Botrytis cinerea is a phytopathogen infecting a broad range of plants including strawberries and grapevine. During infection, the necrotrophic fungus is exposed to reactive oxygen species (ROS) released by the oxidative burst, an early plant defense reaction. B. cinerea even produces ROS itself in planta. This raises questions about how the pathogen senses and responds to the host defense reaction and which role the pathogen's oxidative stress response systems play. Functional analysis of the AP-1 transcription factor Bap1 confirmed its role as a pivotal regulator of ROS detoxification in vitro. Macroarray analysis revealed 99 H(2)O(2)-induced Bap1 target genes, of which several genes encoded ROS-degrading enzymes as well as other central components of the cellular redox status. However, Bap1 is not essential for pathogenesis. In planta analyses revealed that the Bap1 target genes were not expressed 2 days postinoculation although H(2)O(2) was detectable, proving that the normal virulence of the Deltabap1 mutant is not due to alternative regulation of the major oxidative stress response system in planta. The fungus obviously does not suffer H(2)O(2)-induced oxidative stress in planta, questioning classical ideas about the role of the oxidative burst in the infection process.

Atf1-homologous basic region leucine zipper (bZIP) transcription factors are known to act downstream of the stress-activated mitogen-activated protein kinase (SAPK) cascade in mammals, as well as in several fungi; they regulate the transcription of genes involved in the general stress response. Functional analyses of BcAtf1 in Botrytis cinerea show that it is also connected to the SAPK BcSak1, as it shares several stress response target genes. However, Δbcatf1 mutants are not hypersensitive to osmotic or oxidative stress, as are Δbcsak1 mutants. Both BcSak1 and BcAtf1 are regulators of differentiation, but their roles in these processes are almost inverse as, in contrast with Δbcsak1, Δbcatf1 mutants are significantly impaired in conidia production and do not differentiate any sclerotia. They show extremely vigorous growth in axenic culture, with a thick layer of aerial hyphae and a marked increase in colonization efficiency on different host plants and tissues. In addition, the sensitivity to cell wall-interfering agents is increased strongly. Microarray analyses demonstrate that the loss of BcAtf1 leads to extensive transcriptional changes: apart from stress response genes, the expression of a broad set of genes, probably involved in primary metabolism, cell wall synthesis and development, is affected by BcAtf1. Unexpectedly, BcAtf1 also controls secondary metabolism: the mutant contains significantly elevated levels of phytotoxins. These data indicate that BcAtf1 controls a diversity of cellular processes and has broad regulatory functions.

The thioredoxin system is of great importance for maintenance of cellular redox homeostasis. Here, we show that it has a severe influence on virulence of Botrytis cinerea, demonstrating that redox processes are important for host-pathogen interactions in this necrotrophic plant pathogen. The thioredoxin system is composed of two enzymes, the thioredoxin and the thioredoxin reductase. We identified two genes encoding for thioredoxins (bctrx1, bctrx2) and one gene encoding for a thioredoxin reductase (bctrr1) in the genome of B. cinerea. Knockout mutants of bctrx1 and bctrr1 were severely impaired in virulence and more sensitive to oxidative stress. Additionally, Δbctrr1 showed enhanced H2O2 production and retarded growth. To investigate the impact of the second major cellular redox system, glutathione, we generated deletion mutants for two glutathione reductase genes. The effects were only marginal; deletion of bcglr1 resulted in reduced germination and, correspondingly, to retarded infection as well as reduced growth on minimal medium, whereas bcglr2 deletion had no distinctive phenotype. In summary, we showed that the balanced redox status maintained by the thioredoxin system is essential for development and pathogenesis of B. cinerea, whereas the second major cellular redox system, the glutathione system, seems to have only minor impact on these processes.