An expanded regulatory network temporally controls Candida albicans biofilm formation.
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SUBMITTER: Fox EP
PROVIDER: S-EPMC4464956 | biostudies-literature |
REPOSITORIES: biostudies-literature
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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.
| S-EPMC3622937 | biostudies-literature
Project description:A biofilm is an organized, resilient group of microbes in which individual cells acquire properties, such as drug resistance, that are distinct from those observed in suspension cultures. Here, we describe and analyze the transcriptional network controlling biofilm formation in the pathogenic yeast Candida albicans, whose biofilms are a major source of medical device-associated infections. We have combined genetic screens, genome-wide approaches, and two in vivo animal models to describe a master circuit controlling biofilm formation, composed of six transcription regulators that form a tightly woven network with ?1,000 target genes. Evolutionary analysis indicates that the biofilm network has rapidly evolved: genes in the biofilm circuit are significantly weighted toward genes that arose relatively recently with ancient genes being underrepresented. This circuit provides a framework for understanding many aspects of biofilm formation by C. albicans in a mammalian host. It also provides insights into how complex cell behaviors can arise from the evolution of transcription circuits.
| S-EPMC3266547 | biostudies-literature
Project description:Biofilm formation on medically implanted devices by Candida albicans 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 50 genes most highly expressed in biofilms relative to planktonic (suspension-grown) cells, we were able to recover insertion mutations in 25 genes. We observed that 20 of the 25 mutants have altered biofilm-related properties, including cell-substrate adherence, cell-cell signaling, and azole susceptibility. We focused on the most highly up-regulated gene in biofilms, RHR2, which specifies the glycerol biosynthetic enzyme glycerol-3-phosphate phosphatase. Glycerol is 5-fold more abundant in biofilm cells than planktonic cells, and an rhr2D/D strain accumulates 2-fold less biofilm glycerol than the wild type. Under in vitro growth conditions, the rhr2D/D mutant has reduced biofilm biomass and reduced adherence to silicone. The rhr2D/D mutant is severely defective in biofilm formation in vivo, in a rat catheter infection model. Expression profiling of the rhr2D/D mutant indicates that it has reduced expression of cell surface adhesin genes ALS1, ALS3, and HWP1, as well as a large fraction of all other biofilm up-regulated genes. Reduced adhesin expression is the cause of the rhr2D/D 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.
2013-04-12 | GSE45141 | GEO
Project description:Biofilm formation on medically implanted devices by Candida albicans 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 50 genes most highly expressed in biofilms relative to planktonic (suspension-grown) cells, we were able to recover insertion mutations in 25 genes. We observed that 20 of the 25 mutants have altered biofilm-related properties, including cell-substrate adherence, cell-cell signaling, and azole susceptibility. We focused on the most highly up-regulated gene in biofilms, RHR2, which specifies the glycerol biosynthetic enzyme glycerol-3-phosphate phosphatase. Glycerol is 5-fold more abundant in biofilm cells than planktonic cells, and an rhr2D/D strain accumulates 2-fold less biofilm glycerol than the wild type. Under in vitro growth conditions, the rhr2D/D mutant has reduced biofilm biomass and reduced adherence to silicone. The rhr2D/D mutant is severely defective in biofilm formation in vivo, in a rat catheter infection model. Expression profiling of the rhr2D/D mutant indicates that it has reduced expression of cell surface adhesin genes ALS1, ALS3, and HWP1, as well as a large fraction of all other biofilm up-regulated genes. Reduced adhesin expression is the cause of the rhr2D/D 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. Gene expression profiles, in duplicate; (1) for biofilm vs. planktonic growth conditions for the two wild-type clinical isolates of Candida albicans (SC5314 and WO1-white/WO1-opaque), and (2) for rhr2M-NM-^T/M-NM-^T mutant and complemented strain, via RNA-deep sequencing using Illumina GA2 and HiSeq2000 platforms, respectively
