Evolution and Diversity of Fungal Pathogens and Molecular Mechanisms of Host Defense

NON-TECHNICAL SUMMARY: Plant pathogens such as Fusarium, Phytophthora and Magnaporthe cause extensive damage to a wide range of agriculturally important plants every year. Certain species of Fusarium also infect skins of healthy humans and produce toxins that affect food safety and animal/human health. This project will focus on the evolution, phylogeny and identification of plant pathogens and the mechanism of host defense response. With the aid of the Fusarium and Phytophthora databases, accurate and rapid identification of causal agents will help guide disease management strategies. On the other hand, understanding the mechanism of rice defense response will help improve breeding strategies and cultural practices and consequently benefits rice growers in the U.S. and abroad. Since rice is not only an important food crop, but also an excellent model for monocots, the discovery from this project will have broader impacts on breeding and disease control in other cereals, turf grasses and biofuel crops such as switchgrass.<P>

APPROACH: To study the evolution and phylogeny of fungi and oomycetes, DNA primers will be designed corresponding to sites within genes that are conserved across the taxon of interest. Phylogenetic relationships are inferred using a consensus approach, utilizing distance, maximum parsimony, Bayesian and Maximum Likelihood methods. Bootstrap as well as Bayesian posterior probabilities are used as indicators of node support. Intraspecific polymorphism identified in these markers can be used for population analyses, as well as nucleotide polymorphism identified in other loci and tandem repeats. Species boundaries are inferred using principles of genealogical concordance. Resulting data, along with any available phenotypic traits of each species and individual isolates within species will be archived in appropriate databases in a format that can be easily searched and updated through the internet. While multiple loci are generally required to infer robust phylogenies and to recognize species boundaries, a single locus can be implemented as a useful identification tool. A ~700 bp portion of the translation elongation factor 1-alpha gene has proven to be a very powerful tool for identifying Fusarium isolates to phylogenetically defined species. In the genus Phytophthora, the internal transcribed spacer (ITS) regions of the nuclear ribosomal RNA gene repeat have proven to be the best frontline tool for identification, but protein-coding genes such as HSP90 have potential for greater resolution with no loss of breadth of utility. Sequence based identification of unknown pathogens using these loci involves DNA sequencing of the target locus, followed by BLAST searches to identify best hits against a database of sequences corresponding to vouchered, publicly available specimens. A combination of molecular,genetic, genomic and proteomic approaches will be used to study the signal perception and transduction of rice defense response. Putative effectors from the rice blast fungus will be tested for their role in elicitation or suppression of plant defense response using particle bombardment and bacteria-based delivery system. In addition, attempts will be made to identify their interacting proteins and potential targets in rice cells using various approaches such as yeast two-hybrid, tandem affinity purification tag and bimolecular fluorescence complementation. Recently, transgenic analysis reveals that stress-responsive rice MAP kinases may modulate hormone signaling and mediate cross-talks between biotic and abiotic stress responses. To gain insights into the rice MAP kinase signaling, putative MAP kinase substrates will be identified by phosphorylation screening and protein-protein interaction. In vivo phosphorylation and functional analysis will be conducted to further elucidate the role of these MAP substrates in rice stress signaling. In addition, molecular, biochemical, physiological and pathological analyses will be conducted to determine the role of ethylene, jasmonic acid and abscisic acid signal pathways in rice biotic and abiotic cross-talks.