Molecular Biology of Host-Specific Toxin Biosynthesis

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A number of plant pathogens produce low molecular weight chemicals which are toxic to plants. Some toxins, called "host-specific", have the same host-range in vitro as the producing organism does in the field; that is, plants resistant to the pathogen are insensitive to that pathogen's toxin, and plant susceptible to the pathogen are sensitive to the toxin. Host-specific toxins produced by fungi in the genus Cochliobolus (also known as Helminthosporium or Bipolaris) have been implicated in two catastrophic epidemics of cereal crops in the U.S. in the last forty years. While in the last few years the chemical structures of most host- specific toxins have been determined, and their modes of action in some cases partly clarified, nothing is currently known about the biosynthesis of phytotoxins, nor about the molecular processes by which pathogens acquire the ability to make them. The objectives of this project are to identify and isolate enzymes involved in the biosynthesis of a host-specific toxin, to raise antibodies to these enzymes, and to isolate the genes which code for those enzymes. The antibodies and cloned genes will be tools to study the molecular basis of race specificity and toxin production in plant pathogenic fungi. Initial efforts will focus on the purification and characterization of two enzymes recently identified as being involved in the biosynthesis of a cyclic peptide toxin, HC- toxin, made by the fungal pathogen of maize, Cochliobolus carbonum. The purified enzymes will be used to raise antibodies, and, following amino acid sequencing, to make synthetic oligonucleotides. The antibodies and oligonucleotides will be used to clone the genes for these enzymes. The cloned genes will be used to compare isolates of the fungus which can an cannot make HC-toxin, and ultimately, to study at the molecular level the evolutionary processes by which pathogenic fungi acquire the ability to make toxins.***//