The galactose-induced decrease in phosphate levels leads to toxicity in yeast models of galactosemia

Machado, Caio M.; De-Souza, Evandro A.; De-Queiroz, Ana Luiza F V; Pimentel, Felipe S. A.; Silva, Guilherme F S; Gomes, Fabio M.; Montero-Lomelı́, Mónica; Masuda, Cláudio A.

doi:10.1016/j.bbadis.2017.02.014

Cited by 15 publications

(12 citation statements)

References 44 publications

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“…We considered whether accumulation of toxic metabolites might account for cell wall biosynthesis defects. However, toxic metabolites that accumulate, e.g., in S. cerevisiae models of galactosemia (64,65) and fructose intolerance (65), are sugar phosphates, i.e., their biosynthesis requires a phosphorylation step whose substrate was scarce in cells lacking Pho84. Accordingly, we did not identify potentially toxic metabolites among the significantly dysregulated metabolites in these cells (Table S1).…”

Section: Discussionmentioning

confidence: 99%

Phosphoric Metabolites Link Phosphate Import and Polysaccharide Biosynthesis for Candida albicans Cell Wall Maintenance

Liu

Acosta-Zaldívar

et al. 2020

mBio

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The Candida albicans high-affinity phosphate transporter Pho84 is required for normal Target of Rapamycin (TOR) signaling, oxidative stress resistance, and virulence of this fungal pathogen. It also contributes to C. albicans’ tolerance of two antifungal drug classes, polyenes and echinocandins. Echinocandins inhibit biosynthesis of a major cell wall component, beta-1,3-glucan. Cells lacking Pho84 were hypersensitive to other forms of cell wall stress beyond echinocandin exposure, while their cell wall integrity signaling response was weak. Metabolomics experiments showed that levels of phosphoric intermediates, including nucleotides like ATP and nucleotide sugars, were low in pho84 mutant compared to wild-type cells recovering from phosphate starvation. Nonphosphoric precursors like nucleobases and nucleosides were elevated. Outer cell wall phosphomannan biosynthesis requires a nucleotide sugar, GDP-mannose. The nucleotide sugar UDP-glucose is the substrate of enzymes that synthesize two major structural cell wall polysaccharides, beta-1,3- and beta-1,6-glucan. Another nucleotide sugar, UDP-N-acetylglucosamine, is the substrate of chitin synthases which produce a stabilizing component of the intercellular septum and of lateral cell walls. Lack of Pho84 activity, and phosphate starvation, potentiated pharmacological or genetic perturbation of these enzymes. We posit that low substrate concentrations of beta-d-glucan- and chitin synthases, together with pharmacologic inhibition of their activity, diminish enzymatic reaction rates as well as the yield of their cell wall-stabilizing products. Phosphate import is not conserved between fungal and human cells, and humans do not synthesize beta-d-glucans or chitin. Hence, inhibiting these processes simultaneously could yield potent antifungal effects with low toxicity to humans. IMPORTANCE Candida species cause hundreds of thousands of invasive infections with high mortality each year. Developing novel antifungal agents is challenging due to the many similarities between fungal and human cells. Maintaining phosphate balance is essential for all organisms but is achieved completely differently by fungi and humans. A protein that imports phosphate into fungal cells, Pho84, is not present in humans and is required for normal cell wall stress resistance and cell wall integrity signaling in C. albicans. Nucleotide sugars, which are phosphate-containing building block molecules for construction of the cell wall, are diminished in cells lacking Pho84. Cell wall-constructing enzymes may be slowed by lack of these building blocks, in addition to being inhibited by drugs. Combined targeting of Pho84 and cell wall-constructing enzymes may provide a strategy for antifungal therapy by which two sequential steps of cell wall maintenance are blocked for greater potency.

