Comparison of Gene Expression Profiles of Candida albicans Azole-Resistant Clinical Isolates and Laboratory Strains Exposed to Drugs Inducing Multidrug Transporters

ABSTRACT Azole resistance in Candida albicans can be due to upregulation of multidrug transporters belonging to ABC (ATP-binding cassette) transporters (CDR1 and CDR2) or major facilitators (CaMDR1). Upregulation of these genes can also be achieved by exposure to fluphenazine, resulting in specific upregulation of CDR1 and CDR2 and by exposure to benomyl, resulting in specific CaMDR1 upregulation. In this study, these two different states of gene upregulation were used to determine coregulated genes that often share similar functions or similar regulatory regions. The transcript profiles of a laboratory strain exposed to these drugs were therefore determined and compared with those of two matched pairs of azole-susceptible and -resistant strains expressing CDR1 and CDR2 (CDR strains) or CaMDR1 (MDR isolates). The results obtained revealed that, among 42 commonly regulated genes (8.6% of all regulated genes) between fluphenazine-exposed cells and CDR isolates, the most upregulated were CDR1 and CDR2 as expected, but also IFU5, RTA3 (which encodes putative membrane proteins), HSP12 (which encodes heat shock protein), and IPF4065 (which is potentially involved in stress response). Interestingly, all but HSP12 and IPF4065 contain a putative cis-acting drug responsive element in their promoters. Among the 57 genes (11.5% of all regulated genes) commonly regulated between benomyl-exposed cells and MDR isolates, the most upregulated were CaMDR1 as expected but also genes with oxido-reductive functions such as IFD genes, IPF5987, GRP2 (all belonging to the aldo-keto reductase family), IPF7817 [NAD(P)H oxido-reductase], and IPF17186. Taken together, these results show that in vitro drug-induced gene expression only partially mimics expression profiles observed in azole-resistant clinical strains. Upregulated genes in both drug-exposed conditions and clinical strains are drug resistance genes but also genes that could be activated under cell damage conditions.

[1]  A. Feinberg,et al.  A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. , 1983, Analytical biochemistry.

[2]  D. Irwin,et al.  Isogenic strain construction and gene mapping in Candida albicans. , 1993, Genetics.

[3]  A. Brown,et al.  Expression of seven members of the gene family encoding secretory aspartyl proteinases in Candida albicans , 1994, Molecular microbiology.

[4]  K. Kuchler,et al.  Mechanisms of resistance to azole antifungal agents in Candida albicans isolates from AIDS patients involve specific multidrug transporters , 1995, Antimicrobial agents and chemotherapy.

[5]  J. Rex,et al.  Resistance of Candida species to fluconazole , 1995, Antimicrobial agents and chemotherapy.

[6]  J. Goudet,et al.  Typing Candida albicans oral isolates from human immunodeficiency virus-infected patients by multilocus enzyme electrophoresis and DNA fingerprinting , 1996, Journal of clinical microbiology.

[7]  D. Snydman,et al.  The changing face of candidemia: emergence of non-Candida albicans species and antifungal resistance. , 1996, The American journal of medicine.

[8]  T. C. White,et al.  Increased mRNA levels of ERG16, CDR, and MDR1 correlate with increases in azole resistance in Candida albicans isolates from a patient infected with human immunodeficiency virus , 1997, Antimicrobial agents and chemotherapy.

[9]  K. Struhl,et al.  Yap, a novel family of eight bZIP proteins in Saccharomyces cerevisiae with distinct biological functions , 1997, Molecular and cellular biology.

[10]  D. Sanglard,et al.  Cloning of Candida albicans genes conferring resistance to azole antifungal agents: characterization of CDR2, a new multidrug ABC transporter gene. , 1997, Microbiology.

[11]  T. C. White,et al.  The presence of an R467K amino acid substitution and loss of allelic variation correlate with an azole-resistant lanosterol 14alpha demethylase in Candida albicans , 1997, Antimicrobial agents and chemotherapy.

[12]  L. Kitchen,et al.  Diethylcarbamazine-related antimicrobial activity in Mycobacterium tuberculosis-infected blood. , 1998, The Journal of antimicrobial chemotherapy.

[13]  V. Gupta,et al.  Identification of polymorphic mutant alleles of CaMDR1, a major facilitator of Candida albicans which confers multidrug resistance, and its in vitro transcriptional activation , 1998, Current Genetics.

