Like many bacteria, yeast species can form biofilms on several surfaces. Candida albicans colonizes the surfaces of catheters, prostheses, and epithelia, forming biofilms that are extremely resistant to antifungal drugs. We have used transcript profiling to investigate the specific properties of C. albicans biofilms. Biofilm and planktonic cultures produced under different conditions of nutrient flow, aerobiosis, or glucose concentration were compared by overall gene expression correlation. Correlation was much higher between biofilms than planktonic populations irrespective of the growth conditions, indicating that biofilm populations formed in different environments display very similar and specific transcript profiles. A first cluster of 325 differentially expressed genes was identified. In agreement with the overrepresentation of amino acid biosynthesis genes in this cluster, Gcn4p, a regulator of amino acid metabolism, was shown to be required for normal biofilm growth. To identify biofilm-related genes that are independent of mycelial development, we studied the transcriptome of biofilms produced by a wild-type, hypha-producing strain and a cph1/cph1 efg1/efg1 strain defective for hypha production. This analysis identified a cluster of 317 genes expressed independently of hypha formation, whereas 86 genes were dependent on mycelial development. Both sets revealed the activation of the sulfur-amino acid biosynthesis pathway as a feature of C. albicans biofilms.

SummaryBiofilm is the predominant type of microbial development in natural environments, and potentially represents a major form of resistance or source of recurrence during host infection. Although a large number of studies have focussed on the genetics of bacterial biofilm formation, very little is known about the genes involved in this type of growth in fungi. A genetic screen for Candida glabrata Biofilm mutants was performed using a 96-well plate model of biofilm formation. Study of the isolated mutant strains allowed the identification of four genes involved in biofilm formation ( RIF1 , SIR4 , EPA6 and YAK1 ). Epa6p is a newly identified adhesin required for biofilm formation in this pathogenic yeast. EPA6 and its close paralogue EPA7 are located in subtelomeric regions and their transcription is regulated by Sir4p and Rif1p, two proteins involved in subtelomeric silencing. Biofilm growth conditions induce the transcription of EPA6 and EPA7 : this is dependent on the presence of an intact subtelomeric silencing machinery and is independent of the Mpk1p signalling pathway. Finally, the kinase Yak1p is required for expression of both adhesin genes and acts through a subtelomeric silencing machinery-dependent pathway.

In budding yeast, Tup1 and Ssn6/Cyc8 form a corepressor that regulates a large number of genes. This Tup1-Ssn6 corepressor appears to be conserved from yeast to man. In the pathogenic fungus Candida albicans, Tup1 regulates cellular morphogenesis, phenotypic switching, and metabolism, but the role of Ssn6 remains unclear. We show that there are clear differences in the morphological and invasive phenotypes of C. albicans ssn6 and tup1 mutants. Unlike Tup1, Ssn6 depletion promoted morphological events reminiscent of phenotypic switching rather than filamentous growth. Transcript profiling revealed minimal overlap between the Ssn6 and Tup1 regulons. Hypha-specific genes, which are repressed by Tup1 and Nrg1, were not derepressed in ssn6 cells under the conditions studied. In contrast, the phase specific gene WH11 was derepressed in ssn6 cells, but not in tup1 or nrg1 cells. Hence Ssn6 and Tup1 play distinct roles in C. albicans. Nevertheless, both Ssn6 and Tup1 were required for the Nrg1-mediated repression of an artificial NRE promoter, and lexA-Nrg1 mediated repression in the C. albicans one-hybrid system. These observations are explained in models that are generally consistent with the Tup1-Ssn6 paradigm in budding yeast. INTRODUCTIONTranscriptional repression plays a key role in controlling the growth and differentiation of eukaryotic cells. The fundamental importance of negative transcriptional regulators is reflected in their conservation during eukaryotic evolution. For example, orthologues of the Ssn6 (Cyc8) and Tup1 proteins, which act as global repressors in Saccharomyces cerevisiae, have been identified in humans, flies, worms, slime molds, and fungi (reviewed by Smith and Johnson, 2000).Our understanding of the mechanisms of action of Ssn6 and Tup1 is based largely on studies in budding yeast. The S. cerevisiae paradigm suggests that Ssn6 and Tup1 interact physically to form a corepressor complex that actively represses the transcriptional machinery (Williams et al., 1991;Redd et al., 1997;Gounalaki et al., 2000). The relevance of this paradigm to eukaryotes in general is emphasized by the observation that yeast Ssn6 can interact with human Tup1-like proteins to mediate transcriptional repression in human cells (Grbavec et al., 1999).The Ssn6-Tup1 corepressor is thought to repress transcription in yeast by two main mechanisms. First, the corepressor is thought to remodel chromatin on yeast promoters by recruiting histone deacetylases to these promoters and by positioning nucleosomes via direct interactions with histone tails (Edmondson et al., 1996;Davie et al., 2002Davie et al., , 2003Zhang and Reese, 2004a). Second, Tup1 is thought to interact directly with the transcriptional machinery to attenuate its activity (Carlson, 1997;Redd et al., 1997;Gromoller and Lehming, 2000;Papamichos-Chronakis et al., 2000). Recent experiments suggest that both mechanisms operate in yeast, but that they are utilized differentially to regulate distinct sets of yeast genes (Green and Johnson, 2004). However, both mechan...

