Date of Award
Master of Science
Molecular Biology Microbiology and Biochemistry
Aspergillus flavusis a filamentous fungusthat causes an ear and kernel rot in maize (Zea mays L.). It produces a toxic secondary metabolite, aflatoxin, on the colonized maize kernels. Aflatoxin is a carcinogen to humans and animals. The toxin is also an immunosuppressant and causes aspergillosis in immune compromised individuals. Therefore, the presence of aflatoxin in food is strictly regulated by governmental agencies. Contaminated food leads to severe loss in profit and in marketable yield. There has been extensive research to investigate resistance of maize toA. flavus. Certain lines of maize exhibit increased resistance to A. flavus and aflatoxin accumulation compared to others and correlated with that are proteins and metabolites that differ in abundance in those lines. Among them are members of the cupin superfamily of proteins and products of special nitrogen metabolism (derived from glutamate). The goal here was to identify networks underlying disease resistance indifferent maize genotypes through the identification of protein-protein interactions and the analysis of transcript abundance profiles realting to cupins and glutamate. The outcome will be an understanding of host resistance to A. flavussufficient to develop methods to prevent pre-harvest contamination by aflatoxin. A protein abundant in resistant maize was identified as a cupin and named ZmCUP1. The cDNA isolation, expression in E. coliand characterization of the protein encoded by the mRNA, Zmcup1, lead to the discovery that the ZmCUP1 protein had anti fungal properties and oxalate decarboxylase activity (EC 18.104.22.168). Another part of the project aimed at understanding the involvement of a transgene that encoded bacterial NADPH-glutamate dehydrogenase (GDHA; EC 22.214.171.124) that reduced aflatoxin accumulation by half. A maize partial predicted protein to protein interactome was built and used to identify potential interactions between proteins expressed differentially in lines of maize resistant to A. flavus. These interactions were characterized in-silico and one specific interaction, between Zmcup1 and a maize zinc finger protein was characterized in vitro.
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