.6 million people annually and that 600 million people could be fed each year by halting the spread of fungal diseases in the five most important crops. A range of antifungals, although relatively limited, are currently used to control fungal pathogens, but resistance to those is growing, underscoring the urgent need for new molecular targets and treatments.
Iron-sulphur (Fe-S) clusters are inorganic cofactors found in all kingdoms of life and are required for several essential cellular processes such as oxidative phosphorylation, DNA synthesis and repair, iron metabolism. Pathways involved in the biogenesis of Fe-S proteins were suggested to represent possible novel antifungal targets. The main objective of the project was to establish the essential mitochondrial and cytoplasmic NADPH-dependent electron transfer chains (Arh1-Yah1 and Tah18-Dre2, respectively) as novel antifungal targets by discovering compounds that can disrupt the interaction and therefore the function of Arh1-Yah1 and Tah18-Dre2. By screening the libraries of the Institut de Chimie des Substances Naturelles (ICSN, CNRS, Gif-sur-Yvette, France) against the yeast Saccharomyces cerevisiae, several natural compounds and extracts have been found to potentially inhibit the interaction between the mitochondrial ferredoxin Yah1 and its reductase Arh1. Additionally, synergy was found between these compounds and the antimalarial primaquine, a drug that, as we showed previously, targets Fe-S proteins such as the aconitase of the tricarboxylic acid cycle and Rli1, a protein essential for ribosome biogenesis and protein translation. Synergistic combinations allow to lower cost and potential toxicity by reducing doses needed to inhibit fungal growth. This strategy is particularly useful when the protein targeted is relatively well conserved between the organism pathogens and their hosts, like here. The paucity of current antifungal treatments can actually be partly explained by the common eukaryotic nature of fungal pathogens and their hosts. Therefore, finding fungal-specific agents is challenging and synergistic combinations of agents can be a way to minimize this challenge. Synergistic effects are seen where agents target a common process but by different mechanisms or pathways. Therefore, targeting for instance the ferredoxin Yah1 and a protein of its protein network could be a promising strategy to inhibit fungal pathogens. Unfortunately, we have currently very limited information regarding the protein network of the ferredoxin. Another objective of this project was therefore to develop a range of tools to establish this network and identified novel antifungal targets within.
The data collectively support the mitochondrial NADPH-dependent electron transfer chain as a potential antifungal target and highlight promising drug candidates.
Iron-sulphur (Fe-S) clusters are inorganic cofactors found in all kingdoms of life and are required for several essential cellular processes such as oxidative phosphorylation, DNA synthesis and repair, iron metabolism. Pathways involved in the biogenesis of Fe-S proteins were suggested to represent possible novel antifungal targets. The main objective of the project was to establish the essential mitochondrial and cytoplasmic NADPH-dependent electron transfer chains (Arh1-Yah1 and Tah18-Dre2, respectively) as novel antifungal targets by discovering compounds that can disrupt the interaction and therefore the function of Arh1-Yah1 and Tah18-Dre2. By screening the libraries of the Institut de Chimie des Substances Naturelles (ICSN, CNRS, Gif-sur-Yvette, France) against the yeast Saccharomyces cerevisiae, several natural compounds and extracts have been found to potentially inhibit the interaction between the mitochondrial ferredoxin Yah1 and its reductase Arh1. Additionally, synergy was found between these compounds and the antimalarial primaquine, a drug that, as we showed previously, targets Fe-S proteins such as the aconitase of the tricarboxylic acid cycle and Rli1, a protein essential for ribosome biogenesis and protein translation. Synergistic combinations allow to lower cost and potential toxicity by reducing doses needed to inhibit fungal growth. This strategy is particularly useful when the protein targeted is relatively well conserved between the organism pathogens and their hosts, like here. The paucity of current antifungal treatments can actually be partly explained by the common eukaryotic nature of fungal pathogens and their hosts. Therefore, finding fungal-specific agents is challenging and synergistic combinations of agents can be a way to minimize this challenge. Synergistic effects are seen where agents target a common process but by different mechanisms or pathways. Therefore, targeting for instance the ferredoxin Yah1 and a protein of its protein network could be a promising strategy to inhibit fungal pathogens. Unfortunately, we have currently very limited information regarding the protein network of the ferredoxin. Another objective of this project was therefore to develop a range of tools to establish this network and identified novel antifungal targets within.