2013-04-12 | E-GEOD-45141 | biostudies-arrayexpress
Project description:Candida albicans is an opportunistic fungal pathogen that can infect oral mucosal surfaces while being under continuous flow from saliva. Under specific conditions, C. albicans will form microcolonies that more closely resemble the biofilms formed in vivo than standard in vitro biofilm models. However, very little is known about these microcolonies, particularly genomic differences between these specialized biofilm structures and the traditional in vitro biofilms. In this study, we used a novel flow system, in which C. albicans spontaneously forms microcolonies, to further characterize the architecture of fungal microcolonies and their genomics compared to non-microcolony conditions. Fungal microcolonies arose from radially branching filamentous hyphae that increasingly intertwined with one another to form extremely dense biofilms, and closely resembled the architecture of in vivo oropharyngeal candidiasis. We identified 20 core microcolony genes that were differentially regulated in flow-induced microcolonies using RNA-seq. These genes included HWP1, ECE1, IHD1, PLB1, HYR1, PGA10, and SAP5. A predictive algorithm was utilized to identify ten transcriptional regulators potentially involved in microcolony formation. Of these transcription factors, we found that Rob1, Ndt80, Sfl1 and Sfl2, played a key role in microcolony formation under both flow and static conditions and to epithelial surfaces. Expression of core microcolony genes were highly up-regulated in Δsfl1 cells and down-regulated in both Δsfl2 and Δrob1 strains. Microcolonies formed on oral epithelium using C. albicans Δsfl1, Δsfl2 and Δrob1 deletion strains all had altered adhesion, invasion and cytotoxicity. Furthermore, epithelial cells infected with deletion mutants had reduced (SFL2, NDT80, and ROB1) or enhanced (SFL2) immune responses, evidenced by phosphorylation of MKP1 and c-Fos activation, key signal transducers in the hyphal invasion response. This profile of microcolony transcriptional regulators more closely reflects Sfl1 and Sfl2 hyphal regulatory networks than static biofilm regulatory networks, suggesting that microcolonies are a specialized pathogenic form of biofilm.
| S-EPMC6173444 | biostudies-literature
Project description:The planktonic versus biofilm gene expression arrays were performed in a/alpha cell types. Gene expression arrays were performed on planktonic vs biofilm cells grown in Spider medium at 37C. Normalized data is reported in matrix.
2011-07-08 | GSE30474 | GEO
Project description:We determined the changes in transcriptional profiles that occur in the first hour following the transfer of Candida albicans to hypoxic growth conditions. The impressive speed of this response is not compatible with current models of fungal adaptation to hypoxia that depend on the depletion of sterol and heme. Functional analysis using Gene Set Enrichment Analysis (GSEA) identified the Sit4 phosphatase, Ccr4 mRNA deacetylase, and Sko1 transcription factor (TF) as potential regulators of the early hypoxic response. Cells mutated in these and other regulators exhibit a delay in their transcriptional responses to hypoxia. Promoter occupancy data for 29 TFs were combined with the transcriptional profiles of 3,111 in vivo target genes in a Network Component Analysis (NCA) to produce a model of the dynamic and highly interconnected TF network that controls this process. With data from the TF network obtained from a variety of sources, we generated an edge and node model that was capable of separating many of the hypoxia-upregulated and -downregulated genes. Upregulated genes are centered on Tye7, Upc2, and Mrr1, which are associated with many of the gene promoters that exhibit the strongest activations. The connectivity of the model illustrates the high redundancy of this response system and the challenges that lie in determining the individual contributions of specific TFs. Finally, treating cells with an inhibitor of the oxidative phosphorylation chain mimics most of the early hypoxic profile, which suggests that this response may be initiated by a drop in ATP production.
