Carbonic anhydrase and iron acquisition system in the human pathogenic fungus Aspergillus fumigatus

• Grant number: 11/20808-7
• Support Opportunities: Regular Research Grants
• Duration: February 01, 2012 - March 31, 2014
• Field of knowledge: Biological Sciences - Genetics - Molecular Genetics and Genetics of Microorganisms
• Principal Investigator: Márcia Eliana da Silva Ferreira
• Grantee: Márcia Eliana da Silva Ferreira
• Host Institution: Faculdade de Ciências Farmacêuticas de Ribeirão Preto (FCFRP). Universidade de São Paulo (USP). Ribeirão Preto , SP, Brazil

Abstract

Aspergillus fumigatus is a saprophyte mold of great importance in the environment by recycling carbon and nitrogen. However, in the last 20 years, this ascomycete is no longer just a minor saprophyte mold to become one of the major human pathogens. During the infection process, the fungus A. fumigatus is facing a dramatic change in the CO2 concentrations (0.036% in their natural habitat to 1 to 5% in the infected host). Previous studies have shown that deletion of each of the four A. fumigatus carbonic anhydrase did not affect the virulence of this fungus. Despite the double mutant DeltacafADeltacafB strain has not changed the fungus virulence profile, this strain was unable to grow in CO2 environment concentration (0.036%), an effect that is reversed by high concentrations of CO2 (5% - high-CO2-requiring (HCR) phenotype). In addition to this, transcriptional analysis of double mutant DeltacafADeltacafB in different concentrations of CO2 suggested an important role of iron for growth this strain in 5% CO2. In order to verify if there is a direct correlation between the iron acquisition system and the A. fumigatus carbonic anhydrase and for a better characterization of carbonic anhydrase system in this fungus, this project aims to: 1) evaluate the growth of the double mutant DeltacafADeltacafB strain in different concentrations of iron, in 0.036% and 5% CO2; 2) validate the results previously found in the "microarrays" analysis (FAPESP project nº 2008/06493-0), by determination of the siderophores concentration delivered in the media culture by the fungus, and by the determination of the RNA expression level of some relevant genes; 3) delete the sidA gene in the DeltacafADeltacafB strain and functional characterize this mutant, 4) evaluate the effect of different carbonic anhydrase inhibitors in the growth and HCR phenotype of wild-type strain DeltaakuBku80; 5) perform the global analysis of gene expression, by "microarray", of wild-type strain exposed to different concentrations of a specific carbonic anhydrase inhibitor (the one with the most promising up front to phenotypic analysis). This project aims to elucidate the carbonic anhydrase system and its associated metabolic pathways in the A. fumigatus, favoring a better understanding of the involvement of this enzyme in the virulence and pathogenicity mechanisms of this human pathogenic fungus. (AU)