Abstract
BackgroundPostharvest losses of citrus fruit due to green mold decay, caused by the fungus Penicillium digitaum, have a considerable economic impact. However, little is known about the molecular processes underlying the response of citrus fruit to P. digitatum.ResultsHere we describe the construction of a subtracted cDNA library enriched in citrus genes preferentially expressed in response to pathogen infection followed by cDNA macroarray hybridization to investigate gene expression during the early stages of colonization of the fruit's peel by P. digitatum. Sequence annotation of clones from the subtracted cDNA library revealed that induction of secondary and amino acid metabolisms constitutes the major response of citrus fruits to P. digitatum infection. Macroarray hybridization analysis was conducted with RNA from either control, wounded, ethylene treated or P. digitatum infected fruit. Results indicate an extensive overlap in the response triggered by the three treatments, but also demonstrated specific patterns of gene expression in response to each stimulus. Collectively our data indicate a significant presence of isoprenoid, alkaloid and phenylpropanoid biosynthetic genes in the transcriptomic response of citrus fruits to P. digitatum infection. About half of the genes that are up-regulated in response to pathogen infection are also induced by ethylene, but many examples of ethylene-independent gene regulation were also found. Two notable examples of this regulation pattern are the genes showing homology to a caffeine synthase and a berberine bridge enzyme, two proteins involved in alkaloid biosynthesis, which are among the most induced genes upon P. digitatum infection but are not responsive to ethylene.ConclusionsThis study provided the first global picture of the gene expression changes in citrus fruit in response to P. digitatum infection, emphasizing differences and commonalities with those triggered by wounding or exogenous ethylene treatment. Interpretation of the differentially expressed genes revealed that metabolism is redirected to the synthesis of isoprenes, alkaloids and phenylpropanoids.
Highlights
Postharvest losses of citrus fruit due to green mold decay, caused by the fungus Penicillium digitaum, have a considerable economic impact
As a result of the analysis, 352 clones (25% of the total) with differential expression were identified, from which 297 were up-regulated in I over W, whereas 55 were downregulated. These preliminary results demonstrated that RindPdigS is enriched in genes with higher expression in infected than in wounded tissue, and confirmed that the generated cDNA macroarray is an effective efficient tool for analyzing the transcriptional responses of citrus fruit to P. digitatum infection
These two cDNA libraries were obtained from two different Citrus species, C. sinensis and C. clementina for RindPdigS and RindPdig24, respectively, previous work has shown a high level of identity between homologous genes from these closely related Citrus species [20], as we have found by comparing common unigenes between both libraries, whose identity was usually higher than 99%
Summary
Postharvest losses of citrus fruit due to green mold decay, caused by the fungus Penicillium digitaum, have a considerable economic impact. Harvested fruits are usually stored before they reach the market for fresh consumption. During this postharvest period fruits are subjected to both biotic and abiotic stress conditions. It is well known that the flavedo (outer colored part of the rind) is more resistant to P. digitatum than the albedo (inner white part) [1,2]. This fact has been classically associated with the presence of both preformed and induced antifungal compounds in the flavedo [3]. Other responses triggered by this fungus include the induction of PR proteins, such as b-1,3-glucanase and chitinase, and phenylalanine ammonia lyase (PAL), which catalyzes the first step in the phenylpropanoid pathway [1,6,7,8]
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