Proteomic and Biochemical Evidence Support a Role for Transport Vesicles and Endosomes in Stress Response and Secondary Metabolism in <i>Aspergillus parasiticus</i>

Aflatoxin is among the most potent naturally occurring carcinogens known. Previous studies demonstrated that endosomes in the filamentous fungus <i>Aspergillus parasiticus</i> carry enzymes that catalyze the final two steps in aflatoxin synthesis, and these structures also play a role in aflatoxin storage and export. We hypothesized that endosomes house a complete and functional aflatoxin biosynthetic pathway. To address this hypothesis, we purified a cellular fraction containing endosomes, transport vesicles, and vacuoles (V fraction) from <i>A. parasiticus</i> grown under aflatoxin inducing and noninducing conditions. We also added (fed) aflatoxin pathway intermediates to V fraction to test the functional status of aflatoxin pathway enzymes. High throughput LC–MS/MS analysis of proteins in V fraction detected 8 aflatoxin enzymes with high reliability and 8 additional enzymes at lower reliability, suggesting that most aflatoxin pathway enzymes are present. Purified V fraction synthesized aflatoxin and addition of the pathway intermediate versicolorin A increased aflatoxin synthesis, confirming that middle and late aflatoxin enzymes in V fraction are functional. Of particular significance, proteomic and biochemical analysis strongly suggested that additional secondary metabolic pathways as well as proteins involved in response to heat, osmotic, and oxidative stress are housed in V fraction

Proteomic and Biochemical Evidence Support a Role for Transport Vesicles and Endosomes in Stress Response and Secondary Metabolism in <i>Aspergillus parasiticus</i>

Aflatoxin is among the most potent naturally occurring carcinogens known. Previous studies demonstrated that endosomes in the filamentous fungus <i>Aspergillus parasiticus</i> carry enzymes that catalyze the final two steps in aflatoxin synthesis, and these structures also play a role in aflatoxin storage and export. We hypothesized that endosomes house a complete and functional aflatoxin biosynthetic pathway. To address this hypothesis, we purified a cellular fraction containing endosomes, transport vesicles, and vacuoles (V fraction) from <i>A. parasiticus</i> grown under aflatoxin inducing and noninducing conditions. We also added (fed) aflatoxin pathway intermediates to V fraction to test the functional status of aflatoxin pathway enzymes. High throughput LC–MS/MS analysis of proteins in V fraction detected 8 aflatoxin enzymes with high reliability and 8 additional enzymes at lower reliability, suggesting that most aflatoxin pathway enzymes are present. Purified V fraction synthesized aflatoxin and addition of the pathway intermediate versicolorin A increased aflatoxin synthesis, confirming that middle and late aflatoxin enzymes in V fraction are functional. Of particular significance, proteomic and biochemical analysis strongly suggested that additional secondary metabolic pathways as well as proteins involved in response to heat, osmotic, and oxidative stress are housed in V fraction

Galdieria_in_Richmond_Mine_FISH

Micrographs of two G. sulphuraria cells detected using Cya1208 rRNA probe in an A-drift biofilm from the Richmond Min

208_Genomes

List of 208 sequenced genomes used to generate protein families and phylogenetic tree