Time-Course Proteome Analysis Reveals the Dynamic Response of Cryptococcus gattii Cells to Fluconazole

Chong, Hin Siong; Campbell, Leona T.; Padula, Matthew P.; Hill, Charles; Harry, Elizabeth J.; Li, Simone S.; Wilkins, Marc R.; Herbert, Ben R.; Carter, Dee

doi:10.1371/journal.pone.0042835

Cited by 17 publications

(12 citation statements)

References 77 publications

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“…Over the last few years, these approaches have been helpful in identifying both novel and specific targets, and to describe the molecular interactions between drugs and fungi, as previously reported for C . albicans [17,27], Cryptococcus gattii [28], or Aspergillus fumigatus [29,30]. Surprisingly, our results showed no significant differential protein abundance between VRC-treated and non-treated cell whole proteomes.…”

Section: Discussionmentioning

confidence: 67%

“…Regarding the 60S ribosomal proteins that were found down-regulated upon VRC exposure, these are involved in protein translation, as structural parts forming ribosomes. Interestingly, they have been associated with antifungal stress by either being down- [40] or over-expressed [28], which may be due to differences in the mechanisms pathways of action used by the drugs. It is worth highlighting the fact that both Srp1 and 60S ribosomal proteins are related to protein synthesis, and may be related to the morphological changes occurring in L .…”

Section: Discussionmentioning

confidence: 99%

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Molecular and cellular responses of the pathogenic fungus Lomentospora prolificans to the antifungal drug voriconazole

et al. 2017

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The filamentous fungus Lomentospora (Scedosporium) prolificans is an emerging opportunistic pathogen associated with fatal infections in patients with disturbed immune function. Unfortunately, conventional therapies are hardly of any use against this fungus due to its intrinsic resistance. Therefore, we performed an integrated study of the L. prolificans responses to the first option to treat these mycoses, namely voriconazole, with the aim of unveiling mechanisms involved in the resistance to this compound. To do that, we used a wide range of techniques, including fluorescence and electron microscopy to study morphological alterations, ion chromatography to measure changes in cell-wall carbohydrate composition, and proteomics-based techniques to identify the proteins differentially expressed under the presence of the drug. Significantly, we showed drastic changes occurring in cell shape after voriconazole exposure, L. prolificans hyphae being shorter and wider than under control conditions. Interestingly, we proved that the architecture and carbohydrate composition of the cell wall had been modified in the presence of the drug. Specifically, L. prolificans constructed a more complex organelle with a higher presence of glucans and mannans. In addition to this, we identified several differentially expressed proteins, including Srp1 and heat shock protein 70 (Hsp70), as the most overexpressed under voriconazole-induced stress conditions. The mechanisms described in this study, which may be directly related to L. prolificans antifungal resistance or tolerance, could be used as targets to improve existing therapies or to develop new ones in order to successfully eliminate these mycoses.

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Section: Discussionmentioning

confidence: 67%

Section: Discussionmentioning

confidence: 99%

Molecular and cellular responses of the pathogenic fungus Lomentospora prolificans to the antifungal drug voriconazole

et al. 2017

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“…In addition to the capacity for proteomics to provide insight into the pathways affected by antifungals, it can also inform on promising targets for antifungal activity. Evaluation of proteomic changes in response to a time-course with the antifungal drug fluconazole revealed potential synergistic drug targets in Cryptococcus gattii following compilation of protein interaction networks (Chong et al 2012). An MS-based comparative proteomic strategy involving the use of labels has been used to elucidate the mechanism of action of the human AFP histatin 5 (Komatsu et al 2011).…”

Section: Analytical Tools For Characterizing the Effects Of Afpsmentioning

confidence: 99%

Manuscript title: antifungal proteins from moulds: analytical tools and potential application to dry-ripened foods

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Doyle

et al. 2016

Appl Microbiol Biotechnol

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Moulds growing on the surface of dry-ripened foods contribute to their sensory qualities, but some of them are able to produce mycotoxins that pose a hazard to consumers. Small cysteine-rich antifungal proteins (AFPs) from moulds are highly stable to pH and proteolysis and exhibit a broad inhibition spectrum against filamentous fungi, providing new chances to control hazardous moulds in fermented foods. The analytical tools for characterizing the cellular targets and affected pathways are reviewed. Strategies currently employed to study these mechanisms of action include 'omics' approaches that have come to the forefront in recent years, developing in tandem with genome sequencing of relevant organisms. These techniques contribute to a better understanding of the response of moulds against AFPs, allowing the design of complementary strategies to maximize or overcome the limitations of using AFPs on foods. AFPs alter chitin biosynthesis, and some fungi react inducing cell wall integrity (CWI) pathway. However, moulds able to increase chitin content at the cell wall by increasing proteins in either CWI or calmodulin-calcineurin signalling pathways will resist AFPs. Similarly, AFPs increase the intracellular levels of reactive oxygen species (ROS), and moulds increasing G-protein complex β subunit CpcB and/or enzymes to efficiently produce glutathione may evade apoptosis. Unknown aspects that need to be addressed include the interaction with mycotoxin production by less sensitive toxigenic moulds. However, significant steps have been taken to encourage the use of AFPs in intermediate-moisture foods, particularly for mould-ripened cheese and meat products.

