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. 2016 Dec 2;13(1):202.
doi: 10.1186/s12985-016-0663-7.

RNA-seq-based digital gene expression analysis reveals modification of host defense responses by rice stripe virus during disease symptom development in Arabidopsis

Affiliations

RNA-seq-based digital gene expression analysis reveals modification of host defense responses by rice stripe virus during disease symptom development in Arabidopsis

Feng Sun et al. Virol J. .

Abstract

Background: Virus infection induces and suppresses host gene expression on a global level. Rice stripe virus (RSV) is the type species of the genus Tenuivirus and infects rice and Arabidopsis plants. Microarray-based and next generation sequencing-based transcriptomic approaches have been used to study rice-RSV interactions. However, our knowledge of the response of Arabidopsis plants to RSV infection is limited, and it requires further investigation to determine the similarities (or differences) in virus-host interactions between monocot and dicot hosts infected with RSV.

Methods: We characterized transcriptome changes in Arabidopsis thaliana infected with rice stripe virus (RSV) with RNA-seq based digital gene expression (DGE) analysis. The transcriptomes of RSV-infected samples were compared to those of mock-treated samples at 14 and 21 days post-infection (dpi) during different stages of symptom development.

Results: We identified 624 differentially expressed genes (DEGs) in Arabidopsis influenced by RSV at 14 dpi and 21 dpi, among which at 14 dpi, 255 transcripts were induced, and 38 were repressed; at 21 dpi, 146 were induced, and 237 were repressed. Functional annotation indicated that these DEGs were related to multiple biological functions, including defense response, secondary metabolism, protein amino acid phosphorylation and response to abiotic stress.

Conclusions: Importantly, the transcription of genes related to host defense systems was activated by RSV infection at an early stage of symptom development (14 dpi), whereas over the infection period (21 dpi), the host defense response systems were suppressed. A total of 52 genes were continuously differentially expressed between the two time points, indicating that the majority of DEGs were transient and unique to a particular time point during symptom development. The DEGs, particularly the defense response genes, identified in this study are candidates suitable for further functional analysis during the RSV-Arabidopsis interaction.

Keywords: Defense response; Digital gene expression (DGE); RNA-seq; Rice stripe virus.

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Figures

Fig. 1
Fig. 1
Rice stripe virus (RSV) infection in Arabidopsis thaliana. a The left panel shows symptom of A. thaliana plants inoculated with RSV, and the right panel shows the mock-inoculated plants. b RSV accumulation was estimated in Arabidopsis plants using Western blotting with a RSV specific antibody. The actin protein level served as a loading control. c qRT-PCR for expression of RSV CP and SP genes in infected Arabidopsis plants. Signal intensities for each transcript were normalized with EF1-α and actin2. d Accumulation of RSV titer in infected Arabidopsis plants by ELISA
Fig. 2
Fig. 2
Venn diagram depicting the distribution of 624 differentially expressed genes (p<0.05) in RSV-infected leaf tissue at two time points post infection. a 388 induced transcripts. b 271 repressed transcripts
Fig. 3
Fig. 3
Functional distribution of DEGs in RSV-infected Arabidopsis plants at 14 and 21 dpi
Fig. 4
Fig. 4
DAVID functional annotation categories of DEGs in RSV-infected Arabidopsis plants. Significantly enriched categories for (a) up-regulated genes at 14 dpi; (b) down-regulated genes at 14 dpi; (c) up-regulated genes at 21 dpi and (d) down-regulated genes at 21 dpi
Fig. 5
Fig. 5
Singular enrichment analysis (SEA) of the DEGs involved in defense response processes at 14 dpi (a) and 21 dpi (b) using agriGO
Fig. 6
Fig. 6
Heat map showing hierarchical clustering of 26 transcripts differentially expressed during both time points (14 and 21 dpi). Red bars indicate induction (>2.0), and green bars indicate repression (<2.0)
Fig. 7
Fig. 7
Validation of Illumina RNA-seq expression data by quantitative reverse-transcription RT-PCR (qRT-PCR). Expression patterns selected transcripts that were similar between the two technologies are shown. Signal intensities for each transcript were normalized with EF1-α and actin2. The x-axis shows the validated genes at 14 and 21 dpi. The y-axis is the normalized fold-change expression values for each transcript

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