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Section: Discussionmentioning

confidence: 99%

Phosphoric Metabolites Link Phosphate Import and Polysaccharide Biosynthesis for Candida albicans Cell Wall Maintenance

Liu

Acosta-Zaldívar

et al. 2020

mBio

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“…Though the null mutant could grow in glucose, its growth rate was significantly reduced ( Figure 5). The gal1D/D mutant was able to grow in the medium with 1% glucose and 1% galactose with a much lower rate and a half efficiency as compared with the wild-type 11-3002, suggesting that galactose was probably unable to enter into the cells of the mutant without GAL1; otherwise, the unmetabolized galactose would be lethal to the yeast cells [34][35][36]. HPLC analysis showed that the concentration of galactose remained unchanged during the growth course of the 11-3002gal1D/D mutant in the medium with 1% glucose and 1% galactose, though the cells grew well, and glucose was exhausted ( Figure S4A), confirming that galactose was unable to be transferred into the cells of the mutant.…”

Section: The Function Of Gal2 Requires Other Structural Gal Genesmentioning

confidence: 99%

Reverse Evolution of a Classic Gene Network in Yeast Offers a Competitive Advantage

et al. 2019

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“…This cyclical phosphorylation and dephosphorylation of galactose could deplete cellular ATP levels, leading to cellular toxicity. 19,20 Machado et al 21 have suggested that cellular toxicity may be due to trapping of phosphate in Gal-1P, leading to intracellular inorganic phosphate depletion. Using a yeast model of Classic Galactosaemia, the authors showed that supplementation of phosphate suppressed the toxic effects of galactose in yeast cells.…”

Section: Cellular Toxicity Of Gal-1pmentioning

confidence: 99%

Galactose 1‐phosphate accumulates to high levels in galactose‐treated cells due to low GALT activity and absence of product inhibition of GALK

Cheng

Zhou

et al. 2019

J of Inher Metab Disea

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Classic Galactosaemia is a genetic disorder, characterised by galactose intolerance in newborns. It occurs due to recessive mutations in the galactose-1-phosphate uridylyltransferase (GALT) gene. One of the main alterations caused by GALT deficiency is the accumulation of galactose 1-phosphate (Gal-1P) in cells. Studies have suggested that Gal-1P exerts cellular toxicity, possibly by inhibiting cellular metabolism. However, the exact significance of Gal-1P in disease pathogenesis remains unclear. In this study, we tested the hypothesis that Gal-1P inhibits cellular glucose utilisation by competing with substrates in the glycolytic pathway. We also investigated the metabolism of both galactose and glucose in GALT-expressing HEK293T and 143B cells to identify critical reactions steps contributing to the metabolic toxicity of galactose. Notably, we found that galactose-treated HEK293T and 143B cells, which express endogenous GALT, accumulate markedly high intracellular Gal-1P concentrations. Despite very high intracellular Gal-1P concentrations, no inhibition of cellular glucose uptake and no significant changes in the intracellular concentrations of glycolytic metabolites were observed. This indicates that Gal-1P does not exert an inhibitory effect on glycolysis in cells and rules out one potential hypothesis for cellular Gal-1P toxicity. We also investigated the mechanism responsible for the observed Gal-1P accumulation. Our results suggest that Gal-1P accumulation is a result of both low GALT activity and the absence of product inhibition by Gal-1P on galactokinase (GALK1), the enzyme responsible for phosphorylating galactose to Gal-1P. These findings provide a better understanding of the disease mechanisms underlying Classic Galactoaemia. K E Y W O R D SClassic Galactosaemia, galactose metabolism, GALT deficiency, Leloir pathway, metabolomics

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The galactose-induced decrease in phosphate levels leads to toxicity in yeast models of galactosemia

Cited by 15 publications

References 44 publications

Phosphoric Metabolites Link Phosphate Import and Polysaccharide Biosynthesis for Candida albicans Cell Wall Maintenance

Phosphoric Metabolites Link Phosphate Import and Polysaccharide Biosynthesis for Candida albicans Cell Wall Maintenance

Reverse Evolution of a Classic Gene Network in Yeast Offers a Competitive Advantage

Galactose 1‐phosphate accumulates to high levels in galactose‐treated cells due to low GALT activity and absence of product inhibition of GALK

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