Comparative analysis of ESTs involved in grape responses to

Department of Viticulture and Enology, University of California, Davis, Davis, California, United States
BMC Plant Biology (Impact Factor: 3.81). 02/2007; 7(1):8. DOI: 10.1186/1471-2229-7-8
Source: PubMed


The gram-negative bacterium Xylella fastidiosa (Xf) is the causal agent of Pierce's disease (PD) in grape as well as diseases of many fruit and ornamental plants. The current molecular breeding efforts have identified genetic basis of PD resistance in grapes. However, the transcriptome level characterization of the host response to this pathogen is lacking.
Twelve tissue specific subtractive suppression hybridization (SSH) cDNA libraries derived from a time course sampling scheme were constructed from stems, leaves and shoots of PD resistant and susceptible sibling genotypes (V. rupestris x V. arizonica) in response to Xf infection. A total of 5,794 sequences were obtained from these cDNA libraries from which 993 contigs and 949 singletons were derived. Using Gene Ontology (GO) hierarchy, the non-redundant sequences were classified into the three principal categories: molecular function (30%), cellular components (9%) and biological processes (7%). Comparative analysis found variations in EST expression pattern between infected and non-infected PD resistant and PD susceptible grape genotypes. Among the three tissues, libraries from stem tissues showed significant differences in transcript quality suggesting their important role in grape-Xylella interaction.
This study constitutes the first attempt to characterize the Vitis differential transcriptome associated with host-pathogen interactions from different explants and genotypes. All the generated ESTs have been submitted to GenBank and are also available through our website for further functional studies.

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Available from: M. Andrew Walker, Nov 02, 2015
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    • "In grapevine, more than 350,000 EST sequences have been generated and analyzed to identify gene expression related to a wide range of processes including berry development in wine grape [30,45] and in table grape [46], tissue-specific gene expression [6,42], the fulfillment of chilling requirements in dormant grape buds [34], and the characterization of resistance to pathogens such as Xylella fastidiosa [47] and Rhizobium vitis [44]. To discern how steady-state transcript accumulation changes in response to multiple environmental stress treatments, we generated a total of 45,784 ESTs from leaf and berry tissues from vines subjected to abiotic stresses (e.g., salinity, cold, heat, water deficit, and anoxia). "
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    ABSTRACT: Abiotic stresses, such as water deficit and soil salinity, result in changes in physiology, nutrient use, and vegetative growth in vines, and ultimately, yield and flavor in berries of wine grape, Vitis vinifera L. Large-scale expressed sequence tags (ESTs) were generated, curated, and analyzed to identify major genetic determinants responsible for stress-adaptive responses. Although roots serve as the first site of perception and/or injury for many types of abiotic stress, EST sequencing in root tissues of wine grape exposed to abiotic stresses has been extremely limited to date. To overcome this limitation, large-scale EST sequencing was conducted from root tissues exposed to multiple abiotic stresses. A total of 62,236 expressed sequence tags (ESTs) were generated from leaf, berry, and root tissues from vines subjected to abiotic stresses and compared with 32,286 ESTs sequenced from 20 public cDNA libraries. Curation to correct annotation errors, clustering and assembly of the berry and leaf ESTs with currently available V. vinifera full-length transcripts and ESTs yielded a total of 13,278 unique sequences, with 2302 singletons and 10,976 mapped to V. vinifera gene models. Of these, 739 transcripts were found to have significant differential expression in stressed leaves and berries including 250 genes not described previously as being abiotic stress responsive. In a second analysis of 16,452 ESTs from a normalized root cDNA library derived from roots exposed to multiple, short-term, abiotic stresses, 135 genes with root-enriched expression patterns were identified on the basis of their relative EST abundance in roots relative to other tissues. The large-scale analysis of relative EST frequency counts among a diverse collection of 23 different cDNA libraries from leaf, berry, and root tissues of wine grape exposed to a variety of abiotic stress conditions revealed distinct, tissue-specific expression patterns, previously unrecognized stress-induced genes, and many novel genes with root-enriched mRNA expression for improving our understanding of root biology and manipulation of rootstock traits in wine grape. mRNA abundance estimates based on EST library-enriched expression patterns showed only modest correlations between microarray and quantitative, real-time reverse transcription-polymerase chain reaction (qRT-PCR) methods highlighting the need for deep-sequencing expression profiling methods.
    Full-text · Article · May 2011 · BMC Plant Biology
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    • "Lipid-derived molecules act as signals in plantpathogen interactions, and the roles of jasmonic acid and related oxylipins that are produced from membrane-derived fatty acids through beta-oxidation, are particularly important [21]. During infection, low level defence responses can be activated in susceptible plants [22,23]. Therefore, it is likely that well-established "Ca. "
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    Full-text · Article · Jan 2011 · BMC Microbiology
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    • "Based on the previous report [14], a genomic DNA sequence coding for a specific cytochrome P450 gene was identified from a grape genomic database (GenBank Accessions: AM475392.1, CAAP02000243.1, "
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    ABSTRACT: Plant cytochrome P450 monooxygenases (CYP) mediate synthesis and metabolism of many physiologically important primary and secondary compounds that are related to plant defense against a range of pathogenic microbes and insects. To determine if cytochrome P450 monooxygenases are involved in defense response to Xylella fastidiosa (Xf) infection, we investigated expression and regulatory mechanisms of the cytochrome P450 monooxygenase CYP736B gene in both disease resistant and susceptible grapevines. Cloning of genomic DNA and cDNA revealed that the CYP736B gene was composed of two exons and one intron with GT as a donor site and AG as an acceptor site. CYP736B transcript was up-regulated in PD-resistant plants and down-regulated in PD-susceptible plants 6 weeks after Xf inoculation. However, CYP736B expression was very low in stem tissues at all evaluated time points. 5'RACE and 3'RACE sequence analyses revealed that there were three candidate transcription start sites (TSS) in the upstream region and three candidate polyadenylation (PolyA) sites in the downstream region of CYP736B. Usage frequencies of each transcription initiation site and each polyadenylation site varied depending on plant genotype, developmental stage, tissue, and treatment. These results demonstrate that expression of CYP736B is regulated developmentally and in response to Xf infection at both transcriptional and post-transcriptional levels. Multiple transcription start and polyadenylation sites contribute to regulation of CYP736B expression. This report provides evidence that the cytochrome P450 monooxygenase CYP736B gene is involved in defense response at a specific stage of Xf infection in grapevines; multiple transcription initiation and polyadenylation sites exist for CYP736B in grapevine; and coordinative and selective use of transcription initiation and polyadenylation sites play an important role in regulation of CYP736B expression during growth, development and response to Xf infection.
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