Segregation of Proteinase-negative Mutants from Heterozygous Candida albicans

Crandall, Marjorie; Edwards, John E.

doi:10.1099/00221287-133-10-2817

Cited by 58 publications

(62 citation statements)

References 22 publications

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“…Nenrospora crassa produces both neutral and acid proteases if starved of either carbon, nitrogen or sulphur when grown on a protein substrate (Drucker, 1975;Cohen e t al., 1975). Similar results have been described for several Candida species (Crandall & Edwards, 1987;Banerjee etal., 1991). Proteases of certain other fungi, e.g.…”

Section: Discussionsupporting

confidence: 79%

Specific induction of a cuticle-degrading protease of the insect pathogenic fungus Metarhizium anisopliae

et al. 1994

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~The insect pathogenic fungus Metarhizium anisopliae produces several extracellular cuticle-degrading proteases and evidence is consistent that one of these, a chymoelastase PRl, is a determinant of pathogenicity. We have shown previously that the wide-domain regulatory circuits of carbon and nitrogen derepression regulate PRI production. In the present work we have established in addition that PRI is specifically induced by insect cuticle, but not by other soluble or insoluble proteinaceous substrates. The feeding of elastin or collagen to derepressed established mycelium (starved for carbon and nitrogen) did not enhance PR1 production significantly and the soluble proteins BSA and gelatin rapidly and completely repressed PRI. The carbohydrate polymers cellulose and xylan gave derepressed basal levels of PR1. However, addition of locust cuticle enhanced PRI production to a level approximately 10-fold that of derepressed mycelium. In order to establish if the enhancing effect of insect cuticle on PRI production was due to specific induction or merely a reflection of enhanced growth on this insoluble dual carbon and nitrogen source, ergosterol was used as a measure of fungal growth. Expressing enzyme activity per mg dry weight showed that PRI production in cuticle cultures increased approximately five-and ninefold after 12 and 24 h growth compared with elastin-grown cultures. Thus, the substantial increase in PR1 production on cuticle was shown not to be a function of fungal growth and this confirms that PRI is induced by a component of insect cuticle; we believe this is the first report of induction by a specific substrate for any microbial protease.

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Section: Discussionsupporting

confidence: 79%

Specific induction of a cuticle-degrading protease of the insect pathogenic fungus Metarhizium anisopliae

et al. 1994

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“…Extracellular protease secretion was assayed on unbuffered bovine serum albumin (BSA) plates (29), and lipase secretion was assayed on plates containing unbuffered YNB plus 2.5% Tween 80 (30). All plates were prepared with or without DOX.…”

Section: Methodsmentioning

confidence: 99%

The Contribution of Candida albicans Vacuolar ATPase Subunit V ₁ B, Encoded by VMA2 , to Stress Response, Autophagy, and Virulence Is Independent of Environmental pH

