15 research outputs found
Roles of non-coding RNA in sugarcane-microbe interaction
Studies have highlighted the importance of non-coding RNA regulation in plant-microbe
interaction. However, the roles of sugarcane microRNAs (miRNAs) in the regulation of disease
responses have not been investigated. Firstly, we screened the sRNA transcriptome of sugarcane
infected with Acidovorax avenae. Conserved and novel miRNAs were identified. Additionally,
small interfering RNAs (siRNAs) were aligned to differentially expressed sequences from the
sugarcane transcriptome. Interestingly, many siRNAs aligned to a transcript encoding a coppertransporter
gene whose expression was induced in the presence of A. avenae, while the siRNAs were
repressed in the presence of A. avenae. Moreover, a long intergenic non-coding RNA was identified
as a potential target or decoy of miR408. To extend the bioinformatics analysis, we carried out
independent inoculations and the expression patterns of six miRNAs were validated by quantitative
reverse transcription-PCR (qRT-PCR). Among these miRNAs, miR408âa copper- microRNAâwas
downregulated. The cleavage of a putative miR408 target, a laccase, was confirmed by a modified
50RACE (rapid amplification of cDNA ends) assay. MiR408 was also downregulated in samples
infected with other pathogens, but it was upregulated in the presence of a beneficial diazotrophic
bacteria. Our results suggest that regulation by miR408 is important in sugarcane sensing whether
microorganisms are either pathogenic or beneficial, triggering specific miRNA-mediated regulatory
mechanisms accordingly
Sugarcane transcriptome analysis in response to infection caused by <i>Acidovorax avenae</i> subsp. <i>avenae</i>
<div><p>Sugarcane is an important tropical crop mainly cultivated to produce ethanol and sugar. Crop productivity is negatively affected by <i>Acidovorax avenae</i> subsp <i>avenae</i> (<i>Aaa</i>), which causes the red stripe disease. Little is known about the molecular mechanisms triggered in response to the infection. We have investigated the molecular mechanism activated in sugarcane using a RNA-seq approach. We have produced a <i>de novo</i> transcriptome assembly (TR7) from sugarcane RNA-seq libraries submitted to drought and infection with <i>Aaa</i>. Together, these libraries present 247 million of raw reads and resulted in 168,767 reference transcripts. Mapping in TR7 of reads obtained from infected libraries, revealed 798 differentially expressed transcripts, of which 723 were annotated, corresponding to 467 genes. GO and KEGG enrichment analysis showed that several metabolic pathways, such as code for proteins response to stress, metabolism of carbohydrates, processes of transcription and translation of proteins, amino acid metabolism and biosynthesis of secondary metabolites were significantly regulated in sugarcane. Differential analysis revealed that genes in the biosynthetic pathways of ET and JA PRRs, oxidative burst genes, NBS-LRR genes, cell wall fortification genes, SAR induced genes and pathogenesis-related genes (PR) were upregulated. In addition, 20 genes were validated by RT-qPCR. Together, these data contribute to a better understanding of the molecular mechanisms triggered by the <i>Aaa</i> in sugarcane and opens the opportunity for the development of molecular markers associated with disease tolerance in breeding programs.</p></div
The ten most frequently occurred protein domains in the sequence of sugarcane infected by <i>Aaa</i>.
<p>The ten most frequently occurred protein domains in the sequence of sugarcane infected by <i>Aaa</i>.</p
Validation of RNA-seq analysis by qRT-PCR using genes from different pathways.
<p>Two biological replicates were used. Gene names correspond to those listed in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0166473#pone.0166473.s009" target="_blank">S9</a> and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0166473#pone.0166473.s010" target="_blank">S10</a> Tables. Relative expression by qRT-PCR. The bars represent the relative expression of three technical replicates (n = 3) and standard deviation (Green bars: replicate 1 and blue bars: replicate 2). The relative expression values above the dotted line are upregulated genes, whereas below line correspond to downregulated genes. GAPDH was used as a reference gene for normalization of gene-expression data. These 20 genes validated in replicates were grouped into four categories, <b>(A)</b> genes related to stress, <b>(B)</b> genes that coding to several pathways, <b>(C)</b> primary carbohydrate metabolism pathways genes and <b>(D)</b> genes encoding for PRRs. The values of the quantitative method ÎÎCt can be seen in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0166473#pone.0166473.s010" target="_blank">S10 Table</a></p
Histogram presentation of the GO enrichment analysis of sugarcane plantlets infected by <i>Aaa</i>.
<p>TRAPID system calculated GO enrichment based on the upregulated and downregulated dataset compared to a background (p-value 0.01). The x-axis indicates the percent of genes and the y-axis indicates the GO terms. GO analysis to <b>(A)</b> upregulated and <b>(B)</b> downregulated DEGs under biotic stress. For additional details, see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0166473#pone.0166473.s004" target="_blank">S4 Table</a>.</p
Histogram presentation of the 8 KEGG metabolic pathways significantly enriched to DEGs of sugarcane plantlets infected by pathogen.
<p>The x-axis indicates the number of genes assigned to a specific pathway, the y-axis indicates the KEGG pathway. Enriched metabolic pathways to <b>(A)</b> upregulated and <b>(B)</b> downregulated DEGs. For additional details, see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0166473#pone.0166473.s005" target="_blank">S5 Table</a>.</p
Workflow of analysis of the construction and analyses of the sugarcane reference TR7.
<p><b>(A)</b> The transcriptome assembly <i>De novo</i> was generated from sugarcane RNA libraries drought treated and libraries <i>Aaa</i> pathogen treated obtained from of Illumina. After was applied to quality filter to all raw reads the quality filter and next, were removed duplicate genome sequences from the dataset. The Velvet and Oases software was used for the <i>de novo</i> assembly of clean reads to generate the sugarcane reference transcriptome TR7 with 168,767 transcripts. <b>(B)</b> Differential expression analysis and annotated functional in TRAPID. About 14 millions of reads were mapping no TR7. Transcripts differentially expressed were selected by Fisherâs exact-test, p-value < 0.01 and transcripts that have the same expression on both replicas.</p
The top six up- and downregulated differentially expressed genes in sugarcane infected by <i>Aaa</i>.
<p>The top six up- and downregulated differentially expressed genes in sugarcane infected by <i>Aaa</i>.</p
Regulation of photosynthesis and carbohydrate metabolism in sugarcane infected with <i>Aaa</i>.
<p>Altered expression of genes in photosynthesis, glycolysis, metabolism starch, metabolism of raffinose, metabolism trehalose, sucrose metabolism, transport of sugar and mitochondrial respiratory chain. Red for downregulated and blue for upregulated genes. For additional details of the genes, see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0166473#pone.0166473.s007" target="_blank">S7 Table</a>.</p