| S-EPMC4060469 | biostudies-literature
Project description:Candida albicans is a leading pathogen in infections of central venous catheters, which are frequently infused with heparin. Binding of C. albicans to medically relevant concentrations of soluble and plate-bound heparin was demonstrable by confocal microscopy and enzyme-linked immunosorbent assay (ELISA). A sequence-based search identified 34 C. albicans surface proteins containing ≥1 match to linear heparin-binding motifs. The virulence factor Int1 contained the most putative heparin-binding motifs (n = 5); peptides encompassing 2 of 5 motifs bound to heparin-Sepharose. Alanine substitution of lysine residues K805/K806 in 804QKKHQIHK811 (motif 1 of Int1) markedly attenuated biofilm formation in central venous catheters in rats, whereas alanine substitution of K1595/R1596 in 1593FKKRFFKL1600 (motif 4 of Int1) did not impair biofilm formation. Affinity-purified immunoglobulin G (IgG) recognizing motif 1 abolished biofilm formation in central venous catheters; preimmune IgG had no effect. After heparin treatment of C. albicans, soluble peptides from multiple C. albicans surface proteins were detected, such as Eno1, Pgk1, Tdh3, and Ssa1/2 but not Int1, suggesting that heparin changes candidal surface structures and may modify some antigens critical for immune recognition. These studies define a new mechanism of biofilm formation for C. albicans and a novel strategy for inhibiting catheter-associated biofilms.
| S-EPMC4038792 | biostudies-literature
Project description:BACKGROUND: Candida albicans can form biofilms on intravenous catheters; this process plays a key role in the pathogenesis of catheter infections. This study evaluated the effect of human serum (HS) on C. albicans biofilm formation and the expression of adhesion-related genes in vitro. A C. albicans laboratory strain (ATCC90028) and three clinical strains were grown for 24 h in RPMI 1640 supplemented with HS or RPMI 1640 alone (as a control). The growth of biofilm cells of four strains was monitored by a Live Cell Movie Analyzer, and by XTT reduction assay. The expression of the adhesion-related genes BCR1, ALS1, ALS3, HWP1 and ECE1 was analyzed by RT-PCR at three time points (60 min, 90 min, and 24 h). RESULTS: In the adhesion phase, C. albicans cells kept a Brownian movement in RPMI medium containing HS until a large number of germ tubes were formed. In the control group, C. albicans cells quickly adhered to the bottom of the reaction plate. Compared with RPMI 1640, medium supplemented with 3-50% HS caused a significant decrease in biofilm development (all p < 0.001). However, the presence of HS had no significant inhibitory effect on the pre-adhered biofilms (all p > 0.05). Biofilm formation was also inhibited by heat-inactivated and proteinase K pre-treated HS. The presence of 50% HS did not significantly affect the planktonic growth of C. albicans (p > 0.05). At three time points, HS inhibited expression of the ALS1 and ALS3 genes and promoted expression of the HWP1 and ECE1 genes. Significant up-regulation of BCR1 was observed only at the 90-min point. CONCLUSIONS: Human serum reduces biofilm formation by inhibiting the adhesion of C. albicans cells. This response may be associated with the down-regulation of adhesion-related genes ALS1, ALS3 and BCR1. The inhibitory serum component is protease-resistant and heat stable.
| S-EPMC4101872 | biostudies-other
Project description:Aggregatibacter actinomycetemcomitans, a Gram-negative bacterium, and Candida albicans, a polymorphic fungus, are both commensals of the oral cavity but both are opportunistic pathogens that can cause oral diseases. A. actinomycetemcomitans produces a quorum-sensing molecule called autoinducer-2 (AI-2), synthesized by LuxS, that plays an important role in expression of virulence factors, in intra- but also in interspecies communication. The aim of this study was to investigate the role of AI-2 based signaling in the interactions between C. albicans and A. actinomycetemcomitans. A. actinomycetemcomitans adhered to C. albicans and inhibited biofilm formation by means of a molecule that was secreted during growth. C. albicans biofilm formation increased significantly when co-cultured with A. actinomycetemcomitans luxS, lacking AI-2 production. Addition of wild-type-derived spent medium or synthetic AI-2 to spent medium of the luxS strain, restored inhibition of C. albicans biofilm formation to wild-type levels. Addition of synthetic AI-2 significantly inhibited hypha formation of C. albicans possibly explaining the inhibition of biofilm formation. AI-2 of A. actinomycetemcomitans is synthesized by LuxS, accumulates during growth and inhibits C. albicans hypha- and biofilm formation. Identifying the molecular mechanisms underlying the interaction between bacteria and fungi may provide important insight into the balance within complex oral microbial communities.
| S-EPMC4104835 | biostudies-literature