da Silva Ferreira Márcia Eliana, Malavazi Iran, Savoldi Marcela, Brakhage Axel A, Goldman Maria Helena S, Kim H Stanley, Nierman William C, Goldman Gustavo H
Departamento de Ciências Farmacêuticas, Faculdade de Ciências Farmacêuticas de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, Brazil.
Curr Genet. 2006 Jul;50(1):32-44. doi: 10.1007/s00294-006-0073-2. Epub 2006 Apr 19.
For a comprehensive evaluation of genes that have their expression modulated during exposure of the mycelia to voriconazole, we performed a large-scale analysis of gene expression in Aspergillus fumigatus using a microarray hybridization approach. By comparing the expression of genes between the reference time and after addition of voriconazole (30, 60, 120, and 240 min), we identified 2,271 genes differentially expressed in the wild-type strain. To validate the expression of some of these genes during exposure to voriconazole, we analyzed 13 genes showing higher expression in the presence of voriconazole by real-time RT-PCR. Although the magnitudes of induction differed between the two experimental systems, in about 85% of the cases they were in good agreement with the microarray data. To our knowledge this is the first study of microarray hybridization analysis for a filamentous fungus exposed to an antifungal agent. In our study, we have observed: (i) a decreased mRNA expression of various ergosterol biosynthesis genes; (ii) increased mRNA levels of genes involved in a variety of cell functions, such as transporters, transcription factors, proteins involved in cell metabolism, and hypothetical proteins; and (iii) the involvement of the cyclic AMP-protein kinase signaling pathway in the increased mRNA expression of several of these genes.
为全面评估在菌丝体暴露于伏立康唑期间表达受到调控的基因，我们采用微阵列杂交方法对烟曲霉中的基因表达进行了大规模分析。通过比较参考时间与添加伏立康唑后（30、60、120和240分钟）基因的表达情况，我们在野生型菌株中鉴定出2271个差异表达基因。为验证其中一些基因在暴露于伏立康唑期间的表达情况，我们通过实时逆转录聚合酶链反应分析了13个在伏立康唑存在时表达较高的基因。尽管两个实验系统中的诱导程度有所不同，但在约85%的情况下，它们与微阵列数据高度一致。据我们所知，这是首次对丝状真菌暴露于抗真菌剂进行微阵列杂交分析的研究。在我们的研究中，我们观察到：（i）各种麦角固醇生物合成基因的mRNA表达降低；（ii）参与多种细胞功能的基因的mRNA水平升高，如转运蛋白、转录因子、参与细胞代谢的蛋白质以及假定蛋白；（iii）环磷酸腺苷 - 蛋白激酶信号通路参与了其中一些基因mRNA表达的增加。
