Candida albicans is a major fungal pathogen of humans. This yeast is carried by many individuals as a harmless commensal, but when immune defences are perturbed it causes mucosal infections (thrush). Additionally, when the immune system becomes severely compromised, C. albicans often causes life-threatening systemic infections. A battery of virulence factors and fitness attributes promote the pathogenicity of C. albicans. Fitness attributes include robust responses to local environmental stresses, the inactivation of which attenuates virulence. Stress signalling pathways in C. albicans include evolutionarily conserved modules. However, there has been rewiring of some stress regulatory circuitry such that the roles of a number of regulators in C. albicans have diverged relative to the benign model yeasts Saccharomyces cerevisiae and Schizosaccharomyces pombe. This reflects the specific evolution of C. albicans as an opportunistic pathogen obligately associated with warm-blooded animals, compared with other yeasts that are found across diverse environmental niches. Our understanding of C. albicans stress signalling is based primarily on the in vitro responses of glucose-grown cells to individual stresses. However, in vivo this pathogen occupies complex and dynamic host niches characterised by alternative carbon sources and simultaneous exposure to combinations of stresses (rather than individual stresses). It has become apparent that changes in carbon source strongly influence stress resistance, and that some combinatorial stresses exert non-additive effects upon C. albicans. These effects, which are relevant to fungus–host interactions during disease progression, are mediated by multiple mechanisms that include signalling and chemical crosstalk, stress pathway interference and a biological transistor.

Population genetics of the amphibian pathogen Batrachochytrium dendrobatidis (Bd) show that isolates are highly related and globally homogenous, data that are consistent with the recent epidemic spread of a previously endemic organism. Highly related isolates are predicted to be functionally similar due to low levels of heritable genetic diversity. To test this hypothesis, we took a global panel of Bd isolates and measured (i) the genetic relatedness among isolates, (ii) proteomic profiles of isolates, (iii) the susceptibility of isolates to the antifungal drug caspofungin, (iv) the variation among isolates in growth and phenotypic characteristics, and (v) the virulence of isolates against the European common toad Bufo bufo. Our results show (i) genotypic differentiation among isolates, (ii) proteomic differentiation among isolates, (iii) no significant differences in susceptibility to caspofungin, (iv) differentiation in growth and phenotypic/morphological characters, and (v) differential virulence in B. bufo. Specifically, our data show that Bd isolates can be profiled by their genotypic and proteomic characteristics, as well as by the size of their sporangia. Bd genotypic and phenotypic distance matrices are significantly correlated, showing that less-related isolates are more biologically unique. Mass spectrometry has identified a set of candidate genes associated with inter-isolate variation. Our data show that, despite its rapid global emergence, Bd isolates are not identical and differ in several important characters that are linked to virulence. We argue that future studies need to clarify the mechanism(s) and rate at which Bd is evolving, and the impact that such variation has on the host-pathogen dynamic.

Microbes must assimilate carbon to grow and colonize their niches. Transcript profiling has suggested that Candida albicans, a major pathogen of humans, regulates its carbon assimilation in an analogous fashion to the model yeast Saccharomyces cerevisiae, repressing metabolic pathways required for the use of alterative nonpreferred carbon sources when sugars are available. However, we show that there is significant dislocation between the proteome and transcriptome in C. albicans. Glucose triggers the degradation of the ICL1 and PCK1 transcripts in C. albicans, yet isocitrate lyase (Icl1) and phosphoenolpyruvate carboxykinase (Pck1) are stable and are retained. Indeed, numerous enzymes required for the assimilation of carboxylic and fatty acids are not degraded in response to glucose. However, when expressed in C. albicans, S. cerevisiae Icl1 (ScIcl1) is subjected to glucose-accelerated degradation, indicating that like S. cerevisiae, this pathogen has the molecular apparatus required to execute ubiquitin-dependent catabolite inactivation. C. albicans Icl1 (CaIcl1) lacks analogous ubiquitination sites and is stable under these conditions, but the addition of a ubiquitination site programs glucose-accelerated degradation of CaIcl1. Also, catabolite inactivation is slowed in C. albicans ubi4 cells. Ubiquitination sites are present in gluconeogenic and glyoxylate cycle enzymes from S. cerevisiae but absent from their C. albicans homologues. We conclude that evolutionary rewiring of ubiquitination targets has meant that following glucose exposure, C. albicans retains key metabolic functions, allowing it to continue to assimilate alternative carbon sources. This metabolic flexibility may be critical during infection, facilitating the rapid colonization of dynamic host niches containing complex arrays of nutrients.

The unfolded protein response (UPR) regulates the expression of genes involved in the protein secretory pathway and in endoplasmic reticulum (ER) stress in yeasts and filamentous fungi. We have characterized the global transcriptional response of Candida albicans to ER stresses (dithiothreitol and tunicamycin) and established the impact of the transcription factor Hac1 upon this response. Expression of C. albicans Hac1, which is the functional homologue of Saccharomyces cerevisiae Hac1p, is predicted to be translationally regulated via an atypical mRNA splicing event during ER stress. C. albicans genes involved in secretion, vesicle trafficking, stress responses and cell wall biogenesis are up-regulated in response to ER stress, and translation and ribosome biogenesis genes are down-regulated. Hac1 is not essential for C. albicans viability, but plays a major role in this stress-related transcriptional response and is required for resistance to ER stress. In addition, we show that Hac1 plays an important role in regulating the morphology of C. albicans and in the expression of genes encoding cell surface proteins during ER stress, factors that are important in virulence of this fungal pathogen.

