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. 2008 Jul 9:9:325.
doi: 10.1186/1471-2164-9-325.

Altered gene expression changes in Arabidopsis leaf tissues and protoplasts in response to Plum pox virus infection

Mohan Babu  1 , Jonathan S Griffiths, Tyng-Shyan Huang, Aiming Wang
Affiliations

Altered gene expression changes in Arabidopsis leaf tissues and protoplasts in response to Plum pox virus infection

Mohan Babu et al. BMC Genomics. .

Abstract

Background: Virus infection induces the activation and suppression of global gene expression in the host. Profiling gene expression changes in the host may provide insights into the molecular mechanisms that underlie host physiological and phenotypic responses to virus infection. In this study, the Arabidopsis Affymetrix ATH1 array was used to assess global gene expression changes in Arabidopsis thaliana plants infected with Plum pox virus (PPV). To identify early genes in response to PPV infection, an Arabidopsis synchronized single-cell transformation system was developed. Arabidopsis protoplasts were transfected with a PPV infectious clone and global gene expression changes in the transfected protoplasts were profiled.

Results: Microarray analysis of PPV-infected Arabidopsis leaf tissues identified 2013 and 1457 genes that were significantly (Q < or = 0.05) up- (> or = 2.5 fold) and downregulated (< or = -2.5 fold), respectively. Genes associated with soluble sugar, starch and amino acid, intracellular membrane/membrane-bound organelles, chloroplast, and protein fate were upregulated, while genes related to development/storage proteins, protein synthesis and translation, and cell wall-associated components were downregulated. These gene expression changes were associated with PPV infection and symptom development. Further transcriptional profiling of protoplasts transfected with a PPV infectious clone revealed the upregulation of defence and cellular signalling genes as early as 6 hours post transfection. A cross sequence comparison analysis of genes differentially regulated by PPV-infected Arabidopsis leaves against uniEST sequences derived from PPV-infected leaves of Prunus persica, a natural host of PPV, identified orthologs related to defence, metabolism and protein synthesis. The cross comparison of genes differentially regulated by PPV infection and by the infections of other positive sense RNA viruses revealed a common set of 416 genes. These identified genes, particularly the early responsive genes, may be critical in virus infection.

Conclusion: Gene expression changes in PPV-infected Arabidopsis are the molecular basis of stress and defence-like responses, PPV pathogenesis and symptom development. The differentially regulated genes, particularly the early responsive genes, and a common set of genes regulated by infections of PPV and other positive sense RNA viruses identified in this study are candidates suitable for further functional characterization to shed lights on molecular virus-host interactions.

