Location: Corn Insects and Crop Genetics Research
2023 Annual Report
Objectives
Objective 1. Develop an integrated network of barley-powdery mildew interactions as a model to dissect immune signaling in cereals.
Sub-Objective 1A. Identify protein-protein interactions (PPI) among host and pathogen via yeast two-hybrid (Y2H) next-generation-interaction screens (NGIS).
Sub-Objective 1B: Authenticate immune-active genes via phenotyping and functional assays.
Objective 2: Identify regulatory elements associated with immunity to exploit in modern breeding programs.
Sub-Objective 2A: Genome-wide prediction of transcription factor (TF) activation motifs.
Sub-Objective 2B: Validation of immune-active, cis-regulatory elements (CRE)-associated transcription factors (TFs) via highthroughput phenotyping and in-depth molecular characterization.
Approach
Large-scale sequencing of plant and pathogen genomes has provided unprecedented access to the genes and gene networks that underlie diverse outcomes in host-pathogen interactions. Determination of regulatory focal points critical to these interactions will provide the molecular foundation necessary to dissect important disease resistance pathways. This knowledge can be used to guide modern plant breeding efforts in response to pathogens that present diverse challenges to the host.
Progress Report
Plant pathogens are among the greatest threats to crop production worldwide, resulting in yield losses of 10 to 20% (=$100 to 200 billion) each year. Obligate biotrophic fungi, for example, mildews and rusts, require a living host to survive and cause some of the most destructive epidemics. A general view of the regulatory programs that render a plant resistant to pathogens is beginning to emerge in model organisms, but is still in its infancy in large-genome temperate cereals, vital to feeding the world's growing population. The interaction between barley, and the powdery mildew fungus, is central to address this challenge. In this system, the outcome is determined largely by the plant’s response to secreted pathogen effectors. Disease is blocked by host immune receptors encoded by resistance (R) genes, designated by the prefix, Ml (for mildew resistance). As a model for large-genome Triticeae grain crops, for example, barley, wheat, and rye, this research generates the knowledge base to drive scientific advances that will ultimately improve crop productivity, pest and disease resistance, and tolerance to climate change.
In support of Objective 1, Sub-Objective 1A, ARS researchers, in collaboration with Iowa State University and University of Copenhagen in Denmark, employed a technique termed yeast two-hybrid next-generation interaction screening (Y2H-NGIS) to decipher the interactions among pathogen effectors and hosts infected by them. The barley MLA receptor was used as a model regulator to interrogate cereal immune response, as it’s alleles and orthologs confer resistance to several diseases, including powdery mildew, stem rust, stripe rust and rice blast. Several representative powdery mildew effector proteins, including AVRA1, AVRA7, AVRA9, and AVRA13, were used as baits for Y2H-NGIS, followed by next-generation sequencing, quantitation and ranking with custom software to mine for novel protein-protein interactions. These results were integrated with a previously generated, genome-wide barley interactome, comprising 66133 edges and 7181 nodes, to predict MLA localization, signaling response, and effector mechanisms.
Cellular dynamics are regulated by coordinated biological networks that interact in multiple compartments and organelles. Disease phenotypes are the product of dynamic changes in gene and protein interactions. Thus, these results to identify new components of host disease defense and the pathogen effectors that suppress them will enable breeders and growers to produce better crops.
Accomplishments
