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The Candida albicans TOR-Activating GTPases Gtr1 and Rhb1 Coregulate Starvation Responses and Biofilm Formation.

ABSTRACT: Target of rapamycin complex 1 (TORC1) is an essential regulator of metabolism in eukaryotic cells and in the fungal pathogen Candida albicans regulates morphogenesis and nitrogen acquisition. Gtr1 encodes a highly conserved GTPase that in Saccharomyces cerevisiae regulates nitrogen sensing and TORC1 activation. Here, we characterize the role of C. albicans GTR1 in TORC1 activation and compare it with the previously characterized GTPase Rhb1. A homozygous gtr1/gtr1 mutant exhibited impaired TORC1-mediated phosphorylation of ribosomal protein S6 and increased susceptibility to rapamycin. Overexpression of GTR1 impaired nitrogen starvation-induced filamentous growth, MEP2 expression, and growth in bovine serum albumin as the sole nitrogen source. Both GTR1 and RHB1 were shown to regulate genes involved in ribosome biogenesis, amino acid biosynthesis, and expression of biofilm growth-induced genes. The rhb1/rhb1 mutant exhibited a different pattern of expression of Sko1-regulated genes and increased susceptibility to Congo red and calcofluor white. The homozygous gtr1/gtr1 mutant exhibited enhanced flocculation phenotypes and, similar to the rhb1/rhb1 mutant, exhibited enhanced biofilm formation on plastic surfaces. In summary, Gtr1 and Rhb1 link nutrient sensing and biofilm formation and this connectivity may have evolved to enhance the competitiveness of C. albicans in niches where there is intense competition with other microbes for space and nutrients. IMPORTANCECandida albicans is the major fungal pathogen of humans and is responsible for a wide range of infections, including life-threatening systemic infections in susceptible hosts. Target of rapamycin complex 1 (TORC1) is an essential regulator of metabolism in this fungus, and components of this complex are under increased investigation as targets for new antifungal drugs. The present study characterized the role of GTR1, encoding a putative GTPase, in TORC1 activation. This study shows that GTR1 encodes a protein required for activation of TORC1 activity in response to amino acids and regulation of nitrogen starvation responses. GTR1 mutants show increased cell-cell adhesion and biofilm formation and increased expression of genes involved in these processes. This study demonstrates that starvation responses and biofilm formation are coregulated by GTR1 and suggests that these responses are linked to compete with the microbiome for space and nutrients.

SUBMITTER: Flanagan PR

PROVIDER: S-EPMC5687921 | biostudies-literature | 2017 Nov-Dec

REPOSITORIES: biostudies-literature

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The Candida albicans TOR-Activating GTPases Gtr1 and Rhb1 Coregulate Starvation Responses and Biofilm Formation.

Flanagan Peter R PR Liu Ning-Ning NN Fitzpatrick Darren J DJ Hokamp Karsten K Köhler Julia R JR Moran Gary P GP

mSphere 20171115 6

Target of rapamycin complex 1 (TORC1) is an essential regulator of metabolism in eukaryotic cells and in the fungal pathogen Candida albicans regulates morphogenesis and nitrogen acquisition. Gtr1 encodes a highly conserved GTPase that in Saccharomyces cerevisiae regulates nitrogen sensing and TORC1 activation. Here, we characterize the role of C. albicans GTR1 in TORC1 activation and compare it with the previously characterized GTPase Rhb1. A homozygous gtr1/gtr1 mut ...[more]

PMID: 29152581

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Project description:UNLABELLED:Biofilm formation by Candida albicans on medically implanted devices poses a significant clinical challenge. Here, we compared biofilm-associated gene expression in two clinical C. albicans isolates, SC5314 and WO-1, to identify shared gene regulatory responses that may be functionally relevant. Among the 62 genes most highly expressed in biofilms relative to planktonic (suspension-grown) cells, we were able to recover insertion mutations in 25 genes. Twenty mutants had altered biofilm-related properties, including cell substrate adherence, cell-cell signaling, and azole susceptibility. We focused on one of the most highly upregulated genes in our biofilm proles, RHR2, which specifies the glycerol biosynthetic enzyme glycerol-3-phosphatase. Glycerol is 5-fold-more abundant in biofilm cells than in planktonic cells, and an rhr2?/? strain accumulates 2-fold-less biofilm glycerol than does the wild type. Under in vitro conditions, the rhr2?/? mutant has reduced biofilm biomass and reduced adherence to silicone. The rhr2?/? mutant is also severely defective in biofilm formation in vivo in a rat catheter infection model. Expression profiling indicates that the rhr2?/? mutant has reduced expression of cell surface adhesin genes ALS1, ALS3, and HWP1, as well as many other biofilm-upregulated genes. Reduced adhesin expression may be the cause of the rhr2?/? mutant biofilm defect, because overexpression of ALS1, ALS3, or HWP1 restores biofilm formation ability to the mutant in vitro and in vivo. Our findings indicate that internal glycerol has a regulatory role in biofilm gene expression and that adhesin genes are among the main functional Rhr2-regulated genes. IMPORTANCE:Candida albicans is a major fungal pathogen, and infection can arise from the therapeutically intractable biofilms that it forms on medically implanted devices. It stands to reason that genes whose expression is induced during biofilm growth will function in the process, and our analysis of 25 such genes confirms that expectation. One gene is involved in synthesis of glycerol, a small metabolite that we find is abundant in biofilm cells. The impact of glycerol on biofilm formation is regulatory, not solely metabolic, because it is required for expression of numerous biofilm-associated genes. Restoration of expression of three of these genes that specify cell surface adhesins enables the glycerol-synthetic mutant to create a biofilm. Our findings emphasize the significance of metabolic pathways as therapeutic targets, because their disruption can have both physiological and regulatory consequences.

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The Candida albicans TOR-Activating GTPases Gtr1 and Rhb1 Coregulate Starvation Responses and Biofilm Formation.

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The <i>Candida albicans</i> TOR-Activating GTPases Gtr1 and Rhb1 Coregulate Starvation Responses and Biofilm Formation.

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