Homeobox transcription factor CoHox3 is essential for appressorium formation in the plant pathogenic fungus Colletotrichum orbiculare

The plant pathogenic fungus Colletotrichum orbiculare, which is the causal agent of cucumber anthracnose disease, forms dome-shaped, melanized appressoria as a host invasion structure. We found 10 conserved homeobox transcription factors based on genome analysis. In this study, we functionally analyzed one of the C. orbiculare homeobox gene named CoHox3 (Colletotrichum orbiculare Homeobox transcription factor 3) during the infection process. CoHox3 knockout mutants were non-pathogenic on host leaves (inoculated with 1.0 × 104 conidia/mL). Conidia of CoHox3 mutants germinated and/or formed lightly pigmented structures, but did not form mature appressoria on glass slides, artificial membranes, or host leaves. Only 0.3-0.6% of germinated conidia of CoHox3 mutants formed infection hyphae on host leaves. Green fluorescent protein-based expression experiments showed that germinated conidia of the CoHox3 mutant did not express CoGAS1 gene, an appressorium-specific gene, although wild-type strain 104-T expressed the CoGAS1 gene during appressorium formation. When conidia of the CoHox3 mutant was incubated on glass slides, nuclear division was significantly later than that of wild-type, which formed appressorium. Thus, homeobox transcription factor CoHox3 is required to form normal appressoria.