Both the generation and the analysis of proteome data are becoming increasingly widespread, and the field of proteomics is moving incrementally toward high-throughput approaches. Techniques are also increasing in complexity as the relevant technologies evolve. A standard representation of both the methods used and the data generated in proteomics experiments, analogous to that of the MIAME (minimum information about a microarray experiment) guidelines for transcriptomics, and the associated MAGE (microarray gene expression) object model and XML (extensible markup language) implementation, has yet to emerge. This hinders the handling, exchange, and dissemination of proteomics data. Here, we present a UML (unified modeling language) approach to proteomics experimental data, describe XML and SQL (structured query language) implementations of that model, and discuss capture, storage, and dissemination strategies. These make explicit what data might be most usefully captured about proteomics experiments and provide complementary routes toward the implementation of a proteome repository.

SummaryGlucose exerts profound effects upon yeast physiology. In general, the effects of high glucose concentrations (>1%) upon Saccharomyces cerevisiae have been studied. In this paper, we have characterized the global responses of yeast cells to very low (0.01%), low (0.1%) and high glucose signals (1.0%) by transcript profiling. We show that yeast is more sensitive to very low glucose signals than was previously thought, and that yeast displays different responses to these different glucose signals. Genes involved in central metabolic pathways respond rapidly to very low glucose signals, whereas genes involved in the biogenesis of cytoplasmic ribosomes generally respond only to glucose concentrations of >0.1%. We also show that cytoplasmic ribosomal protein mRNAs are transiently stabilized by glucose, indicating that both transcriptional and post-transcriptional mechanisms combine to accelerate the accumulation of ribosomal protein mRNAs. Presumably, this facilitates rapid ribosome biogenesis after exposure to glucose. However, our data indicate that yeast activates ribosome biogenesis only when sufficient glucose is available to make this metabolic investment worthwhile. In contrast, the regulation of metabolic functions in response to very low glucose signals presumably ensures that yeast can exploit even minute amounts of this preferred nutrient.

Immune cells exploit reactive oxygen species (ROS) and cationic fluxes to kill microbial pathogens, such as the fungus Candida albicans. Yet, C. albicans is resistant to these stresses in vitro. Therefore, what accounts for the potent antifungal activity of neutrophils? We show that simultaneous exposure to oxidative and cationic stresses is much more potent than the individual stresses themselves and that this combinatorial stress kills C. albicans synergistically in vitro. We also show that the high fungicidal activity of human neutrophils is dependent on the combinatorial effects of the oxidative burst and cationic fluxes, as their pharmacological attenuation with apocynin or glibenclamide reduced phagocytic potency to a similar extent. The mechanistic basis for the extreme potency of combinatorial cationic plus oxidative stress—a phenomenon we term stress pathway interference—lies with the inhibition of hydrogen peroxide detoxification by the cations. In C. albicans this causes the intracellular accumulation of ROS, the inhibition of Cap1 (a transcriptional activator that normally drives the transcriptional response to oxidative stress), and altered readouts of the stress-activated protein kinase Hog1. This leads to a loss of oxidative and cationic stress transcriptional outputs, a precipitous collapse in stress adaptation, and cell death. This stress pathway interference can be suppressed by ectopic catalase (Cat1) expression, which inhibits the intracellular accumulation of ROS and the synergistic killing of C. albicans cells by combinatorial cationic plus oxidative stress. Stress pathway interference represents a powerful fungicidal mechanism employed by the host that suggests novel approaches to potentiate antifungal therapy.

The transcription of the yeast FBP1 and PCK1 genes, which encode the gluconeogenic enzymes fructose-1,6-bisphosphatase and phosphoenolpyruvate carboxykinase, is repressed by glucose. Here, we show that this repression is both very strong and exceptionally sensitive to glucose, being triggered by glucose at concentrations less than 0.005% (0.27 mM). This repression remains operative in yeast mutants carrying any one of the three hexose kinases, but is lost in a triple hxk1, hxk2, glk1 mutant. In addition, 2-deoxyglucose can trigger the repression, but 6-deoxyglucose cannot, suggesting that internalization and phosphorylation of the glucose is essential for repression to occur. While gluconeogenic gene transcription is subject to the Mig 1p-dependent pathway of glucose repression, the exquisite response to glucose is maintained in hxk2 and mig1 mutants, suggesting that this pathway is not essential for the response. The response can also be triggered by the addition of exogenous cAMP, suggesting that the Ras/cAMP pathway can mediate repression of the FPB1 and PCK1 mRNAs. However, the response is not dependent upon this pathway because it remains intact in Ras, adenyl cyclase and protein kinase A mutants. The data show that yeast cells can detect very low glucose concentrations in the environment, and suggest that several distinct signalling pathways operate to repress FPB1 and PCK1 transcription in the presence of glucose.