BackgroundThe impact of nano-scaled materials on photosynthetic organisms needs to be evaluated. Plants represent the largest interface between the environment and biosphere, so understanding how nanoparticles affect them is especially relevant for environmental assessments. Nanotoxicology studies in plants allude to quantum size effects and other properties specific of the nano-stage to explain increased toxicity respect to bulk compounds. However, gene expression profiles after exposure to nanoparticles and other sources of environmental stress have not been compared and the impact on plant defence has not been analysed.ResultsArabidopsis plants were exposed to TiO2-nanoparticles, Ag-nanoparticles, and multi-walled carbon nanotubes as well as different sources of biotic (microbial pathogens) or abiotic (saline, drought, or wounding) stresses. Changes in gene expression profiles and plant phenotypic responses were evaluated. Transcriptome analysis shows similarity of expression patterns for all plants exposed to nanoparticles and a low impact on gene expression compared to other stress inducers. Nanoparticle exposure repressed transcriptional responses to microbial pathogens, resulting in increased bacterial colonization during an experimental infection. Inhibition of root hair development and transcriptional patterns characteristic of phosphate starvation response were also observed. The exogenous addition of salicylic acid prevented some nano-specific transcriptional and phenotypic effects, including the reduction in root hair formation and the colonization of distal leaves by bacteria.ConclusionsThis study integrates the effect of nanoparticles on gene expression with plant responses to major sources of environmental stress and paves the way to remediate the impact of these potentially damaging compounds through hormonal priming.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-1530-4) contains supplementary material, which is available to authorized users.

CandidaDB is a database dedicated to the genome of the most prevalent systemic fungal pathogen of humans, Candida albicans. CandidaDB is based on an annotation of the Stanford Genome Technology Center C.albicans genome sequence data by the European Galar Fungail Consortium. CandidaDB Release 2.0 (June 2004) contains information pertaining to Assembly 19 of the genome of C.albicans strain SC5314. The current release contains 6244 annotated entries corresponding to 130 tRNA genes and 5917 protein-coding genes. For these, it provides tentative functional assignments along with numerous pre-run analyses that can assist the researcher in the evaluation of gene function for the purpose of specific or large-scale analysis. CandidaDB is based on GenoList, a generic relational data schema and a World Wide Web interface that has been adapted to the handling of eukaryotic genomes. The interface allows users to browse easily through genome data and retrieve information. CandidaDB also provides more elaborate tools, such as pattern searching, that are tightly connected to the overall browsing system. As the C.albicans genome is diploid and still incompletely assembled, CandidaDB provides tools to browse the genome by individual supercontigs and to examine information about allelic sequences obtained from complementary contigs. CandidaDB is accessible at http://genolist.pasteur.fr/CandidaDB.

