Transcriptome analysis of an incompatible Persea americana-Phytophthora cinnamomi interaction reveals the involvement of SA- and JA-pathways in a successful defense response

Abstract

Phytophthora cinnamomi Rands (Pc) is a hemibiotrophic oomycete and the causal agent of Phytophthora root rot (PRR) of the commercially important fruit crop avocado (Persea americana Mill.). Plant defense against pathogens is modulated by phytohormone signaling pathways such as salicylic acid (SA), jasmonic acid (JA), ethylene (ET), auxin and abscisic acid. The role of specific signaling pathways induced and regulated during hemibiotroph-plant interactions has been widely debated. Some studies report SA mediated defense while others hypothesize that JA responses restrict the spread of pathogens. This study aimed to identify the role of SA- and JA- associated genes in the defense strategy of a resistant avocado rootstock, Dusa in response to Pc infection. Transcripts associated with SA-mediated defense pathways and lignin biosynthesis were upregulated at 6 hours post-inoculation (hpi). Results suggest that auxin, reactive oxygen species (ROS) and Ca2+ signaling was also important during this early time point, while JA signaling was absent. Both SA and JA defense responses were shown to play a role during defense at 18 hpi. Induction of genes associated with ROS detoxification and cell wall digestion (β-1-3-glucanase) was also observed. Most genes induced at 24 hpi were linked to JA responses. Other processes at play in avocado at 24 hpi include cell wall strengthening, the formation of phenolics and induction of arabinogalactan, a gene linked to Pc zoospore immobility. This study represents the first transcriptome wide analysis of a resistant avocado rootstock treated with SA and JA compared to Pc infection. The results provide evidence of a biphasic defense response against the hemibiotroph, which initially involves SA-mediated gene expression followed by the enrichment of JA-mediated defense from 18 to 24 hpi. Genes and molecular pathways linked to Pc resistance are highlighted and may serve as future targets for manipulation in the development of PRR resistant avocado rootstocks.

Fig 1. Phytophthora cinnamomi (Pc) infection of resistant avocado roots.

A. Scanning electron micrograph of two Pc cysts germinating, with hyphae entering the root at the same site via direct penetration 1 hpi. B. Confocal image of Pc hyphae (white arrow) penetrating through the root epidermis at 3 hpi. C. Confocal image of lignified root cortex cells with hyphae (white arrow) at 12 hpi. D. Confocal image of root cortex cells demonstrating extensive lignification at 24 hpi. E. Brown necrotic lesions are visible on avocado roots six weeks after Pc inoculation.

Fig 2. SA and JA-linked transcripts induced by P. cinnamomi (Pc) infection of avocado.

Transcripts differentially expressed by either SA of MeJA treatments were identified in P. cinnamomi infected datasets. The values for the transcripts with multiple probes on the microarray was first averaged, and transcripts with poor Pearson correlation was removed. A. The total number of differentially expressed transcripts at 6 hours post treatment/infection. B. The total number of differentially expressed transcripts at 18 hours post treatment/infection. C. The total number of differentially expressed transcripts at 24 hours post treatment/infection. Note that no 24 hrs time point post SA treatment was analyzed.

Fig 3. HCL performed on fold-change filtered avocado transcripts in the SA (salicylic acid), PC (Phytophthora cinnamomi) and JA (MeJA) datasets.

The colour-scale indicates Log₂ fold-change and the branches of the trees are ordered according to the Pearson correlation coefficient (r), with nodes closer to 1 indicative of a positive correlation.

Fig 4. GO classification differentially expressed avocado transcripts in response to Phytophthora cinnamomi infection.

A) Induced molecular function; B) repressed molecular function; C) induced biological process and D) repressed biological process. Second and third level GO terms are indicated with the corresponding percentage of transcripts for molecular function and biological processes respectively.

Fig 5. Model of SA and JA pathway temporal regulation induced by Phytophthora cinnamomi in tolerant avocado roots.

At 6 hrs an induction of signaling pathways and transcription factors occurred, which was linked to SA signaling. Subsequently at 18 hrs, both SA and JA-linked induction of ROS, cell wall strengthening and PR transcripts were observed. At 24 hrs transcripts related to lignin formation, flavonoid and volatile compound generation, as well as JA-linked ROS were up regulated. Text shaded in pink are SA-linked, text shaded in green are JA-linked. The pathogen is indicated in grey, nucleus in blue and lignin deposition in maroon.