Cation-Stress-Responsive Transcription Factors SItA and CrzA Regulate Morphogenetic Processes and Pathogenicity of Colletotrichurn gloeosporioides

Growth of Colletotrichum gloeosporioides in the presence of cation salts NaCI and KCI inhibited fungal growth and anthracnose symptom of colonization. Previous reports indicate that adaptation of Aspergillus nidulans to salt- and osmotic-stress conditions revealed the role of zinc-finger transcription factors SItA and CrzA in cation homeostasis. Homologs of A. nidulans SItA and CrzA were identified in C. gloeosporioides. The C. gloeosporioides CrzA homolog is a 682-amino acid protein, which contains a C2H2 zinc finger DNA-binding domain that is highly conserved among CrzA proteins from yeast and filamentous fungi. The C. gloeosporioides SItA homolog encodes a 775-amino acid protein with strong similarity to A. nidulans SItA and Trichoderma reeseiACE1, and highest conservation in the three zinc finger regions with almost no changes compared to ACE1 sequences. Knockout of C. gloeosporioides crzA (AcrzA) resulted in a phenotype with inhibited growth, sporulation, germination and appressorium formation, indicating the importance of this calciu006D-activated transcription factor in regulating these morphogenetic processes. In contrast, knockout of C. gloeosporioides sltA (AsItA) mainly inhibited appressorium formation. Both mutants had reduced pathogenicity on mango and avocado fruit. Inhibition of the different morphogenetic stages in the AcrzA mutant was accompanied by drastic inhibition of chitin synthase A and B and glucan synthase, which was partially restored with Ca2+ supplementation. Inhibition of appressorium formation in AsItA mutants was accompanied by downregulation of the MAP kinase pmkl and carnitine acetyl transferase (call), genes involved in appressorium formation and colonization, which was restored by Ca2+ supplementation. Furthermore, exposure of C. gloeosporioides AcrzA or AsItA mutants to cations such as Na+, K+ and Li+ at concentrations that the wild type C. gloeosporioides is not affected had further adverse morphogenetic effects on C. gloeosporioides which were partially or fully restored by Ca2+. Overall results suggest that both genes modulating alkali cation homeostasis have significant morphogenetic effects that reduce C. gloeosporioides colonization.