-
[show abstract]
[hide abstract]
ABSTRACT: Drug susceptible clinical isolates of Candida albicans frequently become highly tolerant to drugs during chemotherapy, with dreadful consequences to patient health. We used RNA sequencing (RNA-seq) to analyze the transcriptomes of a CDR (Candida Drug Resistance) strain and its isogenic drug sensitive counterpart.
RNA-seq unveiled differential expression of 228 genes including a) genes previously identified as involved in CDR, b) genes not previously associated to the CDR phenotype, and c) novel transcripts whose function as a gene is uncharacterized. In particular, we show for the first time that CDR acquisition is correlated with an overexpression of the transcription factor encoding gene CZF1. CZF1 null mutants were susceptible to many drugs, independently of known multidrug resistance mechanisms. We show that CZF1 acts as a repressor of β-glucan synthesis, thus negatively regulating cell wall integrity. Finally, our RNA-seq data allowed us to identify a new transcribed region, upstream of the TAC1 gene, which encodes the major CDR transcriptional regulator.
Our results open new perspectives of the role of Czf1 and of our understanding of the transcriptional and post-transcriptional mechanisms that lead to the acquisition of drug resistance in C. albicans, with potential for future improvements of therapeutic strategies.
BMC Genomics 08/2012; 13:396. · 4.07 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Gross chromosomal rearrangements (GCRs) such as aneuploidy are key factors in genome evolution as well as being common features of human cancer. Their role in tumour initiation and progression has not yet been completely elucidated and the effects of additional chromosomes in cancer cells are still unknown. Most previous studies in which Saccharomyces cerevisiae has been used as a model for cancer cells have been carried out in the haploid context. To obtain new insights on the role of ploidy, the cellular effects of GCRs were compared between the haploid and diploid contexts.
A total number of 21 haploid and diploid S. cerevisiae strains carrying various types of GCRs (aneuploidies, nonreciprocal translocations, segmental duplications and deletions) were studied with a view to determining the effects of ploidy on the cellular responses. Differences in colony and cell morphology as well as in the growth rates were observed between mutant and parental strains. These results suggest that cells are impaired physiologically in both contexts. We also investigated the variation in genomic expression in all the mutants. We observed that gene expression was significantly altered. The data obtained here clearly show that genes involved in energy metabolism, especially in the tricarboxylic acid cycle, are up-regulated in all these mutants. However, the genes involved in the composition of the ribosome or in RNA processing are down-regulated in diploids but up-regulated in haploids. Over-expression of genes involved in the regulation of the proteasome was found to occur only in haploid mutants.
The present comparisons between the cellular responses of strains carrying GCRs in different ploidy contexts bring to light two main findings. First, GCRs induce a general stress response in all studied mutants, regardless of their ploidy. Secondly, the ploidy context plays a crucial role in maintaining the stoichiometric balance of the proteins: the translation rates decrease in diploid strains, whereas the excess protein synthesized is degraded in haploids by proteasome activity.
BMC Genomics 06/2011; 12:331. · 4.07 Impact Factor
-
Mathieu Rougemaille,
Guennaelle Dieppois,
Elena Kisseleva-Romanova,
Rajani Kanth Gudipati, Sophie Lemoine,
Corinne Blugeon,
Jocelyne Boulay,
Torben Heick Jensen,
Françoise Stutz,
Frédéric Devaux,
Domenico Libri
[show abstract]
[hide abstract]
ABSTRACT: During transcription, proteins assemble sequentially with nascent RNA to generate a messenger ribonucleoprotein particle (mRNP). The THO complex and its associated Sub2p helicase are functionally implicated in both transcription and mRNP biogenesis but their precise function remains elusive. We show here that THO/Sub2p mutation leads to the accumulation of a stalled intermediate in mRNP biogenesis that contains nuclear pore components and polyadenylation factors in association with chromatin. Microarray analyses of genomic loci that are aberrantly docked to the nuclear pore in mutants allowed the identification of approximately 400 novel validated target genes that require THO /Sub2p for efficient expression. Our data strongly suggests that the THO complex/Sub2p function is required to coordinate events leading to the acquisition of export competence at a step that follows commitment to 3'-processing.
Cell 11/2008; 135(2):308-21. · 32.40 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Stress responses provide valuable models for deciphering the transcriptional networks controlling the adaptation of the cell to its environment. We analyzed the transcriptome response of yeast to toxic concentrations of selenite. We used gene network mapping tools to identify functional pathways and transcription factors involved in this response. We then used chromatin immunoprecipitation and knock-out experiments to investigate the role of some of these regulators and the regulatory connections between them.
