Use of an isocitrate lyase promoter–GFP fusion to monitor carbon metabolism of the plant pathogen Tapesia yallundae during infection of wheat

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Abstract
Green fluorescent protein (GFP) has been used as a vital marker in a variety of species. Here, we present the use of a GFP–promoter fusion to visualize carbon metabolism in a pathogenic fungus during growth on defined medium and during infection of plants. Isocitrate lyase (ICL), a key enzyme in carbon metabolism, is tightly regulated at the transcriptional level, with high levels of expression during 2-carbon growth and no expression during growth on glucose. A GFP–ICL promoter fusion was used to visualize carbon metabolism in the plant pathogenic fungus Tapesia yallundae during growth in vitro and in the host plant. The ICL promoter from Neurospora crassa retained its native induction and repression characteristics in T. yallundae. Loss of GFP fluorescence from hyphae after repression of the ICL promoter suggested a rapid turnover rate for GFP in T. yallundae. Regulation of this promoter was observed during infection, with expression occurring only on the plant surface, suggesting that 2-carbon metabolism occurs during this phase. These data suggest that GFP is a useful vital marker for the in planta imaging of fungal metabolism.