2013 Annual Report
1a.Objectives (from AD-416):
Objective 1: Identify changes in host gene expression and small RNA-mediated regulation associated with viroid and bacterial infection and disease development as potential targets for disease management. Objective 2: Identify key metabolites that are involved in the early stages of pathogenesis and may have global effects on disease resistance through either their bioactive nature or effect on redox-status. Sub-objective 2.A. Identify and quantify secondary metabolites induced upon infection of tomato with either P. syringae or potato spindle tuber viroid (PSTVd). Sub-objective 2.B. Determine bioactivity of secondary metabolites induced upon infection of tomato with either P. syringae or PSTVd. Sub-objective 2.C. Determine the effect on redox status of secondary metabolites induced upon infection of tomato with either P. syringae or PSTVd. Objective 3: Identify the molecular signals and pathways used by viroids to move through the cytoplasm, enter the nucleus or chloroplast of the host cell, and begin replication. Sub-objective 3.A. Determine the role of host protein 4/1 (and other proteins interacting with 4/1) in the intra- and intercellular movement of PSTVd. Sub-objective 3.B. Use sequence motifs derived from Eggplant latent viroid (ELVd) to redirect mRNAs encoding enzymes involved in terpenoid biosynthesis into the chloroplast.
1b.Approach (from AD-416):
This project seeks to elucidate specific signaling mechanisms that are involved in plant disease resistance. Plants respond to biotic stress by the integration of responses located in two different cellular compartments, the symplast and the apoplast. Improvement of existing resistance to plant disease requires a more comprehensive understanding of key apoplastic and symplastic responses to pathogen invasion and how they are integrated. In the apoplast, we will examine the complex interplay between secondary metabolites and redox signaling that controls early events in bacterial and viroid pathogenesis as well as other responses triggered by long distance signaling. In the symplast, we will examine the molecular interactions between viroids and their hosts to determine how these small RNA molecules replicate and move between organelles within a single cell or over long distances between cells. The long-term objective of this multidisciplinary project is to understand the role of plant-pathogen signaling in disease resistance in sufficient detail that novel strategies can be developed to render plants resistant/immune to pathogen infection. In the intermediate term, we will test the ability of certain viroid-derived targeting signals to redirect mRNAs to the chloroplast, thereby adding novel biosynthetic capabilities to chloroplast metabolism with the need for chloroplast genome transformation.
Determination of the role played by redox (oxidants/antioxidants) metabolites produced by the plant upon detection of bacterial pathogens was advanced by the development of a new technique to monitor redox potential during these interactions. We now know the precise timing (within minutes) of the interactions which allows precision sampling to examine metabolic changes. We have been able to identify several different outcomes depending on whether the plant/bacterial combination results in a disease, resistance, or saprophytic interaction. In addition, we have identified specific metabolites that are oxidized during the plant/bacterial response producing highly lethal radicals that kill the pathogen and possibly the plant cell. We also confirmed that Potato spindle tuber viroid is present in phloem sap in tomato, and is not present in xylem sap, suggesting that long-distance viroid RNA transport is exclusively through the phloem.
Few options are available for management of bacterial diseases in plants. Antimicrobial resistance continues to be a high priority issue in agriculture and the routine use of antibiotics against bacterial plant pathogens is likely to be eliminated as it has been with animals. Here we investigated an alternative approach, using a technique, which we developed this year, to monitor redox (oxidant/antioxidant) mechanisms. The improved precision of the technique revealed that certain metabolites that plants excrete into the apoplast, can be converted into highly effective “natural” antimicrobial products. These data and findings will be used by researchers in developing “antibiotic-free” bacterial-resistant plants in the future.
Averyanov, A.A., Lapikova, V.P., Pasechnik, T.D., Zakharenkova, T.S., Baker, C.J. 2012. Self-inhibition of spore germination via reactive oxygen in the fungus Cladosporium cucumerinum, causal agent of cucurbit scab. European Journal of Plant Pathology. 131:541-550.
Baker, C.J., Kovalskaya, N.Y., Mock, N.M., Deahl, K.L., Owens, R.A., Whitaker, B.D., Roberts, D.P., Hammond, R., Averyanov, A.A. 2013. Real-time monitoring of the extracellular redox potential of cell suspensions during plant/bacterial interactions. Physiological and Molecular Plant Pathology. 82:20-27.