Determining Aspergillus fumigatus transcription factor expression and function during invasion of the mammalian lung

Abstract

To gain a better understanding of the transcriptional response ofAspergillus fumigatusduring invasive pulmonary infection, we used a NanoString nCounter to assess the transcript levels of 467A.fumigatusgenes during growth in the lungs of immunosuppressed mice. These genes included ones known to respond to diverse environmental conditions and those encoding most transcription factors in theA.fumigatusgenome. We found that invasive growthin vivoinduces a unique transcriptional profile as the organism responds to nutrient limitation and attack by host phagocytes. Thisin vivotranscriptional response is largely mimicked byin vitrogrowth inAspergillusminimal medium that is deficient in nitrogen, iron, and/or zinc. From the transcriptional profiling data, we selected 9 transcription factor genes that were either highly expressed or strongly up-regulated duringin vivogrowth. Deletion mutants were constructed for each of these genes and assessed for virulence in mice. Two transcription factor genes were found to be required for maximal virulence. One wasrlmA, which is required for the organism to achieve maximal fungal burden in the lung. The other wassltA, which regulates of the expression of multiple secondary metabolite gene clusters and mycotoxin genes independently oflaeA. Using deletion and overexpression mutants, we determined that the attenuated virulence of the ΔsltAmutant is due in part to decreased expressionaspf1, which specifies a ribotoxin, but is not mediated by reduced expression of the fumigaclavine gene cluster or the fumagillin-pseruotin supercluster. Thus,in vivotranscriptional profiling focused on transcription factors genes provides a facile approach to identifying novel virulence regulators.