The secreted acid trehalase encoded by the CgATH1 gene is involved in Candida glabrata virulence.

BACKGROUND: Candida glabrata yeast is the second cause of candidiasis worldwide. Differs from other yeasts since assimilates only glucose and trehalose (a characteristic used in rapid identification tests for this pathogen) by secreting into the medium a highly active acid trehalase encoded by the CgATH1 gene. OBJECTIVE: This study aimed to characterise the function of the acid trehalase in the physiopathology of C. glabrata. METHODS: Gene deletion was performed to obtain a mutant ath1Δ strain, and the ability of the ath1Δ strain to grow in trehalase, or the presence of trehalase activity in the ath1Δ yeast cells, was verified. We also tested the virulence of the ath1Δ strain in a murine model of infection. FINDINGS: The ath1Δ mutant strain grows normally in the presence of glucose, but loses its ability to grow in trehalose. Due to the high acid trehalase activity present in wild-type cells, the cytoplasmic neutral trehalase activity is only detected in the ath1Δ strain. We also observed a significantly lower virulence of the ath1Δ strain in a murine model of infection with either normal or immunocompromised mice. MAIN CONCLUSIONS: The acid trehalase is involved in the hydrolysis of external trehalose by C. glabrata, and the enzyme also plays a major virulence role during infectivity.

The secreted acid trehalase encoded by the CgATH1 gene is involved in Candida glabrata virulence

BACKGROUND Candida glabrata yeast is the second cause of candidiasis worldwide. Differs from other yeasts since assimilates only glucose and trehalose (a characteristic used in rapid identification tests for this pathogen) by secreting into the medium a highly active acid trehalase encoded by the CgATH1 gene. OBJECTIVE This study aimed to characterise the function of the acid trehalase in the physiopathology of C. glabrata. METHODS Gene deletion was performed to obtain a mutant ath1Δ strain, and the ability of the ath1Δ strain to grow in trehalase, or the presence of trehalase activity in the ath1Δ yeast cells, was verified. We also tested the virulence of the ath1Δ strain in a murine model of infection. FINDINGS The ath1Δ mutant strain grows normally in the presence of glucose, but loses its ability to grow in trehalose. Due to the high acid trehalase activity present in wild-type cells, the cytoplasmic neutral trehalase activity is only detected in the ath1Δ strain. We also observed a significantly lower virulence of the ath1Δ strain in a murine model of infection with either normal or immunocompromised mice. MAIN CONCLUSIONS The acid trehalase is involved in the hydrolysis of external trehalose by C. glabrata, and the enzyme also plays a major virulence role during infectivity.

Insect trehalase: physiological significance and potential applications.

Trehalose, a non-reducing disaccharide, is widespread throughout the biological world. It is the major blood sugar in insects playing a crucial role as an instant source of energy and in dealing with abiotic stresses. The hydrolysis of trehalose is under the enzymatic control of trehalase. The enzyme trehalase is gaining interest in insect physiology as it regulates energy metabolism and glucose generation via trehalose catabolism. The two forms of insect trehalase namely, Tre-1 and Tre-2, are important in energy supply, growth, metamorphosis, stress recovery, chitin synthesis and insect flight. Insect trehalase has not been reviewed in depth and the information available is quite scattered. The present mini review discusses our recent understanding of the regulation, mechanism and biochemical characterization of insect trehalase with respect to its physiological role in vital life functions. We also highlight the molecular and biochemical properties of insect trehalase that makes it amenable to competitive inhibition by most glycosidase inhibitors. Due to its crucial role in carbon metabolism in insects, application of inhibitors against trehalose can form a promising area towards formulating strategies for insect pest control.

Disruption of the Candida albicans ATC1 gene encoding a cell-linked acid trehalase decreases hypha formation and infectivity without affecting resistance to oxidative stress.

In Candida albicans, the ATC1 gene, encoding a cell wall-associated acid trehalase, has been considered as a potentially interesting target in the search for new antifungal compounds. A phenotypic characterization of the double disruptant atc1Delta/atc1Delta mutant showed that it was unable to grow on exogenous trehalose as sole carbon source. Unlike actively growing cells from the parental strain (CAI4), the atc1Delta null mutant displayed higher resistance to environmental insults, such as heat shock (42 degrees C) or saline exposure (0.5 M NaCl), and to both mild and severe oxidative stress (5 and 50 mM H(2)O(2)), which are relevant during in vivo infections. Parallel measurements of intracellular trehalose and trehalose-metabolizing enzymes revealed that significant amounts of the disaccharide were stored in response to thermal and oxidative challenge in the two cell types. The antioxidant activities of catalase and glutathione reductase were triggered by moderate oxidative exposure (5 mM H(2)O(2)), whereas superoxide dismutase was inhibited dramatically by H(2)O(2), where a more marked decrease was observed in atc1Delta cells. In turn, the atc1Delta mutant exhibited a decreased capacity of hypha and pseudohypha formation tested in different media. Finally, the homozygous null mutant in a mouse model of systemic candidiasis displayed strongly reduced pathogenicity compared with parental or heterozygous strains. These results suggest not only a novel role for the ATC1 gene in dimorphism and infectivity, but also that an interconnection between stress resistance, dimorphic conversion and virulence in C. albicans may be reconsidered. They also support the hypothesis that Atc1p is not involved in the physiological hydrolysis of endogenous trehalose.

