Location: Corn Insects and Crop Genetics Research2014 Annual Report
Objective 1: Discover and functionally characterize host genes and pathways that respond to pathogen infection in barley and maize. Sub-Objective 1A: Identification of barley Blufensin1 (Bln1)-mediated response pathways. Sub-Objective 1B: Functional confirmation via integrated reverse genetic analysis. Sub-Objective 1C: Identify and functionally characterize mechanisms of defense against leaf blights with hemi-biotrophic and necrotrophic lifestyles in maize. Objective 2: Identify and analyze the role of pathogen effectors that influence host response in barley and maize. Sub-Objective 2A: Isolation of Blumeria graminis f. sp. hordei (Bgh) effectors to identify plant targets that promote or suppress defense in host and nonhost interactions. Sub-Objective 2B: Identification of candidate effectors produced by S. turcica and C. heterostrophus during interactions with maize.
Large scale sequencing of pathogen genomes, as well as their plant hosts, has provided unprecedented access to the genes and gene networks that underlie host-pathogen interactions. Regulatory focal points critical to these interactions will be investigated. Determination of these focal points will enable the molecular dissection of important disease resistance pathways, as well as the creation of a molecular toolbox from which to apply modern plant breeding methodologies.
Plant diseases pose significant threats to crop production worldwide. To control diseases that can devastate our food, fiber, and fuel crops, it is necessary to know how pathogens cause diseases and how hosts defend against them. Using pathosystems of both barley and corn, this project aims to identify both host disease defense components and pathogen signaling molecules that suppress them. Cereal grains are one of the most important nutritional foundations of humans and domesticated animals world-wide. Hence, controlling existing and emerging cereal crop diseases is vital to food security. In addition to their economic importance, cereal hosts to fungal pathogens also present unique models for cellular biology. Effector proteins secreted by these pathogens take over host processes to enable their colonization. As such, effectors are optimal probes to help understand host pathways, particularly disease resistance. As part of Objective 1A and 1B, we have been investigating a barley pathogen susceptibility factor, designated Blufensin1. Blufensin1 was silenced and plants were subjected to genome-wide expression analysis, to identify potential interacting partners. The resulting gene set was highly enriched for proteins associated with nuclear import and the secretory pathway, two sub-cellular systems that are critical to plants being parasitized by pathogens. These results highlight the role of the Blufensin family in pathogen colonization on its barley host. As part of Objective 1C, year 1 of replicated, multi-location genetic mapping experiments was executed for both Southern corn leaf blight (Raleigh NC only) and Northern corn leaf blight (Ithaca, NY and Ames, IA) using a high resolution mapping resource curated by this team. Seeds and experimental designs were provided to collaborators at Cornell University and USDA-ARS in Raleigh, NC. Two replicates of 380 corn genotypes were inoculated with local pathogen isolates and phenotyped for disease symptoms in order to identify genetic factors involved in quantitative disease defense. It is expected that these experiments will provide lists of genes that can be tested for roles in defense. If the highest resolution we expect is achieved, the lists of gene candidates for each identified resistance factor will be smaller than 20 genes. Using the previous best resources, such lists ranged from 100 to 1000 genes, precluding advancement to testing. Histological analysis of the infection kinetics was also conducted in order to determine which time points following inoculation will be best for identifying host and pathogen genes involved in disease signaling. This was particularly important for Northern corn leaf blight because it has a variable latent period before causing strong and sudden disease symptoms. As part of Objective 2A, large-scale gene expression profiling experiments identified powdery mildew effector candidates that may function in suppressing host defense. Functional assays will be performed in subsequent years to establish their role in disease development and to identify host interacting proteins that can be exploited for disease defense. As part of Objective 2B, local isolates of Northern corn leaf blight were collected, purified, and raised for artificial inoculation of the 24,000 plants that are part of the genetic mapping experiment associated with Objective 1C. Since a local isolate was used in Ithaca, NY as well, failures to detect the same set of genetic components of defense at the two locations could either be due to environment or to differences in pathogen virulence. Particular attention will be paid to these cases, since they offer opportunities to dissect pathogen signaling effects on a specific host defense mechanism. The pathogen isolates collected are available for use in the gene expression experiments planned under this objective in the coming years. Active plant defense to microbial attack is highly dependent upon recognition events involving associated gene products in the host and pathogen. Host perception of pathogen-associated molecules result in signal transduction cascades ultimately leading to disease resistance. Knowledge gained from this project will promote broadly applicable disease control strategies.
1. Completed development of a new type of genetic analysis resource for corn that informs important breeding practices. Corn flowering is highly sensitive to day length, which does not vary dramatically across the seasons at low latitudes where corn was domesticated. During pre-breeding for line development in corn, sensitivity to day length is selected against so strongly that it prevents harnessing an unknown wealth of disease and pest resistance alleles harbored in tropical germplasm. Scientists at USDA-ARS in Ames, Iowa and at University of Delaware used DNA-marker-assisted breeding on nearly 150,000 plants to provide saturated coverage at each of the four most significant day length responsive genes in corn. The resource will be used by the project team and others to characterize how many and what kind of alleles are lost by present breeding practices, permitting several longstanding questions about artificial selection to be addressed for the first time. This research will enhance the availability of maize genetic diversity in the tropics for improvement of this crop.
Cernadas, R.A., Doyle, E.L., Nino-Liu, D., Wilkins, K.E., Bancroft, T., Wang, L., Schmidt, C., Caldo, R., Yang, B., White, F.F., Nettleton, D., Wise, R.P., Bogdanove, A. 2014. Code-assisted discovery of TAL effector targets in bacterial leaf streak of rice reveals contrast with bacterial blight and a novel susceptibility gene. PLoS Pathogens. DOI: 10.1371/journal.ppat.1003972.
Wise, R.P., Surana, P., Fuerst, G.S., Xu, R., Mistry, D., Dickerson, J., Nettleton, D. 2014. Flor revisited (again): eQTL and mutational analysis of NB-LRR mediated immunity to powdery mildew in barley. Journal of Integrative Agriculture. 13(2):237-243.
Xu, W., Meng, Y., Wise, R.P. 2014. Mla- and Rom1-mediated control of microRNA398 and chloroplast copper/zinc superoxide dismutase regulates cell death in response to the barley powdery mildew fungus. New Phytologist. 201(4):1396-1412.
Hayes, N., Maffin, L., McGhee, L., Hall, G., Hubbard, T., Whigham, E., Wise, R.P. 2013. iTAG Barley: A 9-12 curriculum to explore inheritance of traits and genes using Oregon Wolfe barley. iTunes. Available: https://itunes.apple.com/us/book/itag-barley/id715260619?mt=11.
Mascher, M., Muehlbauer, G.J., Rokhsar, D.S., Chapman, J., Schmutz, J., Barry, K., Munoz-Amatriain, M., Close, T.J., Wise, R.P., Schulman, A.H., Himmelbach, A., Mayer, K.F., Scholz, U., Poland, J.A., Stein, N., Waugh, R. 2013. Anchoring and ordering NGS contig assemblies by population sequencing (POPSEQ). Plant Journal. DOI: 10.1111/tpj.12319.
Kronmiller, B.A., Wise, R.P. 2013. TEnest 2.0: Computational annotation and visualization of nested transposable elements. In: Peterson, T., editor. Methods in Molecular Biology. New York, NY: Springer. p. 305-320.