et al. 2014

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Candida albicans vacuoles are central to many critical biological processes, including filamentation and in vivo virulence. The V-ATPase proton pump is a multisubunit complex responsible for organellar acidification and is essential for vacuolar biogenesis and function. To study the function of the V 1 B subunit of C. albicans V-ATPase, we constructed a tetracycline-regulatable VMA2 mutant, tetR-VMA2. Inhibition of VMA2 expression resulted in the inability to grow at alkaline pH and altered resistance to calcium, cold temperature, antifungal drugs, and growth on nonfermentable carbon sources. Furthermore, V-ATPase was unable to fully assemble at the vacuolar membrane and was impaired in proton transport and ATPase-specific activity. VMA2 repression led to vacuolar alkalinization in addition to abnormal vacuolar morphology and biogenesis. Key virulence-related traits, including filamentation and secretion of degradative enzymes, were markedly inhibited. These results are consistent with previous studies of C. albicans V-ATPase; however, differential contributions of the V-ATPase V o and V 1 subunits to filamentation and secretion are observed. We also make the novel observation that inhibition of C. albicans V-ATPase results in increased susceptibility to osmotic stress. Notably, V-ATPase inhibition under conditions of nitrogen starvation results in defects in autophagy. Lastly, we show the first evidence that V-ATPase contributes to virulence in an acidic in vivo system by demonstrating that the tetR-VMA2 mutant is avirulent in a Caenorhabditis elegans infection model. This study illustrates the fundamental requirement of V-ATPase for numerous key virulence-related traits in C. albicans and demonstrates that the contribution of VATPase to virulence is independent of host pH.T he fungal pathogen Candida albicans is the fourth most common cause of hospital-acquired bloodstream infections and is a major cause of catheter-associated infections, sepsis, and devicerelated infections. It is also an extremely common cause of urinary and mucosal infections. Despite its clinical significance, the diagnosis and treatment of disseminated candidiasis remain limited by an incomplete understanding of its molecular pathogenesis. The fungal vacuole, a degradative organelle roughly equivalent to the mammalian lysosome, plays an important role in numerous biological processes in C. albicans, including key aspects of pathogenesis. Previous studies have established that C. albicans mutants compromised in vacuolar function are defective in yeast-to-hypha transitioning, a major virulence-related trait, and exhibit reduced virulence in vivo (1, 2).An essential component of vacuolar biogenesis and function is the vacuolar H ϩ -ATPase (V-ATPase) proton pump, which is a multisubunit complex responsible for the acidification of internal organelles. V-ATPase is located at the vacuolar membrane and throughout the endomembrane system, including prevacuolar compartments and the Golgi complex (1). The acidification of these organelles has sever...

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“…Extracellular aspartyl protease secretion/activity was assayed on BSA plates after incubation at 30°C for 3 days (36). Lipase secretion/activity was assayed on yeast nitrogen base plates containing 2.5% Tween 80 after incubation at 37°C for 6 days (37).…”

Section: Methodsmentioning

confidence: 99%

Deletion of Vacuolar Proton-translocating ATPase Voa Isoforms Clarifies the Role of Vacuolar pH as a Determinant of Virulence-associated Traits in Candida albicans*

Raines¹,

Rane

Bernardo

et al. 2013

Journal of Biological Chemistry

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Background: V-ATPase regulates pH, and Candida albicans virulence is pH-dependent. Results: Deletion of V-ATPase V o a subunit Vph1p, but not Stv1p, alkalinizes vacuoles; several virulence-related traits remain unaffected. Conclusion: Vacuolar acidification is not essential for in vitro filamentation, biofilm formation, and macrophage killing in C. albicans. Significance: Stv1p in non-vacuolar organelles may play important roles in C. albicans infectivity, particularly if Vph1p is not functional.

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Segregation of Proteinase-negative Mutants from Heterozygous Candida albicans

Cited by 58 publications

References 22 publications

Specific induction of a cuticle-degrading protease of the insect pathogenic fungus Metarhizium anisopliae

Specific induction of a cuticle-degrading protease of the insect pathogenic fungus Metarhizium anisopliae

The Contribution of Candida albicans Vacuolar ATPase Subunit V ₁ B, Encoded by VMA2 , to Stress Response, Autophagy, and Virulence Is Independent of Environmental pH

Deletion of Vacuolar Proton-translocating ATPase Voa Isoforms Clarifies the Role of Vacuolar pH as a Determinant of Virulence-associated Traits in Candida albicans*

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Segregation of Proteinase-negative Mutants from Heterozygous Candida albicans

Cited by 58 publications

References 22 publications

Specific induction of a cuticle-degrading protease of the insect pathogenic fungus Metarhizium anisopliae

Specific induction of a cuticle-degrading protease of the insect pathogenic fungus Metarhizium anisopliae

The Contribution of Candida albicans Vacuolar ATPase Subunit V 1 B, Encoded by VMA2 , to Stress Response, Autophagy, and Virulence Is Independent of Environmental pH

Deletion of Vacuolar Proton-translocating ATPase Voa Isoforms Clarifies the Role of Vacuolar pH as a Determinant of Virulence-associated Traits in Candida albicans*

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The Contribution of Candida albicans Vacuolar ATPase Subunit V ₁ B, Encoded by VMA2 , to Stress Response, Autophagy, and Virulence Is Independent of Environmental pH