Genetic and Proteomic Analysis of Fungal Virulence
This chapter explores the genetic and proteomic approaches that are now feasible for many fungal systems. Post-genomic approaches to the analysis of biological function, networks, and processes often include techniques that permit global analysis of gene expression at the protein level. Analysis of protein expression during modulation of external conditions or during particular developmental states can provide useful clues about which genes might be important for a particular function. These proteomic approaches are complementary to techniques, such as serial analysis of gene expression (SAGE) and microarrays, that measure gene expression at the RNA level. Signature-tagged mutagenesis (STM) was also used to identify Cryptococcus neoformans var. grubii mutants with altered virulence in a mouse model of disseminated disease. Restriction enzyme-mediated insertion has also been used with fungi to promote relatively random insertions in fungal genomes. It has been commonly used in phytopathogens, but the only published studies in a human pathogen are those done with C. albicans and Aspergillus fumigatus. Proteomic analysis of the C. albicans response to several antifungal drugs showed that inhibition of specific targets resulted in a unique profile. Mechanisms of antifungal resistance can be elucidated by identification of proteins induced by treatment with antifungal agents. To determine mechanisms of fungal virulence, it is important to study the expression of proteins during the interaction of a pathogen with host cells, either in vitro or in vivo.