Many physiological responses are regulated by serpins (serine protease inhibitors) in mammals, including the blood clotting, inflammatory, complement activation, and angiogenesis pathways (15,48). Disorders in serpin metabolism are responsible for a wide range of human diseases, such as emphysema, cirrhosis, blood coagulation disorders, and dementia (36, 38). For this reason, the inhibitory mechanism of serpins has been extensively studied, mainly in humans and mammalian model organisms.Serpins are large-molecular-mass protease inhibitors with a core of three ␤-sheets connected by short, ␣-helical linkers. In the native state, a reactive center loop (RCL) extends out from the serpin core and presents an ideal bait to the target protease (48). The native serpin is in a metastable (stressed) configuration. Following cleavage, the structure adopts a stable (relaxed) configuration. The rapid insertion of the RCL into the serpin ␤-sheet A distorts the protease's catalytic site so that the esterification step of the hydrolysis reaction cannot be completed. As a result, the protease and serpin form a covalently linked complex. The protease is translocated through 70 Å and crushed against the serpin core (25). This process denatures the serpin/protease complex, which is targeted for proteolytic destruction. This "suicide-inhibition" mechanism destroys the activity of both the serpin and target protease, both of which have a high turnover. Reduction in serpin levels results in explosive activation of signaling pathways as inhibition of the target protease is lost. Each subsequent zymogen in the proteolytic cascade is activated by its upstream protease. This mechanism gives a rapid, amplified response. With continued activation, both serpin and downstream proteases tend to be upregulated at the transcriptional level.

Microbe-Associated Molecular Patterns and virulence effectors are recognized by plants as a first step to mount a defence response against potential pathogens. This recognition involves a large family of extracellular membrane receptors and other immune proteins located in different sub-cellular compartments. We have used phage-display technology to express and select for Arabidopsis proteins able to bind bacterial pathogens. To rapidly identify microbe-bound phage, we developed a monitoring method based on microarrays. This combined strategy allowed for a genome-wide screening of plant proteins involved in pathogen perception. Two phage libraries for high-throughput selection were constructed from cDNA of plants infected with Pseudomonas aeruginosa PA14, or from combined samples of the virulent isolate DC3000 of Pseudomonas syringae pv. tomato and its avirulent variant avrRpt2. These three pathosystems represent different degrees in the specificity of plant-microbe interactions. Libraries cover up to 2×107 different plant transcripts that can be displayed as functional proteins on the surface of T7 bacteriophage. A number of these were selected in a bio-panning assay for binding to Pseudomonas cells. Among the selected clones we isolated the ethylene response factor ATERF-1, which was able to bind the three bacterial strains in competition assays. ATERF-1 was rapidly exported from the nucleus upon infiltration of either alive or heat-killed Pseudomonas. Moreover, aterf-1 mutants exhibited enhanced susceptibility to infection. These findings suggest that ATERF-1 contains a microbe-recognition domain with a role in plant defence. To identify other putative pathogen-binding proteins on a genome-wide scale, the copy number of selected-vs.-total clones was compared by hybridizing phage cDNAs with Arabidopsis microarrays. Microarray analysis revealed a set of 472 candidates with significant fold change. Within this set defence-related genes, including well-known targets of bacterial effectors, are over-represented. Other genes non-previously related to defence can be associated through this study with general or strain-specific recognition of Pseudomonas.

Transcriptomics studies are available to evaluate the potential toxicity of nanomaterials in plants, and many highlight their effect on stress-responsive genes. However, a comparative analysis of overall expression changes suggests a low impact on the transcriptome. Environmental challenges like pathogens, saline, or drought stress induce stronger transcriptional responses than nanoparticles. Clearly, plants did not have the chance to evolve specific gene regulation in response to novel nanomaterials; but they use common regulatory circuits with other stress responses. A shared effect with abiotic stress is the inhibition of genes for root development and pathogen response. Other works are reviewed here, which also converge on these results.