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    Open AcceResearch articleStructure and properties of transcriptional networks driving selenite stress response in yeastsHlne Salin1,3, Vivienne Fardeau1,4, Eugenia Piccini1, Gaelle Lelandais1,5, Vronique Tanty2, Sophie Lemoine2, Claude Jacq1 and Frdric Devaux*1

    Address: 1Laboratoire de gntique molculaire, ENS/CNRS UMR 8541 46 rue d'Ulm, 75005 Paris, France, 2Plate-forme transcriptome, IFR 36, 46 rue d'Ulm, 75005 Paris, France, 3Musum national d'Histoire naturelle, 57 rue Cuvier 75005 PARIS, France, 4Commissariat l'Energie Atomique, Institut de Biologie et de Technologies de Saclay, 91191 Gif sur Yvette Cedex, France and 5Equipe de Bioinformatique Gnomique et Molculaire, INSERM UMR S726, Universit Paris 7, 2 place Jussieu, 75251 Paris cedex 05, France

    Email: Hlne Salin - [email protected]; Vivienne Fardeau - [email protected]; Eugenia Piccini - [email protected]; Gaelle Lelandais - [email protected]; Vronique Tanty - [email protected]; Sophie Lemoine - [email protected]; Claude Jacq - [email protected]; Frdric Devaux* - [email protected]

    * Corresponding author Equal contributors

    AbstractBackground: Stress responses provide valuable models for deciphering the transcriptionalnetworks controlling the adaptation of the cell to its environment. We analyzed the transcriptomeresponse of yeast to toxic concentrations of selenite. We used gene network mapping tools toidentify functional pathways and transcription factors involved in this response. We then usedchromatin immunoprecipitation and knock-out experiments to investigate the role of some ofthese regulators and the regulatory connections between them.

    Results: Selenite rapidly activates a battery of transcriptional circuits, including iron deprivation,oxidative stress and protein degradation responses. The mRNA levels of several transcriptionalregulators are themselves regulated. We demonstrate the existence of a positive transcriptionalloop connecting the regulator of proteasome expression, Rpn4p, to the pleiotropic drug responsefactor, Pdr1p. We also provide evidence for the involvement of this regulatory module in theoxidative stress response controlled by the Yap1p transcription factor and its conservation in thepathogenic yeast C. glabrata. In addition, we show that the drug resistance regulator gene YRR1 andthe iron homeostasis regulator gene AFT2 are both directly regulated by Yap1p.

    Conclusion: This work depicted a highly interconnected and complex transcriptional networkinvolved in the adaptation of yeast genome expression to the presence of selenite in its chemicalenvironment. It revealed the transcriptional regulation of PDR1 by Rpn4p, proposed a new role forthe pleiotropic drug resistance network in stress response and demonstrated a direct regulatoryconnection between oxidative stress response and iron homeostasis.

    BackgroundThe adaptation of genome expression to the chemicalenvironment is a complex but crucial challenge for all liv-

    ing cells. Functional genomics analyses in budding yeasthave shown that environmental stress responses mayinvolve rapid changes in the expression of up to 30% of

