Essential Gene Identification and Drug Target Prioritization in Aspergillus Fumigatus

Overview

Journal PLoS Pathog

Specialty Microbiology

Date 2007 Mar 14

PMID 17352532

Citations 125

Authors

Wenqi Hu

Susan Sillaots

Sebastien Lemieux

John Davison

Sarah Kauffman

Anouk Breton

Annie Linteau

Chunlin Xin

Joel Bowman

Jeff Becker

Bo Jiang

Terry Roemer

Affiliations

Soon will be listed here.

Abstract

Aspergillus fumigatus is the most prevalent airborne filamentous fungal pathogen in humans, causing severe and often fatal invasive infections in immunocompromised patients. Currently available antifungal drugs to treat invasive aspergillosis have limited modes of action, and few are safe and effective. To identify and prioritize antifungal drug targets, we have developed a conditional promoter replacement (CPR) strategy using the nitrogen-regulated A. fumigatus NiiA promoter (pNiiA). The gene essentiality for 35 A. fumigatus genes was directly demonstrated by this pNiiA-CPR strategy from a set of 54 genes representing broad biological functions whose orthologs are confirmed to be essential for growth in Candida albicans and Saccharomyces cerevisiae. Extending this approach, we show that the ERG11 gene family (ERG11A and ERG11B) is essential in A. fumigatus despite neither member being essential individually. In addition, we demonstrate the pNiiA-CPR strategy is suitable for in vivo phenotypic analyses, as a number of conditional mutants, including an ERG11 double mutant (erg11BDelta, pNiiA-ERG11A), failed to establish a terminal infection in an immunocompromised mouse model of systemic aspergillosis. Collectively, the pNiiA-CPR strategy enables a rapid and reliable means to directly identify, phenotypically characterize, and facilitate target-based whole cell assays to screen A. fumigatus essential genes for cognate antifungal inhibitors.

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References

Muro-Pastor M, Gonzalez R, Strauss J, Narendja F, Scazzocchio C . The GATA factor AreA is essential for chromatin remodelling in a eukaryotic bidirectional promoter. EMBO J. 1999; 18(6):1584-97. PMC: 1171246. DOI: 10.1093/emboj/18.6.1584. View

dEnfert C, Diaquin M, Delit A, Wuscher N, Debeaupuis J, Huerre M . Attenuated virulence of uridine-uracil auxotrophs of Aspergillus fumigatus. Infect Immun. 1996; 64(10):4401-5. PMC: 174389. DOI: 10.1128/iai.64.10.4401-4405.1996. View

Mellado E, Garcia-Effron G, Buitrago M, Alcazar-Fuoli L, Cuenca-Estrella M, Rodriguez-Tudela J . Targeted gene disruption of the 14-alpha sterol demethylase (cyp51A) in Aspergillus fumigatus and its role in azole drug susceptibility. Antimicrob Agents Chemother. 2005; 49(6):2536-8. PMC: 1140498. DOI: 10.1128/AAC.49.6.2536-2538.2005. View

De Backer M, Nelissen B, Logghe M, Viaene J, Loonen I, Vandoninck S . An antisense-based functional genomics approach for identification of genes critical for growth of Candida albicans. Nat Biotechnol. 2001; 19(3):235-41. DOI: 10.1038/85677. View

Firon A, Beauvais A, Latge J, Couve E, dEnfert C . Characterization of essential genes by parasexual genetics in the human fungal pathogen Aspergillus fumigatus: impact of genomic rearrangements associated with electroporation of DNA. Genetics. 2002; 161(3):1077-87. PMC: 1462181. DOI: 10.1093/genetics/161.3.1077. View

Amaar Y, Moore M . Mapping of the nitrate-assimilation gene cluster (crnA-niiA-niaD) and characterization of the nitrite reductase gene (niiA) in the opportunistic fungal pathogen Aspergillus fumigatus. Curr Genet. 1998; 33(3):206-15. DOI: 10.1007/s002940050328. View

Verdoes J, Punt P, van der Berg P, Debets F, Stouthamer A, van den Hondel C . Characterization of an efficient gene cloning strategy for Aspergillus niger based on an autonomously replicating plasmid: cloning of the nicB gene of A. niger. Gene. 1994; 146(2):159-65. DOI: 10.1016/0378-1119(94)90288-7. View

