Title: Using an apple (Malus) microarray for expression analysis of responses to compatible and incompatible pathogens Authors
|Schuyler, Korban - U OF IL - URBANA-CHAMPAIG|
|Norelli, John (jay)|
Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: August 13, 2008
Publication Date: August 21, 2008
Citation: Phillips, J.G., Dardick, C.D., Schuyler, K.S., Bassett, C.L., Wisniewski, M.E., Norelli, J.L., Bicsanczy, A.M. 2008. Using an apple (Malus) microarray for expression analysis of responses to compatible and incompatible pathogens. Meeting Abstract. MAPMBX Program Book. p. 31. Technical Abstract: Fire blight is a devastating disease of apple (Malus x domestica) caused by the bacterial pathogen Erwinia amylovora (Ea). Ea enters the plant through blossom nectaries or wounds, multiplies in the apoplast and spreads through the plant via vascular tissues. When infiltrated into host leaves, Ea induces lipid peroxidation, electrolyte leakage, and antioxidant enzyme changes, similar in intensity and kinetics to the oxidative burst observed during the development of a hypersensitive response (HR) following inoculation with an incompatible rather than a compatible pathogen (Venisse et al. 2001. Pl. Physiol. 125:2164-72). Pseudomonas syringae pv. syringae strain B86-6 (Pss) is a broad host range bacterial pathogen that is incapable of producing disease in apple but produces an HR response upon infiltration into apple leaves (incompatible pathogen). The objective of this study was to compare the defense response of Malus to challenges by Ea and Pss using microarray analysis to gain insight into how Ea overcomes defense reactions associated with HR and establishes systemic infection. To this end, we used a 40,000 feature, two-channel, printed Malus microarray that contains 548 control probes and 39,412 long-oligonucleotide (70-mer) probes designed to non-redundant Malus EST contigs (unigenes) obtained from different tissues, genotypes, developmental stages and stress conditions. Leaf tissues were harvested from apple shoots of fire blight susceptible ‘Malling 26’ rootstock inoculated with either phosphate buffer (Mock), virulent Ea strain Ea273, or Pss strain B86-6 at 6 h post-inoculation. RNA was isolated from tissues, and labeled with Alexa Dye 555 and 647. Several protocols were tested to optimize transcript labeling and microarray hybridization. A loop design was used to compare expression profiles of Mock vs. Ea, Mock vs. Pss, and Ea vs. Pss challenged tissues. Differentially expressed genes were identified using two interconnected mixed linear models for normalization of array data and determination of gene effects (Wolfinger et al. 2001 J. Comput. Biol. 8:625-37). A total of 1541 genes were differentially expressed in Mock vs. Ea, 995 in Mock vs. Pss, and 726 in Ea vs. Pss comparisons. Interestingly, a high number of differentially expressed genes (450) were common in both Mock vs. Ea and Mock vs. Pss comparisons, suggesting commonalities between both interactions, while 726 differentially expressed genes in the Ea vs. Pss comparison suggest there are also specific responses to both compatible and incompatible interactions. Bioinformatic analysis of differentially regulated genes is currently in progress to identify gene families and/or enzymatic pathways that are either common or specific to the compatible and incompatible host-pathogen responses in Malus.