    Published: 15 July 2008

    BMC Genomics 2008, 9:333 doi:10.1186/1471-2164-9-333

    Received: 9 June 2008Accepted: 15 July 2008

    This article is available from: http://www.biomedcentral.com/1471-2164/9/333

    2008 Salin et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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    the genome. A common response to all stresses, namedESR (Environmental Stress Response), has beendescribed, which consists in the inhibition of the cytosolictranslation apparatus and the activation of the energystorage pathways [1]. However, pathways responding spe-cifically to the parameters of the environment also form akey part of the stress response. These pathways involvespecific transcriptional modules that rapidly sense theenvironment as a series of chemical and physical features(e.g. redox, pH, osmolarity, temperature, etc.) and acttogether to adapt genome expression to the specific natureof each stress [2]. For instance, at least eight different tran-scription factors act together to define the first-hourresponse of yeast cells to the toxic metalloid arsenite [3].These global and rapid responses are highly dynamic,involving sequential waves of gene activation and repres-sion [1,2,4]. This requires tight temporal coordinationbetween different transcriptional routes, which can beachieved in two complementary ways. First, the transcrip-tion factors involved in stress responses, despite respond-ing to different signals, may have overlapping sets oftargets [5]. Second, cross-regulation between transcriptionfactors may ensure the coordinated activation of differentpathways [6]. We focus here on the cross-talks betweenthree transcriptional modules responsible for the oxida-tive stress response, the ubiquitine-mediated protein deg-radation and the pleiotropic drug resistance, respectively.These cellular pathways exist in all species, from bacteriato mammals and plants. In S. cerevisiae, the oxidativestress response is controlled principally by the Yap1p tran-scription factor of the AP1-like leucine zipper family.Yap1p acts as a secondary sensor for oxidative molecules,and thus responds to a wide spectra of toxic compounds,such as hydrogen peroxide, metals and metalloids,organic nucleophilic molecules and internal metabolicoxidative stress due to the production of toxic by-productsduring glycolysis [3,4,7,8]. Yap1p recognizes YRE (Yap1presponse elements, 5'-TKACTMA-3') in the promoters ofgenes involved in redox homeostasis and in xenobioticexport at the plasma membrane. The proteasome isinvolved in both the degradation of damaged or aggre-gated proteins and in the post-translational regulation ofseveral biological processes, playing a key role in manystress responses [9]. Expression of the genes involved inproteasome biogenesis and activity, and in ubiquitin-dependent proteolysis, is controlled by the C2H2 zinc fin-ger protein Rpn4p, which recognizes the PACE (proteas-ome associated control element, 5'-GGTGGCAAA-3')sequence in the promoter of its target genes [3,10,11].Pleiotropic drug resistance involves the upregulation ofmembrane proteins involved in drug efflux. The corre-sponding genes are controlled principally by two Zn2Cys6Gal4p-like transcription factors: Pdr1p and Pdr3p. Thesetwo transcription factors have largely overlapping sets oftargets and recognize the same DNA motif (named PDRE,

    5'-TCCGYGGR-3'), but have different roles and regulatoryproperties [2,5,12-14]. The Yap1p and Rpn4p pathwaysare simultaneously involved in the yeast response to arse-nate [3]. Yap1p acts together with the Pdr1p/Pdr3p path-way to induce a drug specific response to the antifungaldrug benomyl [4]. No transcriptional regulation has beendescribed for PDR1, but YAP1, RPN4 and PDR3 areinduced by stress [3,5,6]. In this work, we used selenite asa model stress to investigate further the interactionsbetween these three transcriptional modules. Selenium isan essential oligoelement that replaces the sulfur atom ofsome methionine and cysteine in proteins involved in var-ious essential cell functions [15]. Selenium is also a prom-ising agent for cancer therapy and anti-aging treatments[16]. However, high doses of selenium are toxic to eukary-otic cells [17]. In yeast, selenium alters genome stability[18] and is detoxified in the vacuole after reacting withglutathione [19]. Yeast cells have a high level of seleniumtolerance, and yeast enriched in selenium have been usedin therapeutic trials [20]. We showed in this study thattoxic doses of selenite activated various yeast stressresponse pathways, including the proteasome, oxidativestress, iron homeostasis and general stress pathways. Wedemonstrated that, in these growth conditions, theexpression of PDR1 and RPN4 was coordinated through apositive transcriptional loop. This loop contributed to theoptimal Yap1p-dependent oxidative stress induction ofseveral genes encoding membrane proteins, includingFLR1, ATR1 and FRM2. This function seemed to be con-served in the pathogenic yeast species C. glabrata. Finally,our data provide evidence for direct transcriptional regu-lation of the iron homeostasis regulator Aft2p and of themultidrug resistance regulator Yrr1p by Yap1p, indicatinga broader role for this factor in coordination of the oxida-tive stress response.

    ResultsGene ontology mapping of the selenite responseWe analyzed the transcriptome of S. cerevisiae cells treatedwith 1 mM sodium selenite for 2, 5, 10, 20, 40, 60 and 80minutes. The corresponding cDNAs were competitivelyhybridized on DNA microarrays, with cDNAs obtainedfrom cells mock-treated for an equivalent period. Eachexperiment was carried out four times, with independentbiological samples. We used SAM to evaluate the signifi-cance of variations in expression of each gene [21]. Thedose of selenite used was the lowest dose that significantlyaltered cell survival in our preliminary growth assays (datanot shown). This toxic dose of selenite induced largechanges in the transcriptome of the cells (see additionalfile 1). In our data set, about 30% of the yeast ORFs dis

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