Nascimento A, Goldman G, Park S, Marras S, Delmas G, Oza U . Multiple resistance mechanisms among Aspergillus fumigatus mutants with high-level resistance to itraconazole. Antimicrob Agents Chemother. 2003; 47(5):1719-26. PMC: 153329. DOI: 10.1128/AAC.47.5.1719-1726.2003. View

Yang B, Zhao D, Solenov E, Verkman A . Evidence from knockout mice against physiologically significant aquaporin 8-facilitated ammonia transport. Am J Physiol Cell Physiol. 2006; 291(3):C417-23. DOI: 10.1152/ajpcell.00057.2006. View

10.

Tsai H, Wheeler M, Chang Y, Kwon-Chung K . A developmentally regulated gene cluster involved in conidial pigment biosynthesis in Aspergillus fumigatus. J Bacteriol. 1999; 181(20):6469-77. PMC: 103784. DOI: 10.1128/JB.181.20.6469-6477.1999. View

11.

Bruno V, Kalachikov S, Subaran R, Nobile C, Kyratsous C, Mitchell A . Control of the C. albicans cell wall damage response by transcriptional regulator Cas5. PLoS Pathog. 2006; 2(3):e21. PMC: 1401495. DOI: 10.1371/journal.ppat.0020021. View

12.

Galagan J, Calvo S, Cuomo C, Ma L, Wortman J, Batzoglou S . Sequencing of Aspergillus nidulans and comparative analysis with A. fumigatus and A. oryzae. Nature. 2005; 438(7071):1105-15. DOI: 10.1038/nature04341. View

13.

Latge J . The pathobiology of Aspergillus fumigatus. Trends Microbiol. 2001; 9(8):382-9. DOI: 10.1016/s0966-842x(01)02104-7. View

14.

Cove D . The induction and repression of nitrate reductase in the fungus Aspergillus nidulans. Biochim Biophys Acta. 1966; 113(1):51-6. DOI: 10.1016/s0926-6593(66)80120-0. View

15.

Beauvais A, Latge J . Membrane and cell wall targets in Aspergillus fumigatus. Drug Resist Updat. 2001; 4(1):38-49. DOI: 10.1054/drup.2001.0185. View

16.

Firon A, Villalba F, Beffa R, dEnfert C . Identification of essential genes in the human fungal pathogen Aspergillus fumigatus by transposon mutagenesis. Eukaryot Cell. 2003; 2(2):247-55. PMC: 154855. DOI: 10.1128/EC.2.2.247-255.2003. View

17.

Nakayama H, Nakayama N, Arisawa M, Aoki Y . In vitro and in vivo effects of 14alpha-demethylase (ERG11) depletion in Candida glabrata. Antimicrob Agents Chemother. 2001; 45(11):3037-45. PMC: 90779. DOI: 10.1128/AAC.45.11.3037-3045.2001. View

18.

KURTZ M, Heath I, Marrinan J, Dreikorn S, Onishi J, Douglas C . Morphological effects of lipopeptides against Aspergillus fumigatus correlate with activities against (1,3)-beta-D-glucan synthase. Antimicrob Agents Chemother. 1994; 38(7):1480-9. PMC: 284580. DOI: 10.1128/AAC.38.7.1480. View

19.

Romero B, Turner G, Olivas I, Laborda F, De Lucas J . The Aspergillus nidulans alcA promoter drives tightly regulated conditional gene expression in Aspergillus fumigatus permitting validation of essential genes in this human pathogen. Fungal Genet Biol. 2003; 40(2):103-14. DOI: 10.1016/s1087-1845(03)00090-2. View

20.

da Silva Ferreira M, Kress M, Savoldi M, Goldman M, Hartl A, Heinekamp T . The akuB(KU80) mutant deficient for nonhomologous end joining is a powerful tool for analyzing pathogenicity in Aspergillus fumigatus. Eukaryot Cell. 2006; 5(1):207-11. PMC: 1360264. DOI: 10.1128/EC.5.1.207-211.2006. View