The ATC1 gene encodes a cell wall-linked acid trehalase required for growth on trehalose in Candida albicans.

After screening a Candida albicans genome data base, the product of an open reading frame (IPF 19760/CA2574) with 41% identity to Saccharomyces cerevisiae vacuolar acid trehalase (Ath1p) was identified and named Atc1p. The deduced amino acid sequence shows that Atc1p contains an N-terminal hydrophobic signal peptide and 20 potential sites for N-glycosylation. C. albicans homozygous mutants that lack acid trehalase activity were constructed by gene disruption at the two ATC chromosomal alleles. Analysis of these null mutants shows that Atc1p is localized in the cell wall and is required for growth on trehalose as a carbon source. An Atc1p endowed with acid trehalase activity was obtained by an in vtro transcription-translation coupled system. These results strongly suggest that ATC1 is the structural gene encoding cell wall acid trehalase in C. albicans. Determinations of ATC1 mRNA expression as well as acid trehalase activity in the presence and absence of glucose point out that ATC1 gene is regulated by glucose repression.

Response to oxidative stress caused by H(2)O(2) in Saccharomyces cerevisiae mutants deficient in trehalase genes.

The role of trehalose as cell protector against oxidative stress induced by H(2)O(2) has been studied in Saccharomyces cerevisiae mutants in which the two trehalase genes ATH1 and NTH1 are deleted. The addition of low H(2)O(2) concentrations to proliferating cultures of either strain did not harm cell viability and induced a marked activity to Nth1p, but with no significant level of trehalose accumulation. This pattern was reversed after a more severe H(2)O(2) treatment that caused drastic cell killing. The most severe phenotype corresponded to the Delta nth1 mutant. Under these conditions, the increase in Nth1p was abolished and a three-fold rise in trehalose content was recorded concomitant with activation of the trehalose synthase complex. The behavior of the double-disruptant Delta ath1Delta nth1 mutant was identical to that of wild-type cells, although in exponential cultures Ath1p activity was virtually undetectable upon exposure to H(2)O(2). Furthermore, these strains displayed an adaptive response to oxidative stress that was independent of intracellular trehalose synthesis. Our data strongly suggest that trehalose storage in budding yeasts is not an essential protectant in cell defense against oxidative challenge.

The effect of metacestodes of Hymenolepis diminuta on the bean-shaped accessory glands in male Tenebrio molitor.

Metacestodes of Hymenolepis diminuta affect several aspects of female reproductive physiology in Tenebrio molitor and such effects are mediated via the endocrine system. The effects on male reproduction are less well known and were studied with respect to the Bean-Shaped Accessory Glands (BAGs). The size and wet and dry weight of BAGs from infected and uninfected beetles were compared and rose to a plateau from 0-6 days post-emergence in uninfected beetles but in infected individuals continued to increase in both size and weight. These effects were density independent. Glands from both infected and uninfected beetles were assayed for trehalase activity measured by its ability to convert the sugar trehalose to glucose. The activity of this enzyme, per mg wet weight, was not affected by the parasite. However, total activity per gland increased in infected males. Total protein content and electrophoretic profiles of BAGs from infected and uninfected individuals showed no change in profile but showed an increase in all protein subunits per gland over a broad molecular weight range.

Trehalose transport and metabolism in Escherichia coli.

Trehalose metabolism in Escherichia coli is complicated by the fact that cells grown at high osmolarity synthesize internal trehalose as an osmoprotectant, independent of the carbon source, although trehalose can serve as a carbon source at both high and low osmolarity. The elucidation of the pathway of trehalose metabolism was facilitated by the isolation of mutants defective in the genes encoding transport proteins and degradative enzymes. The analysis of the phenotypes of these mutants and of the reactions catalyzed by the enzymes in vitro allowed the formulation of the degradative pathway at low osmolarity. Thus, trehalose utilization begins with phosphotransferase (IITre/IIIGlc)-mediated uptake delivering trehalose-6-phosphate to the cytoplasm. It continues with hydrolysis to trehalose and proceeds by splitting trehalose, releasing one glucose residue with the simultaneous transfer of the other to a polysaccharide acceptor. The enzyme catalyzing this reaction was named amylotrehalase. Amylotrehalase and EIITre were induced by trehalose in the medium but not at high osmolarity. treC and treB encoding these two enzymes mapped at 96.5 min on the E. coli linkage map but were not located in the same operon. Use of a mutation in trehalose-6-phosphate phosphatase allowed demonstration of the phosphoenolpyruvate- and IITre-dependent in vitro phosphorylation of trehalose. The phenotype of this mutant indicated that trehalose-6-phosphate is the effective in vivo inducer of the system.

Differential location of regulatory and nonregulatory trehalases in Candida utilis cells.

The isolation of vacuoles by density gradient centrifugation of protoplast lysates from Candida utilis cells showed a high specific activity for nonregulatory trehalase in vacuoles whereas the regulatory trehalase activatable by phosphorylation behaves as a cytoplasmic enzyme. The vacuolar trehalase is a glycoprotein that can be precipitated by Con A-Sepharose. Treatment of this enzyme with endo H reduced its reactivity with the lectin without loss of enzyme activity and decreased its apparent molecular weight by gel filtration.