Selenite rapidly activates a battery of transcriptional circuits, including iron deprivation, oxidative stress and protein degradation responses. The mRNA levels of several transcriptional regulators are themselves regulated. We demonstrate the existence of a positive transcriptional loop connecting the regulator of proteasome expression, Rpn4p, to the pleiotropic drug response factor, Pdr1p. We also provide evidence for the involvement of this regulatory module in the oxidative stress response controlled by the Yap1p transcription factor and its conservation in the pathogenic yeast C. glabrata. In addition, we show that the drug resistance regulator gene YRR1 and the iron homeostasis regulator gene AFT2 are both directly regulated by Yap1p.
This work depicted a highly interconnected and complex transcriptional network involved in the adaptation of yeast genome expression to the presence of selenite in its chemical environment. It revealed the transcriptional regulation of PDR1 by Rpn4p, proposed a new role for the pleiotropic drug resistance network in stress response and demonstrated a direct regulatory connection between oxidative stress response and iron homeostasis.
BMC Genomics 01/2008; 9:333. · 4.07 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The asymmetric localization of mRNA plays an important role in coordinating posttranscriptional events in eukaryotic cells. We investigated the peripheral mitochondrial localization of nuclear-encoded mRNAs (MLR) in various conditions in which the mRNA binding protein context and the translation efficiency were altered. We identified Puf3p, a Pumilio family RNA-binding protein, as the first trans-acting factor controlling the MLR phenomenon. This allowed the characterization of two classes of genes whose mRNAs are translated to the vicinity of mitochondria. Class I mRNAs (256 genes) have a Puf3p binding motif in their 3'UTR region and many of them have their MLR properties deeply affected by PUF3 deletion. Conversely, mutations in the Puf3p binding motif alter the mitochondrial localization of BCS1 mRNA. Class II mRNAs (224 genes) have no Puf3p binding site and their asymmetric localization is not affected by the absence of PUF3. In agreement with a co-translational import process, we observed that the presence of puromycin loosens the interactions between most of the MLR-mRNAs and mitochondria. Unexpectedly, cycloheximide, supposed to solidify translational complexes, turned out to destabilize a class of mRNA-mitochondria interactions. Classes I and II mRNAs, which are therefore transported to the mitochondria through different pathways, correlated with different functional modules. Indeed, Class I genes code principally for the assembly factors of respiratory chain complexes and the mitochondrial translation machinery (ribosomes and translation regulators). Class II genes encode proteins of the respiratory chain or proteins involved in metabolic pathways. Thus, MLR, which is intimately linked to translation control, and the activity of mRNA-binding proteins like Puf3p, may provide the conditions for a fine spatiotemporal control of mitochondrial protein import and mitochondrial protein complex assembly. This work therefore provides new openings for the global study of mitochondria biogenesis.
PLoS ONE 01/2008; 3(6):e2293. · 4.09 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The widespread pleiotropic drug resistance (PDR) phenomenon is well described as the long term selection of genetic variants expressing constitutively high levels of membrane transporters involved in drug efflux. However, the transcriptional cascades leading to the PDR phenotype in wild-type cells are largely unknown, and the first steps of this phenomenon are poorly understood. We investigated the transcriptional mechanisms underlying the establishment of an efficient PDR response in budding yeast. We show that within a few minutes of drug sensing yeast elicits an effective PDR response, involving tens of PDR genes. This early PDR response (ePDR) is highly dependent on the Pdr1p transcription factor, which is also one of the major genetic determinants of long term PDR acquisition. The activity of Pdr1p in early drug response is not drug-specific, as two chemically unrelated drugs, benomyl and fluphenazine, elicit identical, Pdr1p-dependent, ePDR patterns. Our data also demonstrate that Pdr1p is an original stress response factor, the DNA binding properties of which do not depend on the presence of drugs. Thus, Pdr1p is a promoter-resident regulator involved in both basal expression and rapid drug-dependent induction of PDR genes.
Journal of Biological Chemistry 03/2007; 282(7):5063-74. · 4.77 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Raw data normalization is a critical step in microarray data analysis because it directly affects data interpretation. Most of the normalization methods currently used are included in the R/BioConductor packages but it is often difficult to identify the most appropriate method. Furthermore, the use of R commands for functions and graphics can introduce mistakes that are difficult to trace. We present here a script written in R that provides a flexible means of access to and monitoring of data normalization for two-color microarrays. This script combines the power of BioConductor and R analysis functions and reduces the amount of R programming required.
Goulphar was developed in and runs using the R language and environment. It combines and extends functions found in BioConductor packages (limma and marray) to correct for dye biases and spatial artifacts. Goulphar provides a wide range of optional and customizable filters for excluding incorrect signals during the pre-processing step. It displays informative output plots, enabling the user to monitor the normalization process, and helps adapt the normalization method appropriately to the data. All these analyses and graphical outputs are presented in a single PDF report.
Goulphar provides simple, rapid access to the power of the R/BioConductor statistical analysis packages, with precise control and visualization of the results obtained. Complete documentation, examples and online forms for setting script parameters are available from http://transcriptome.ens.fr/goulphar/.
BMC Bioinformatics 02/2006; 7:467. · 2.75 Impact Factor