The data collectively support the mitochondrial NADPH-dependent electron transfer chain as a potential antifungal target and highlight promising drug candidates.
The yeast Saccharomyces cerevisiae is a powerful model organism to study fundamental cellular processes and has been extensively used to identify the mode of action of antifungals and other chemical compounds.
Here we engineered the yeast in order to identify rapidly compounds among chemical libraries that can target specifically the mitochondrial or cytoplasmic NADPH-dependent electron transfer chains, both essential for the biogenesis of Fe-S proteins. Once the strains obtained, we screened a selection of compounds from the ICSN chemical library and hundreds of extracts from the ICSN strain extract library. The collection contains no commercially available compounds or compounds from commercial chemical libraries enabling the identification of novel agents. Three compounds and one extract were found to potentially inhibit the interaction between Arh1 and Yah1. A 3D model of the Arh1-Yah1 complex was generated using AlphaFold and docking experiment to establish the molecular mechanism of action of the three identified agents.
These compounds inhibited the growth of the yeast synergistically with the antimalarial primaquine known to target several Fe-S proteins. To discover further potential targets for combinatorial treatments with our compounds of interest, we developed tools (proximity-dependent biotin identification, Föster resonance energy transfer, functional genomics screening) to establish the protein network of the ferredoxin for which we have limited information.
Part of this work has been presented during the ICSN 602+ symposium and the Iron-Sulphur Proteins: Biogenesis, Regulation and Functions conference held in in 2022 in Gif-sur-Yvette (France) and Saint-Tropez (France) respectively.
Here we engineered the yeast in order to identify rapidly compounds among chemical libraries that can target specifically the mitochondrial or cytoplasmic NADPH-dependent electron transfer chains, both essential for the biogenesis of Fe-S proteins. Once the strains obtained, we screened a selection of compounds from the ICSN chemical library and hundreds of extracts from the ICSN strain extract library. The collection contains no commercially available compounds or compounds from commercial chemical libraries enabling the identification of novel agents. Three compounds and one extract were found to potentially inhibit the interaction between Arh1 and Yah1. A 3D model of the Arh1-Yah1 complex was generated using AlphaFold and docking experiment to establish the molecular mechanism of action of the three identified agents.
These compounds inhibited the growth of the yeast synergistically with the antimalarial primaquine known to target several Fe-S proteins. To discover further potential targets for combinatorial treatments with our compounds of interest, we developed tools (proximity-dependent biotin identification, Föster resonance energy transfer, functional genomics screening) to establish the protein network of the ferredoxin for which we have limited information.
Part of this work has been presented during the ICSN 602+ symposium and the Iron-Sulphur Proteins: Biogenesis, Regulation and Functions conference held in in 2022 in Gif-sur-Yvette (France) and Saint-Tropez (France) respectively.
In addition to finding novel strategies against fungal pathogens that have a major socio-economic impact, this project provided tools that could advance our current knowledge on the mitochondrial ferredoxin as our comprehension of this essential Fe-S protein is still partial.
Our data and tools will be important in other aspects of human health as humans possess two ferredoxins, FDX1 and FDX2 that were respectively found recently to be essential for the copper-dependent regulated cell death cuproptosis that is suggested to play a role in tumour suppression, and to associated to mitochondrial myopathy and neurological disorders.
Our data and tools will be important in other aspects of human health as humans possess two ferredoxins, FDX1 and FDX2 that were respectively found recently to be essential for the copper-dependent regulated cell death cuproptosis that is suggested to play a role in tumour suppression, and to associated to mitochondrial myopathy and neurological disorders.