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ABSTRACTIn filamentous fungi, the GATA-type transcription factor AreA plays a major role in the transcriptional activation of genes needed to utilize poor nitrogen sources. In Fusarium fujikuroi, AreA also controls genes involved in the biosynthesis of gibberellins, a family of diterpenoid plant hormones. To identify more genes responding to nitrogen limitation or sufficiency in an AreA-dependent or -independent manner, we examined changes in gene expression of F. fujikuroiwild-type and ?areAstrains by use of a Fusarium verticillioidesmicroarray representing ~9,300 genes. Analysis of the array data revealed sets of genes significantly down- and upregulated in the areAmutant under both N starvation and N-sufficient conditions. Among the downregulated genes are those involved in nitrogen metabolism, e.g., those encoding glutamine synthetase and nitrogen permeases, but also those involved in secondary metabolism. Besides AreA-dependent genes, we found an even larger set of genes responding to N starvation and N-sufficient conditions in an AreA-independent manner. To study the impact of NMR on AreA activity, we examined the expression of several AreA target genes in the wild type and in areAand nmrdeletion and overexpression mutants. We show that NMR interacts with AreA as expected but affects gene expression only in early growth stages. This is the first report on genome-wide expression studies examining the influence of AreA on nitrogen-responsive gene expression in a genome-wide manner in filamentous fungi.

ABSTRACTIn Fusarium fujikuroi, the biosynthesis of gibberellins (GAs) and bikaverin is under control of AreA-mediated nitrogen metabolite repression. Thus far, the signaling components acting upstream of AreA and regulating its nuclear translocation are unknown. In Saccharomyces cerevisiae, the target of rapamycin (TOR) proteins, Tor1p and Tor2p, are key players of nutrient-mediated signal transduction to control cell growth. In filamentous fungi, probably only one TOR kinase-encoding gene exists. However, nothing is known about its function. Therefore, we investigated the role of TOR in the GA-producing fungus F. fujikuroiin order to determine whether TOR plays a role in nitrogen regulation, especially in the regulation of GA and bikaverin biosynthesis. We cloned and characterized the F. fujikuroi torgene. However, we were not able to create knockout mutants, suggesting that TOR is essential for viability. Inhibition of TOR by rapamycin affected the expression of AreA-controlled secondary metabolite genes for GA and bikaverin biosynthesis, as well as genes involved in transcriptional and translational regulation, ribosome biogenesis, and autophagy. Deletion of fpr1encoding the FKBP12-homologue confirmed that the effects of rapamycin are due to the specific inhibition of TOR. Interestingly, the expression of most of the TOR target genes has been previously shown to be also affected in the glutamine synthetase mutant, although in the opposite way. We demonstrate here for the first time in a filamentous fungus that the TOR kinase is involved in nitrogen regulation of secondary metabolism and that rapamycin affects also the expression of genes involved in translation control, ribosome biogenesis, carbon metabolism, and autophagy.