[14]  Dominique Sanglard,et al.  Amino Acid Substitutions in the Cytochrome P-450 Lanosterol 14α-Demethylase (CYP51A1) from Azole-Resistant Candida albicans Clinical Isolates Contribute to Resistance to Azole Antifungal Agents , 1998, Antimicrobial Agents and Chemotherapy.

[15]  M. Raymond,et al.  The bZip Transcription Factor Cap1p Is Involved in Multidrug Resistance and Oxidative Stress Response inCandida albicans , 1999, Journal of bacteriology.

[16]  D. Sanglard,et al.  The ATP Binding Cassette Transporter GeneCgCDR1 from Candida glabrata Is Involved in the Resistance of Clinical Isolates to Azole Antifungal Agents , 1999, Antimicrobial Agents and Chemotherapy.

[17]  D. Botstein,et al.  Cluster analysis and display of genome-wide expression patterns. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[18]  A. Goffeau,et al.  Genome microarray analysis of transcriptional activation in multidrug resistance yeast mutants , 2000, FEBS letters.

[19]  D. Sanglard,et al.  A novel multidrug efflux transporter gene of the major facilitator superfamily from Candida albicans (FLU1) conferring resistance to fluconazole. , 2000, Microbiology.

[20]  J. Fostel,et al.  Genome-Wide Expression Patterns inSaccharomyces cerevisiae: Comparison of Drug Treatments and Genetic Alterations Affecting Biosynthesis of Ergosterol , 2000, Antimicrobial Agents and Chemotherapy.

[21]  L. Cowen,et al.  Evolution of Drug Resistance in Experimental Populations of Candida albicans , 2000, Journal of bacteriology.

[22]  D. Sanglard,et al.  Analysis of the oxidative stress regulation of the Candida albicans transcription factor, Cap1p , 2000, Molecular microbiology.

[23]  G. Köhler,et al.  Activation of the Multiple Drug Resistance GeneMDR1 in Fluconazole-Resistant, Clinical Candida albicans Strains Is Caused by Mutations in atrans-Regulatory Factor , 2000, Journal of bacteriology.

[24]  Xiao-Jun Ma,et al.  Genomic Profiling of the Response of Candida albicans to Itraconazole Treatment Using a DNA Microarray , 2001, Antimicrobial Agents and Chemotherapy.

[25]  Yoshiharu Inoue,et al.  Regulation of the Yeast Yap1p Nuclear Export Signal Is Mediated by Redox Signal-Induced Reversible Disulfide Bond Formation , 2001, Molecular and Cellular Biology.

[26]  D. Singleton,et al.  Cloning and Analysis of a Candida albicans Gene That Affects Cell Surface Hydrophobicity , 2001, Journal of bacteriology.

[27]  J. Lopez-Ribot,et al.  Prevalence of Molecular Mechanisms of Resistance to Azole Antifungal Agents in Candida albicans Strains Displaying High-Level Fluconazole Resistance Isolated from Human Immunodeficiency Virus-Infected Patients , 2001, Antimicrobial Agents and Chemotherapy.

[28]  D. Sanglard,et al.  Role of ATP-Binding-Cassette Transporter Genes in High-Frequency Acquisition of Resistance to Azole Antifungals in Candida glabrata , 2001, Antimicrobial Agents and Chemotherapy.

[29]  David Y. Thomas,et al.  Population genomics of drug resistance in Candida albicans , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[30]  D. Sanglard,et al.  A common drug‐responsive element mediates the upregulation of the Candida albicans ABC transporters CDR1 and CDR2, two genes involved in antifungal drug resistance , 2002, Molecular microbiology.

[31]  D. Botstein,et al.  Genome-wide Analysis of Gene Expression Regulated by the Calcineurin/Crz1p Signaling Pathway in Saccharomyces cerevisiae * , 2002, The Journal of Biological Chemistry.

[32]  K. Barker,et al.  Genome-Wide Expression Profile Analysis Reveals Coordinately Regulated Genes Associated with Stepwise Acquisition of Azole Resistance in Candida albicans Clinical Isolates , 2003, Antimicrobial Agents and Chemotherapy.

[33]  M. Whiteway,et al.  Stress-induced gene expression in Candida albicans: absence of a general stress response. , 2003, Molecular biology of the cell.

[34]  J. Cleary,et al.  Genome-wide Expression Profiling of the Response to Polyene, Pyrimidine, Azole, and Echinocandin Antifungal Agents in Saccharomyces cerevisiae* , 2003, Journal of Biological Chemistry.