The Transcription Factor BcLTF1 Regulates Virulence and Light Responses in the Necrotrophic Plant Pathogen Botrytis cinerea

Abstract

Botrytis cinerea is the causal agent of gray mold diseases in a range of dicotyledonous plant species. The fungus can reproduce asexually by forming macroconidia for dispersal and sclerotia for survival; the latter also participate in sexual reproduction by bearing the apothecia after fertilization by microconidia. Light induces the differentiation of conidia and apothecia, while sclerotia are exclusively formed in the absence of light. The relevance of light for virulence of the fungus is not obvious, but infections are observed under natural illumination as well as in constant darkness. By a random mutagenesis approach, we identified a novel virulence-related gene encoding a GATA transcription factor (BcLTF1 for light-responsive TF1) with characterized homologues in Aspergillus nidulans (NsdD) and Neurospora crassa (SUB-1). By deletion and over-expression of bcltf1, we confirmed the predicted role of the transcription factor in virulence, and discovered furthermore its functions in regulation of light-dependent differentiation, the equilibrium between production and scavenging of reactive oxygen species (ROS), and secondary metabolism. Microarray analyses revealed 293 light-responsive genes, and that the expression levels of the majority of these genes (66%) are modulated by BcLTF1. In addition, the deletion of bcltf1 affects the expression of 1,539 genes irrespective of the light conditions, including the overexpression of known and so far uncharacterized secondary metabolism-related genes. Increased expression of genes encoding alternative respiration enzymes, such as the alternative oxidase (AOX), suggest a mitochondrial dysfunction in the absence of bcltf1. The hypersensitivity of Δbctlf1 mutants to exogenously applied oxidative stress - even in the absence of light - and the restoration of virulence and growth rates in continuous light by antioxidants, indicate that BcLTF1 is required to cope with oxidative stress that is caused either by exposure to light or arising during host infection.

Chasing stress signals – Exposure to extracellular stimuli differentially affects the redox state of cell compartments in the wild type and signaling mutants of Botrytis cinerea

“…The transcription factor has an exceptional function in B. cinerea; it is, for instance, required for growth in (excessive) light, to cope with oxidative stress and for virulence. Moreover, the deletion mutants produce more ROS and some of the phenotypes could be restored by applying antioxidants (Schumacher et al, 2014). As previously shown, the cytosolic glutathione pool is oxidized upon exposure to H 2 O 2 in a wild-type-like manner; a similar pattern was found in this study for the redox state of the ER (Fig.…”

“…The changes lead to different spectral properties with excitation wavelengths of 395 and 488 nm facilitating quantitative ratiometric imaging of redox dynamics (Dooley et al, 2004;Jiang et al, 2006;Morgan et al, 2011;Schwarzländer et al, 2008;Yu et al, 2009). The biosensor (cyt-roGFP2) was successfully expressed under control of a constitutive promoter (PoliC) in B. cinerea, and was applied as a tool to compare the cytosolic redox states in wild-type and mutant strains (Heller et al, 2012a(Heller et al, , 2012bSchumacher et al, 2014;Viefhues et al, 2014).…”

“…Hence, the light-responsive transcription factor BcLtf1 plays an essential role in maintaining the ROS homeostasis and mediating the tolerance to exogenous oxidative stress; moreover, it modulates the light responses and the production of secondary metabolites (melanin, botrydial and botcinic acid) in B. cinerea (Schumacher et al, 2014).…”

Unraveling the Function of the Response Regulator BcSkn7 in the Stress Signaling Network of Botrytis cinerea

“…As the mycelium of the mutant looked paler than that of the WT, pigmentation defects of the bcskn7 mutant were assumed. Therefore, the expression of genes involved in melanin biosynthesis (bcpks13 and bcscd1) was examined (50,51). The strains were grown for 4 days in constant light (Fig.…”

“…2D to F), and the overexpression of BcSkn7 in B05.10 resulted in enhanced sporulation, even in constant darkness. Normally, the induction of macroconidia is produced by light and involves the function of light-dependent TFs, such as BcLtf1 (51). It follows, therefore, that overexpression of BcSkn7 leads to an altered light response, maybe due to deregulation of photoreceptor genes.…”