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“…In our previous work on Cryptococcus proteomics, we found conditions optimized to simulate those encountered in the host induced the production of large amounts of shed capsular material, which interfered with the isolation and identification of proteins 4 . Therefore, we developed a novel method of protein capture using BioRad ProteoMinerÔ beads, followed by mass spectroscopy.…”

mentioning

confidence: 99%

“…While there are sporadic reports of T. marneffei infections in 'immunocompetent hosts' the immune status has not been adequately tested in these cases, and the term 'immunocompetent' is often used interchangeably (and incorrectly) in these reports with HIV negative status. The ecological niche of T. marneffei is unclear, but there is a strong association with a number of bamboo rat species in endemic areas 3,4 .…”

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confidence: 99%

You are what you secrete: extracellular proteins and virulence in Cryptococcus

et al. 2015

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Fungal organisms secrete a wide range of biomolecules, including degradative enzymes that are essential for nutrition, toxins, effectors and secondary compounds that modulate interactions with host animals and plants, and a variety of signaling and stress-related proteins 1 . As these are likely to be key determinants of virulence and may also be useful diagnostic and therapeutic targets, we investigated the secretome of different strains of the fungal pathogen Cryptococcus. Virulent strains secreted predominantly hydrolytic and proteolytic enzymes, while the least virulent strain secreted a range of additional non-degradative proteins including many that lacked secretion signals, some that appear to be 'moonlighting', and a number that are known to be allergenic. It appears that in Cryptococcus, the secretome may influence virulence both through the presence of harmful enzymes and through the absence of proteins that alert the host defence mechanisms.Cryptococcus is an encapsulated yeast with two predominant pathogenic species: Cryptococcus neoformans and Cryptococcus gattii.These cause cryptococcosis in animals and humans, with disease ranging from asymptomatic to severe, fatal meningitis. There are a number of differences between C. gattii and C. neoformans including their preferred environmental niche, basidiospore morphology, drug susceptibility, epidemiology, the clinical manifestations of associated disease, and host susceptibility 2 . In addition, there are significant differences among stains within each species. In C. gattii, a hypervirulent sub-genotype designated VGIIa has caused a recent significant outbreak of cryptococcosis on Vancouver Island in British Columbia, Canada and in the Pacific Northwest of the United States.In contrast, a closely related sub-genotype designated VGIIb is globally distributed and hypovirulent 3 . These differences betweenCryptococcus species and sub-genotypes provide an opportunity for understanding pathogenicity and disease progression by what are otherwise very genetically similar fungal organisms.Our laboratory has been using 'omics approaches to understand virulence in Cryptococcus, and used proteomic analysis to characterize the secretome produced by three Cryptococcus strains.Two strains were of high virulence (C. neoformans and C. gattii sub-genotype VGIIa) and the third of low virulence (C. gattii sub-genotype VGIIb). In our previous work on Cryptococcus proteomics, we found conditions optimized to simulate those encountered in the host induced the production of large amounts of shed capsular material, which interfered with the isolation and identification of proteins 4 . Therefore, we developed a novel method of protein capture using BioRad ProteoMinerÔ beads, followed by mass spectroscopy. Sixty-seven cryptococcal proteins were identified and only one was common to all three strains. The secretomes of the high virulence C. neoformans and C. gattii VGIIa strains were similar and mostly consisted of a hydrolytic and proteolytic proteins.In contrast the lower vir...

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Time-Course Proteome Analysis Reveals the Dynamic Response of Cryptococcus gattii Cells to Fluconazole

Cited by 17 publications

References 77 publications

Molecular and cellular responses of the pathogenic fungus Lomentospora prolificans to the antifungal drug voriconazole

Molecular and cellular responses of the pathogenic fungus Lomentospora prolificans to the antifungal drug voriconazole

Manuscript title: antifungal proteins from moulds: analytical tools and potential application to dry-ripened foods

You are what you secrete: extracellular proteins and virulence in Cryptococcus

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