Project Number: 8042-21220-256-00-D
Project Type: In-House Appropriated
Start Date: Apr 2, 2017
End Date: Aug 12, 2018
Objective 1: Discover pathogen gene function through use of functional genomics [NP303; C2, PS2A] Objective 2: Characterize underlying mechanisms of resistance in Solanaceous hosts in response to pathogen infection [NP303; C2, PS2B] Objective 3: Develop novel gene strategies for genetic improvement to manage disease in solanaceous crops. [NP303; C3, PS3A]
Three major diseases of potato will be investigated, late blight (caused by Phytophthora infestans), early blight (caused by Alternaria solani) and common scab (caused by Streptomyces scabies and related species). Two of these diseases, early blight and late blight, are also major disease problems on the related Solanaceous crop tomato. The first objective focuses on pathogen glycosyl hydrolase enzymes which have multiple roles in the initiation of plant disease including thefacilitation of pathogen ingress, generation of carbon substrates, and for direct elicitation of necrosis. Individual genes encoding glycosyl hydrolases from Alternaria and Streptomyces will be identified through analysis of gene expression during stages of host infection using literature analysis and RNA-seq based gene expression profiling. Genes identified as potential candidates will be cloned and introduced into plants individually through Agrobacterium-mediated infiltration and transient expression. Screening will include microscopic examination of plant tissue and screening of upregulated host response genes. In a parallel study, glycosyl hydrolases will be disrupted and/or silenced followed by assessment of changes in pathogenicity or virulence on inoculated hosts. This information will provide targets for suppression using RNAi, leading to reduced disease. The second objective will study the effects of host manipulation on expression of genes within the host and within the pathogen. Plants treated or untreated with supplemental nitrogen will be tested to generate a RNA-seq based gene expression profile to evaluate host and pathogen responses that could reduce infection by Alternaria. Auxin analogues will be applied to potato foliage, followed by assessement of gene expression in both the host and Streptomyces, following the same gene expression profiling. While both the nitrogen treatment and auxin analogue treatments are not directly suitable for disease reduction in cropping systems, the understanding of their effect on genes from the pathogen and the host can provide a basis for altering gene expression in the absence of these host modifying compounds through genetic manipulation. The third objective investigates methods for weakening the cell walls of Phytophthora and Alternaria which can reduce the ability of the pathogens to penetrate and grow through the plant tissue. The weakening may also allow other host defenses to be more effective in limiting the pathogen. Specific enzymes will be derived from numerous sources including enzyme mixes used for protoplast generation, plant defensive enzymes, and enzymes the pathogen may use itself to construct and deconstruct its own cell wall. Enzymes will be tested using Agrobacterium-mediated expression, followed by inoculation of the pathogens to infiltrated tissues. Enzymes will be enhanced through addition of specific cell wall binding motifs such as cellulose binding domains and chitin binding domains, and antimicrobial peptide sequences. Successful development of enzyme mediated pathogen control will add to the methods of disease control and may provide a basis for control of a broader range of pathogens.