The early transcriptional and post-transcriptional responses to fluconazole in sensitive and resistant Candida albicans.

Candida albicans is a leading cause of fungal infections in immunocompromised patients. Management of candidemia relies on a few antifungal agents, with fluconazole being first line therapy. The emergence of fluconazole-resistant strains highlights the pressing need to improve our molecular understanding of the drug response mechanisms.

Diverse mechanisms control amino acid-dependent environmental alkalization by Candida albicans.

Candida albicans has the capacity to neutralize acidic growth environments by releasing ammonia derived from the catabolism of amino acids. The molecular components underlying alkalization and its physiological significance remain poorly understood. Here, we present an integrative model with the cytosolic NAD-dependent glutamate dehydrogenase (Gdh2) as the principal ammonia-generating component.

Proline catabolism is a key factor facilitating Candida albicans pathogenicity.

Candida albicans, the primary etiology of human mycoses, is well-adapted to catabolize proline to obtain energy to initiate morphological switching (yeast to hyphal) and for growth. We report that put1-/- and put2-/- strains, carrying defective Proline UTilization genes, display remarkable proline sensitivity with put2-/- mutants being hypersensitive due to the accumulation of the toxic intermediate pyrroline-5-carboxylate (P5C), which inhibits mitochondrial respiration. The put1-/- and put2-/- mutations attenuate virulence in Drosophila and murine candidemia models and decrease survival in human neutrophils and whole blood.

ER-localized Shr3 is a selective co-translational folding chaperone necessary for amino acid permease biogenesis.

Proteins with multiple membrane-spanning segments (MS) co-translationally insert into the endoplasmic reticulum (ER) membrane of eukaryotic cells. Shr3, an ER membrane-localized chaperone in Saccharomyces cerevisiae, is required for the functional expression of a family of 18 amino acid permeases (AAP) comprised of 12 MS. We have used comprehensive scanning mutagenesis and deletion analysis of Shr3 combined with a modified split-ubiquitin approach to probe chaperone-substrate interactions in vivo.

Amino Acid Sensing and Assimilation by the Fungal Pathogen in the Human Host.

Nutrient uptake is essential for cellular life and the capacity to perceive extracellular nutrients is critical for coordinating their uptake and metabolism. Commensal fungal pathogens, e.g.

A global metagenomic map of urban microbiomes and antimicrobial resistance.

We present a global atlas of 4,728 metagenomic samples from mass-transit systems in 60 cities over 3 years, representing the first systematic, worldwide catalog of the urban microbial ecosystem. This atlas provides an annotated, geospatial profile of microbial strains, functional characteristics, antimicrobial resistance (AMR) markers, and genetic elements, including 10,928 viruses, 1,302 bacteria, 2 archaea, and 838,532 CRISPR arrays not found in reference databases. We identified 4,246 known species of urban microorganisms and a consistent set of 31 species found in 97% of samples that were distinct from human commensal organisms.

Glutamate dehydrogenase (Gdh2)-dependent alkalization is dispensable for escape from macrophages and virulence of Candida albicans.

Candida albicans cells depend on the energy derived from amino acid catabolism to induce and sustain hyphal growth inside phagosomes of engulfing macrophages. The concomitant deamination of amino acids is thought to neutralize the acidic microenvironment of phagosomes, a presumed requisite for survival and initiation of hyphal growth. Here, in contrast to an existing model, we show that mitochondrial-localized NAD+-dependent glutamate dehydrogenase (GDH2) catalyzing the deamination of glutamate to α-ketoglutarate, and not the cytosolic urea amidolyase (DUR1,2), accounts for the observed alkalization of media when amino acids are the sole sources of carbon and nitrogen.

Spatial and temporal regulation of the endoproteolytic activity of the SPS-sensor-controlled Ssy5 signaling protease.

The Ssy5 signaling protease is a core component of the plasma membrane (PM)-localized SPS (Ssy1-Ptr3-Ssy5) sensor In response to extracellular amino acids, the SPS-sensor orchestrates the proteasomal degradation of the inhibitory Ssy5 prodomain. The unfettered catalytic (Cat)-domain cleaves latent transcription factors Stp1 and Stp2, freeing them from negative N-terminal regulatory domains. By studying the spatial and temporal constraints affecting the unfettered Cat-domain, we found that it can cleave substrates not associated with the PM; the Cat-domain efficiently cleaves Stp1 even when fused to the carboxy terminus of the endoplasmic reticulum (ER) membrane protein Shr3.

Ssy1 functions at the plasma membrane as a receptor of extracellular amino acids independent of plasma membrane-endoplasmic reticulum junctions.

Evidence from multiple laboratories has implicated Ssy1, a nontransporting amino acid permease, as the receptor component of the yeast plasma membrane (PM)-localized SPS (Ssy1-Ptr3-Ssy5)-sensor. Upon binding external amino acids, Ssy1 is thought to initiate signaling events leading to the induction of amino acid permease gene expression. In striking contrast, Kralt et al (2015) (Traffic 16:135-147) have questioned the role of Ssy1 in amino acid sensing and reported that Ssy1 is a component of the endoplasmic reticulum (ER), where it reportedly participates in the formation of ER-PM junctions.

Trk1-mediated potassium uptake contributes to cell-surface properties and virulence of Candida glabrata.

The absence of high-affinity potassium uptake in Candida glabrata, the consequence of the deletion of the TRK1 gene encoding the sole potassium-specific transporter, has a pleiotropic effect. Here, we show that in addition to changes in basic physiological parameters (e.g.

Mitochondrial proline catabolism activates Ras1/cAMP/PKA-induced filamentation in Candida albicans.

