Progress 08/15/06 to 08/14/08
Outputs
OUTPUTS: In order to accomplish the research goals of this project, several outputs were necessary. All were accomplished in the appropriate time frame. We generated a fine-scale genetic map of sorghum in the region of the Pc locus. We constructed a fosmid library from sorghum line Pc/Pc Colby. We cloned the Pc region, and also sequenced it, from inbreds BTx623 (on a BAC clone) and a Pc/Pc Colby fosmid. We also cloned and sequenced PCR fragments from several pc derivatives of Pc/Pc Colby, and demonstrated that they were primarily deletion products of unequal homologous recombination. Finally, we also generated expression data by RT-PCR from all of the Pc and pc lines investigated. This information was disseminated in numerous seminars given by the PI and the lead scientist on the project, Dr. Ervin Nagy, a postdoctoral fellow in the PI's lab. In addition, the sequence data of all BACs, fosmids, and PCR products generated in the project were downloaded to GenBank with unrestricted access by all interested parties. Finally, as indicated below, there were two refereed publications that described all of the research that was undertaken. PARTICIPANTS: The primary researcher on this project was postdoctoral fellow Ervin Nagy. He has completed his term in my lab and is currently pursuing a second postdoctoral experience. This project involved a collaboration between Dr. Larry Dunkle and his lab at Purdue University, with significant involvement by Dr. Patricia Klein and the late Dr. Keith Schertz, both at Texas A&M University. The training from this funded project was solely for Dr. Nagy, who learned several molecular biological technologies (e.g., fosmid library construction, RT-PCR, DNA sequencing) and some bioinformatics (e.g., genome annotation) while in my lab. TARGET AUDIENCES: The target audience for this research is other plant pathologists, in both academia and industry. Our dissemination of these research results has significantly altered our understanding of the interaction between different kinds of disease resistance and is leading to new research programs to screen for and characterize the breadth of the phenomena we have discovered. PROJECT MODIFICATIONS: No major modifications were required or undertaken in this project from the research proposed.
Impacts
The cloning of the Pc region indicated that it was comprised of three CC-NBS-LRR genes like those usually associated with gene-for-gene disease resistance. The analysis of many independent pc derivatives of this specific Pc gene indicated that loss of the central gene, either by internal deletion or by unequal recombination, was responsible for the change from susceptibility (Pc) to resistance (pc) to the saprophytic fungus Periconia circinata. Hence, the loss of a disease resistance gene created resistance, the opposite of what is expected for the usual biology of CC-NBS-LRR disease resistance genes. We proposed that the P. circinata toxin mimics an avirulence gene product, thus causing the CC-NBS-LRR to initiate hypersensitive necrosis (HR). However, instead of providing resistance, this HR creates the necrotic tissue that is needed for infection by P. circinata. This is the first case of a known disease susceptibility gene being associated with corruption of a disease resistance pathway, although a similar system has been indicated in the model system Arabidopsis by the Wolpert lab. Most interesting, though, was the observation that the recombination events at the Pc locus that yielded pc alleles were not random. Instead, unequal crossovers were resolved in one of the most variable portions of these genes, within the LRR regions that carry the primary responsibility for differential recognition of different avirulence gene products. The simplest explanation of this result is that it is caused by a site-directed recombination phenomenon, the first discovery of site-directed recombination in any plant. A patent application has been made on the use of the process we discovered, to make plants with specific useful traits This patent application is entitled "Compositions and Methods Useful for Site-Directed Recombination in Plants."
Publications
- Nagy, E. D., T.-S. Lee, W. Ramakrishna, Z. Xu, P. E. Klein, P. SanMiguel, C.-P. Cheng, J. Li, K. M. Devos, K. Schertz, L. Dunkle and J. L. Bennetzen (2007) Fine mapping of the Pc locus of Sorghum bicolor, a gene controlling the reaction of a fungal pathogen and its host-selective toxin. Theor. Appl. Genet. 114:961-970.
- Nagy, E. D. and J. L. Bennetzen (2008) Pathogen corruption and site-directed recombination at a plant disease resistance gene cluster. Genome Res., published online in August, hard copy in press.
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