Aspects of nitrogen and carbon metabolism that regulate fumonisin biosynthesis in Fusarium verticillioides
Fusarium verticillioides is a ubiquitous fungal pathogen that can cause ear, kernel, seedling, and stalk diseases in maize. During colonization of maize kernels, this fungus produces fumonisins, which are mycotoxins that adversely affect human and animal health. My research focused on aspects of fumonisin B1 (FB1) regulation related to the colonization of blister-stage kernels and of the endosperm of mature kernels. First, I discovered that, during growth on blister-stage kernels, expression of AREA (nitrogen metabolite regulator) and production of FB 1 are repressed. In addition, the failure of the fungus to produce FB 1 during growth on the blister kernels is likely due to alkaline pH, which results from fungal metabolism caused by a low carbon:nitrogen ratio in these kernels. I tested the hypothesis that AREA is required for FB1 production during growth on mature kernels with a F. verticillioides strain that constitutively expressed AREA. When grown on mature maize kernels, this strain produced FB 1 even with the addition of concentrations of ammonium phosphate that were repressive in the wild type. These results indicate that AREA is required for FB1 production. With respect to the endosperm of mature kernels, expression of three putative sugar transporter genes ( FST1, FST2, and FST4) in F. verticillioides was higher in colonized endosperm than in germ tissue. I created mutants by disrupting each gene and found that only strain Δ fst1, a disruption of FST1, did not produce FB 1. In culture, the Δfst1 grew similar to the wild type; however, when inoculated onto maize ears, disease symptom expression and fungal growth were delayed. Characterization of the FST1 promoter revealed that regulation of FST1 expression was similar to that of fumonisin biosynthetic (FUM) genes, in that expression was highest during growth on endosperm tissue and repressed by elevated concentrations of ammonium in the growth medium. With a fluorescent tag attached to FST1, the protein transiently localized to the periphery of the cells near the plasma membrane and in vacuoles. Expression of FST1 in a yeast strain lacking hexose transporter genes did not complement the yeast mutation, suggesting that FST1 does not transport glucose, fructose or mannose. Thus, the results of this study indicate that FST1 impacts the colonization of living maize kernels, affecting the development of disease symptoms and mycotoxin production during colonization of maize kernels.^
Charles P. Woloshuk, Purdue University.
Agriculture, Plant Pathology