Identification of eukaryotic UDP-galactopyranose mutase inhibitors using the ThermoFAD assay

Campo, Julia S. Martín del; Eckshtain-Levi, Meital; Sobrado, Pablo

doi:10.1016/j.bbrc.2017.09.074

Cited by 6 publications

(9 citation statements)

References 34 publications

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“…Galactomannan, the production of which is initiated by UGM, is a ubiquitous component in fungal cell walls. The absence of UGM leads to the thinner fungal cell wall and reduced fungal virulence. , In our study, A. fumigatus was cultivated with different concentrations of flavopiridol for 1, 2, 3, 4, and 5 days.…”

Section: Discussionmentioning

confidence: 99%

“…The absence of UGM leads to the thinner fungal cell wall and reduced fungal virulence. 20,37 In our study, A. fumigatus was cultivated with different concentrations of flavopiridol for 1, 2, 3, 4, and 5 days. The absorbance of the medium was reduced after flavopiridol treatment.…”

Section: ■ Discussionmentioning

confidence: 99%

“…4,9,26 In addition, flavopiridol has been considered an inhibitor of UGM, which has antifungal potential. 20 Thus, we hypothesized that flavopiridol might protect against fungal keratitis.…”

Section: ■ Discussionmentioning

confidence: 99%

“…Moreover, flavopiridol has been considered an inhibitor of UDP-galactopyranose mutase (UGM) in A. fumigatus, suggesting its potential antifungal effect. , UGM is a crucial enzyme for fungal cell wall biogenesis, which promotes the transformation from UDP-galactofuranose (Galf) to UDP-galactopyranose (Galp) . The fungal cell wall is involved in the interaction between pathogens and hosts, which determines the pathogenicity of fungal keratitis .…”

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confidence: 99%

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Flavopiridol Protects against Fungal Keratitis due to Aspergillus fumigatus by Alleviating Inflammation through the Promotion of Autophagy

Peng

et al. 2022

ACS Infect. Dis.

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Fungal keratitis is a serious infectious keratopathy related to fungal virulence and excessive inflammatory responses. Autophagy exhibits a potent ability to resolve inflammation during fungal infection. This study aimed to investigate the protective function of flavopiridol in Aspergillus fumigatus keratitis and explore its effects on autophagy. In our study, the corneas of the fungal keratitis mouse model were treated with 5 μM flavopiridol. In vitro, RAW 264.7 cells were pretreated with 200 nM flavopiridol before fungal stimulation. A. fumigatus was incubated with flavopiridol, and the antifungal activity of flavopiridol was detected. Our results indicated that flavopiridol treatment notably reduced clinical scores as well as cytokines expression of infected corneas. In infected RAW 264.7 cells, flavopiridol treatment inhibited IL-1β, IL-6, and TNF-α expression but promoted IL-10 expression. Transmission electron microscopy (TEM) images showed that more autolysosomes were present in infected corneas and RAW 264.7 cells after flavopiridol treatment. Flavopiridol treatment notably upregulated the protein expression of LC3, Beclin-1, and Atg-7. 3-Methyladenine (3-MA, an inhibitor of autophagy) pretreatment counteracted the cytokine regulation induced by flavopiridol. Moreover, flavopiridol promoted the phagocytosis of RAW 264.7 cells. Flavopiridol also exhibited antifungal activity by restricting fungal growth and limiting fungal biofilm formation and conidial adhesion. In conclusion, flavopiridol significantly alleviated the inflammation of fungal keratitis by activating autophagy. In addition, flavopiridol promoted the phagocytosis of RAW 264.7 cells and exhibited antifungal function, indicating the potential therapeutic role of flavopiridol in fungal keratitis.

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Section: Discussionmentioning

confidence: 99%

Section: ■ Discussionmentioning

confidence: 99%

Section: ■ Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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Flavopiridol Protects against Fungal Keratitis due to Aspergillus fumigatus by Alleviating Inflammation through the Promotion of Autophagy

Peng

et al. 2022

ACS Infect. Dis.