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Figures

Figure 1
Figure 1
Systemic infection and detection of PPV in Arabidopsis thaliana plants. (A) Symptoms on mock-inoculated (left) and PPV-infected (right) plants 17 days post inoculation (dpi). (B) RT-PCR and ELISA analysis of PPV-infected and mock-inoculated leaf tissues at 17 dpi. BR1, BR2, BR3, represents three independent biological replicates of PPV-infected and mock-inoculated leaf tissues, respectively. RT-PCR amplification of a 467 bp fragment corresponding to a segment of the PPV genome (nt 9140 to 9607), as tested using 25 cycles of amplification. +, ELISA positive; -; ELISA negative. (C) sqRT-PCR amplification of a cDNA fragment of the PPV genome isolated from Arabidopsis protoplasts transfected with pPPV-SK68 and pPPV-SK68Δ at 3, 6 and 12 hours post transfection (hpt), respectively. Histone 3 gene was used as a loading control. RT-PCR amplification for both panel B and C were carried out using the same RNA samples that were used in microarray hybridizations.
Figure 2
Figure 2
Functional distribution of Arabidopsis genes significantly induced and repressed in PPV-infected leaves. The genes were grouped following the methods of the MIPS Arabidopsis classification scheme. Genes whose function has not been determined were grouped under "unknown function". Number of genes identified in each functional group is indicated on the x-axis. Genes that belong to major functional categories are highlighted in bold and the subcategories within a major functional category are highlighted in italics.
Figure 3
Figure 3
Venn diagrams depicting the distribution of induced (≥ 2.5 fold) and repressed (≤ -2.5 fold) genes in PPV-infected protoplasts at three different time points. A statistical cut-off with a FDR at 5% that corresponds to Q ≤ 0.05, after Benjamini and Hochberg's correction was used to determine genes significantly differentially regulated in protoplasts transfected with a PPV infectious cDNA clone, pPPV-SK68 at 3, 6 and 12 hours post transfection (hpt). The number of genes in the non-overlapping sector represents unique significant genes at each time point, while the overlapping sectors represent genes that are in common at time points indicated. The number in parentheses in the Venn diagram circles corresponds to the genes induced or repressed in the PPV infected leaves.
Figure 4
Figure 4
Functional distribution of Arabidopsis genes significantly induced and repressed in PPV-infected protoplasts. The genes were grouped following the method of the MIPS Arabidopsis classification scheme. Genes whose function has not been determined were grouped under "unknown function". Number of genes identified in each functional group is indicated on the x-axis. Genes that belong to major functional categories are highlighted in bold and the subcategories within a major functional category are highlighted in italics.
Figure 5
Figure 5
Clustering analysis of differentially regulatedgenes in PPV-infected Arabidopsis protoplasts. (A) Hierarchical clustering and changes in gene expression of 411 significantly (Q ≤ 0.05) differentially regulated genes in PPV-infected Arabidopsis protoplasts at 3, 6 and 12 hours post transfection (hpt). Gene tree map generated from 411 significantly differentially regulated Arabidopsis genes are derived from 263 upregulated and 304 downregulated genes. (B) Using k-mean clustering, the gene expression profiles were grouped into twelve major cluster groups. The expression pattern of a gene in a cluster group is indicated in parentheses. Expression levels are color coded with red indicating upregulation by PPV infection; green indicating downregulation by PPV infection; and black indicating no change in expression. The intensity of color represents the degree of gene expression levels. List of genes induced or repressed at each time point in different cluster groups along with their expression levels and putative functions is provided in [Additional file 8].
Figure 6
Figure 6
Confirmation of relative expression levels of the transcripts selected from the microarray analysis with semi quantitative RT-PCR (sqRT-PCR). Expression changes of 8 selected genes were determined by sqRT-PCR and microarray. The signal intensity of each transcript was normalized using At3g18780 (Actin 2). The Arabidopsis Genome Initiative (AGI) locus identifier of each gene is provided. The y-axis indicates the normalized expression level of the transcript. The x-axis represents hours post transfection. Expression ratios are the average of three independent hybridizations ± standard deviation (SD). hpt, hours post transfection; dpi, days post inoculation. The error bars represent the standard deviation for the signals from each of the three independent hybridizations.
Figure 7
Figure 7
Confirmation of relative expression levels of the transcripts selected from the microarray analysis with semi quantitative RT-PCR (sqRT-PCR) or Northern blot hybridizations. Expression changes of 5 selected genes were determined by sqRT-PCR, microarray and Northern blot. The signal intensity of each transcript was normalized using At3g18780 (Actin 2). The Arabidopsis Genome Initiative (AGI) locus identifier of each gene is provided. The y-axis indicates the normalized expression level of the transcript. The x-axis represents hours post transfection or days post inoculation. Expression ratios are the average of three independent hybridizations ± standard deviation (SD). hpt, hours post transfection; dpi, days post inoculation. The error bars represent the standard deviation for the signals from each of the three independent hybridizations.
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References

    1. Maule A, Leh V, Lederer C. The dialogue between viruses and hosts in compatible interactions. Curr Opin Plant Biol. 2002;5:279–284. - PubMed
    1. Whitham SA, Yang C, Goodin MM. Global impact: elucidating plant responses to viral infection. Mol Plant Microbe Interact. 2006;19:1207–1215. - PubMed
    1. Stange C. Plant-virus interactions during the infective process. Cien Inv Agr. 2006;33:1–18.
    1. Culver JN, Padmanabhan MS. Virus-induced disease: Altering host physiology one interaction at a time. Annu Rev Phytopathol. 2007;45:221–243. - PubMed
    1. Whitham SA, Wang Y. Roles for host factors in plant viral pathogenicity. Curr Opin Plant Biol. 2004;7:365–371. - PubMed

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