Cloning and Characterization of a General Amino Acid Control Transcriptional Activator from the Chestnut Blight FungusCryphonectria parasitica

Abstract

We have cloned and characterized a homologue of theNeurospora crassageneral amino acid control genecpc-1from the chestnut blight fungusCryphonectria parasitica.The deduced amino acid sequence ofC. parasiticaCPC1 (cpCPC1) contains regions with significant homology to the transcriptional activation, DNA binding, and dimerization domains previously defined forN. crassaCPC1 (ncCPC1) and the equivalent “b-ZIP” transcription factor fromSaccharomyces cerevisiae,GCN4 (scGCN4). Treatment ofC. parasiticawith low levels of the protein synthesis inhibitor cycloheximide causedcpc-1transcript levels to undergo a rapid, transient increase similar to that reported for the mammalian b-ZIP transactivators, c-Jun and c-Fos. Northern analysis also revealed that amino acid starvation ofC. parasiticaelicits an increase incpc-1transcript levels. Hypovirus infection did not affect this increase, although transcript accumulation for several amino acid biosynthetic genes was slightly diminished in the hypovirus-containing strain. Recombinant cpCPC1 specifically bound to the consensus DNA binding element (AP-1), 5′-A/GTGACTCAT-3′, also located upstream of theC. parasitica cpc-1coding region. Constitutive transgenic expression of a DNA binding defective cpCPC1 mutant impaired the ability ofC. parasiticato adjust to amino acid starvation. Moreover, these transformants showed reduced ability to grow on host chestnut tissue. Our results define a general amino acid control transactivator in a plant pathogenic fungus and suggest that functional modulation of this factor can influence fungal virulence.