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“…Moreover, avopiridol has been considered as an inhibitor of UDP-galactopyranose mutase (UGM) in A. fumigatus, suggesting its potential antifungal effect (Martín et al 2017, Hostetter et al 1994. UGM is a crucial enzyme for fungal cell wall biogenesis, which promotes the transformation from UDPgalactofuranose (Galf) to UDP-galactopyranose (Galp) (Latgé 2010).…”

Section: Introductionmentioning

confidence: 99%

Flavopiridol Protects Against Fungal Keratitis by Alleviating Inflammation Through the Promotion of Autophagy

Cui

Peng

et al. 2022

Preprint

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Background: Fungal keratitis is a serious infectious keratopathy related to fungal virulence and excessive inflammatory responses. Autophagy exhibits a potent ability to resolve inflammation during fungal infection. This study aimed to investigate the protective function of flavopiridol in fungal keratitis and explore its effects on autophagy.Methods: A mouse model of fungal keratitis was established and then treated with 5 μM flavopiridol. RAW 264.7 cells were treated with 200 nM flavopiridol before fungal stimulation. The severity of corneal diseases was evaluated by slit-lamp microscopy. The expression levels of cytokines were detected by RT-PCR and ELISA. The protein levels of LC3, Beclin-1 and Atg7 were determined by western blot and immunofluorescence. A Cell Counting Kit-8 assay was used to test cell viability. Autolysosomes were detected by transmission electron microscopy (TEM). An inhibitor of autophagy, 3-methyladenine (3-MA), was used to pretreat RAW 264.7 cells. Phagocytosis of RAW 264.7 cells was evaluated by counting colony forming units. A. fumigatus was incubated with flavopiridol, and the hyphae were stained with calcofluor white. Absorbance assay, crystal violet staining and adherence assay were used to detect the antifungal activity of flavopiridol.Results: Flavopiridol treatment notably reduced corneal opacity and the clinical scores of infected corneas. Compared with DMSO treatment, flavopiridol treatment greatly downregulated IL-1β, IL-6 and TNF-a expression in infected corneas. In RAW 264.7 cells, flavopiridol treatment inhibited IL-1β, IL-6 and TNF-a expression but promoted IL-10 expression. TEM images showed that more autolysosomes were presented in infected corneas and RAW 264.7 cells after flavopiridol treatment than after DMSO treatment. Flavopiridol treatment notably upregulated the protein expression of LC3, Beclin-1 and Atg7 in infected corneas as well as in RAW 264.7 cells. 3-MA pretreatment counteracted the cytokine regulation induced by flavopiridol. Moreover, flavopiridol promoted the phagocytosis of RAW 264.7 cells. Flavopiridol also exhibited antifungal activity by restricting fungal growth and limiting fungal biofilm formation and conidial adhesion. Conclusions: Flavopiridol significantly alleviated the inflammation of fungal keratitis by activating autophagy. In addition, flavopiridol promoted the phagocytosis of RAW 264.7 cells and exhibited antifungal function, indicating the potential therapeutic role of flavopiridol in fungal keratitis.

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Sirtuin E deacetylase is required for full virulence of Aspergillus fumigatus

Wassano,

da Silva,

Reis

et al. 2024

Commun Biol

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Aspergillus fumigatus represents a public health problem due to the high mortality rate in immunosuppressed patients and the emergence of antifungal-resistant isolates. Protein acetylation is a crucial post-translational modification that controls gene expression and biological processes. The strategic manipulation of enzymes involved in protein acetylation has emerged as a promising therapeutic approach for addressing fungal infections. Sirtuins, NAD+-dependent lysine deacetylases, regulate protein acetylation and gene expression in eukaryotes. However, their role in the human pathogenic fungus A. fumigatus remains unclear. This study constructs six single knockout strains of A. fumigatus and a strain lacking all predicted sirtuins (SIRTKO). The mutant strains are viable under laboratory conditions, indicating that sirtuins are not essential genes. Phenotypic assays suggest sirtuins’ involvement in cell wall integrity, secondary metabolite production, thermotolerance, and virulence. Deletion of sirE attenuates virulence in murine and Galleria mellonella infection models. The absence of SirE alters the acetylation status of proteins, including histones and non-histones, and triggers significant changes in the expression of genes associated with secondary metabolism, cell wall biosynthesis, and virulence factors. These findings encourage testing sirtuin inhibitors as potential therapeutic strategies to combat A. fumigatus infections or in combination therapy with available antifungals.

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Identification of eukaryotic UDP-galactopyranose mutase inhibitors using the ThermoFAD assay

Cited by 6 publications

References 34 publications

Flavopiridol Protects against Fungal Keratitis due to Aspergillus fumigatus by Alleviating Inflammation through the Promotion of Autophagy

Flavopiridol Protects against Fungal Keratitis due to Aspergillus fumigatus by Alleviating Inflammation through the Promotion of Autophagy

Flavopiridol Protects Against Fungal Keratitis by Alleviating Inflammation Through the Promotion of Autophagy

Sirtuin E deacetylase is required for full virulence of Aspergillus fumigatus

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