Amino acids are among the earliest identified inducers of yeast-to-hyphal transitions in Candida albicans, an opportunistic fungal pathogen of humans. Here, we show that the morphogenic amino acids arginine, ornithine and proline are internalized and metabolized in mitochondria via a PUT1- and PUT2-dependent pathway that results in enhanced ATP production. Elevated ATP levels correlate with Ras1/cAMP/PKA pathway activation and Efg1-induced gene expression.

Intersection of phosphate transport, oxidative stress and TOR signalling in Candida albicans virulence.

Phosphate is an essential macronutrient required for cell growth and division. Pho84 is the major high-affinity cell-surface phosphate importer of Saccharomyces cerevisiae and a crucial element in the phosphate homeostatic system of this model yeast. We found that loss of Candida albicans Pho84 attenuated virulence in Drosophila and murine oropharyngeal and disseminated models of invasive infection, and conferred hypersensitivity to neutrophil killing.

Ssy5 is a signaling serine protease that exhibits atypical biogenesis and marked S1 specificity.

Ssy5 is a signaling endoprotease that plays a key role in regulating central metabolism, cellular aging, and morphological transitions important for growth and survival of yeast () cells. In response to extracellular amino acids, Ssy5 proteolytically activates the transcription factors Stp1 and Stp2, leading to enhanced Ssy1-Ptr3-Ssy5 (SPS) sensor-regulated gene expression. Ssy5 comprises a catalytic (Cat) domain and an extensive regulatory prodomain.

Cdc48 and Ubx1 participate in a pathway associated with the inner nuclear membrane that governs Asi1 degradation.

The nuclear envelope is a barrier comprising outer and inner membranes that separate the cytoplasm from the nucleoplasm. The two membranes have different physical characteristics and protein compositions. The processes governing the stability of inner nuclear membrane (INM) proteins are not well characterized.

Diverse nitrogen sources in seminal fluid act in synergy to induce filamentous growth of Candida albicans.

The pathogenic fungus Candida albicans is the leading cause of vulvovaginal candidiasis (VVC). VVC represents a major quality-of-life issue for women during their reproductive years, a stage of life where the vaginal epithelium is subject to periodic hormonally induced changes associated with menstruation and concomitant exposure to serum as well as potential intermittent contact with seminal fluid. Seminal fluid potently triggers Candida albicans to switch from yeastlike to filamentous modes of growth, a developmental response tightly linked to virulence.

Protein quality control at the inner nuclear membrane.

The nuclear envelope is a double membrane that separates the nucleus from the cytoplasm. The inner nuclear membrane (INM) functions in essential nuclear processes including chromatin organization and regulation of gene expression. The outer nuclear membrane is continuous with the endoplasmic reticulum and is the site of membrane protein synthesis.

Atypical ubiquitylation in yeast targets lysine-less Asi2 for proteasomal degradation.

Proteins are typically targeted for proteasomal degradation by the attachment of a polyubiquitin chain to ϵ-amino groups of lysine residues. Non-lysine ubiquitylation of proteasomal substrates has been considered an atypical and rare event limited to complex eukaryotes. Here we report that a fully functional lysine-less mutant of an inner nuclear membrane protein in yeast, Asi2, is polyubiquitylated and targeted for proteasomal degradation.

Latency of transcription factor Stp1 depends on a modular regulatory motif that functions as cytoplasmic retention determinant and nuclear degron.

The Ssy1-Ptr3-Ssy5 (SPS)-sensing pathway enables yeast to respond to extracellular amino acids. Stp1, the effector transcription factor, is synthesized as a latent cytoplasmic precursor with an N-terminal regulatory domain that restricts its nuclear accumulation. The negative regulatory mechanisms impinging on the N-terminal domain are poorly understood.

A nuclear ubiquitin-proteasome pathway targets the inner nuclear membrane protein Asi2 for degradation.

The nuclear envelope consists of inner and outer nuclear membranes. Whereas the outer membrane is an extension of the endoplasmic reticulum, the inner nuclear membrane (INM) represents a unique membranous environment containing specific proteins. The mechanisms of integral INM protein degradation are unknown.

Rts1-protein phosphatase 2A antagonizes Ptr3-mediated activation of the signaling protease Ssy5 by casein kinase I.

Ligand-induced conformational changes of plasma membrane receptors initiate signals that enable cells to respond to discrete extracellular cues. In response to extracellular amino acids, the yeast Ssy1-Ptr3-Ssy5 sensor triggers the endoproteolytic processing of transcription factors Stp1 and Stp2 to induce amino acid uptake. Activation of the processing protease Ssy5 depends on the signal-induced phosphorylation of its prodomain by casein kinase I (Yck1/2).

Regulation of amino acid, nucleotide, and phosphate metabolism in Saccharomyces cerevisiae.

Ever since the beginning of biochemical analysis, yeast has been a pioneering model for studying the regulation of eukaryotic metabolism. During the last three decades, the combination of powerful yeast genetics and genome-wide approaches has led to a more integrated view of metabolic regulation. Multiple layers of regulation, from suprapathway control to individual gene responses, have been discovered.

Wild-type Drosophila melanogaster as a model host to analyze nitrogen source dependent virulence of Candida albicans.

The fungal pathogen Candida albicans is a common cause of opportunistic infections in humans. We report that wild-type Drosophila melanogaster (OrR) flies are susceptible to virulent C. albicans infections and have established experimental conditions that enable OrR flies to serve as model hosts for studying C.

A phosphodegron controls nutrient-induced proteasomal activation of the signaling protease Ssy5.

Regulated proteolysis serves as a mechanism to control cellular processes. The SPS (Ssy1-Ptr3-Ssy5) sensor in yeast responds to extracellular amino acids by endoproteolytically activating transcription factors Stp1 and Stp2 (Stp1/2). The processing endoprotease Ssy5 is regulated via proteasomal degradation of its noncovalently associated N-terminal prodomain.