Global Analysis of Transcriptome Responses and Gene Expression Profiles to Cold Stress of <i>Jatropha curcas</i> L.

<div><p>Background</p><p><i>Jatropha curcas</i> L., also called the Physic nut, is an oil-rich shrub with multiple uses, including biodiesel production, and is currently exploited as a renewable energy resource in many countries. Nevertheless, because of its origin from the tropical MidAmerican zone, <i>J. curcas</i> confers an inherent but undesirable characteristic (low cold resistance) that may seriously restrict its large-scale popularization. This adaptive flaw can be genetically improved by elucidating the mechanisms underlying plant tolerance to cold temperatures. The newly developed Illumina Hiseq™ 2000 RNA-seq and Digital Gene Expression (DGE) are deep high-throughput approaches for gene expression analysis at the transcriptome level, using which we carefully investigated the gene expression profiles in response to cold stress to gain insight into the molecular mechanisms of cold response in <i>J. curcas</i>.</p> <p>Results</p><p>In total, 45,251 unigenes were obtained by assembly of clean data generated by RNA-seq analysis of the <i>J. curcas</i> transcriptome. A total of 33,363 and 912 complete or partial coding sequences (CDSs) were determined by protein database alignments and ESTScan prediction, respectively. Among these unigenes, more than 41.52% were involved in approximately 128 known metabolic or signaling pathways, and 4,185 were possibly associated with cold resistance. DGE analysis was used to assess the changes in gene expression when exposed to cold condition (12°C) for 12, 24, and 48 h. The results showed that 3,178 genes were significantly upregulated and 1,244 were downregulated under cold stress. These genes were then functionally annotated based on the transcriptome data from RNA-seq analysis.</p> <p>Conclusions</p><p>This study provides a global view of transcriptome response and gene expression profiling of <i>J. curcas</i> in response to cold stress. The results can help improve our current understanding of the mechanisms underlying plant cold resistance and favor the screening of crucial genes for genetically enhancing cold resistance in <i>J. curcas</i>.</p> </div

The differentially expressed genes of <i>Jatropha curcas</i> after cold exposure for schemed time points.

<p>A: Venn diagram indicating the total number of differentially expressed genes after 12, 24, and 48 h of 12°C (≥2-fold change in expression). B: Classified number of upregulated and downregulated genes for each time point of the three cold treatments.</p

Gene Ontology classification of differentially expressed genes of <i>Jatropha curcas</i>.

<p>A: Biological Process. B: Molecular Function. C: Cellular Component. The vertical line indicates the percentage of corresponding genes.</p

Differential expression analysis of tags and unigenes by DGE.

<p>The expression level for each tag and unigene is shown in the volcano plots (a, c, e) and (b, d, f) after 12 (a, b), 24 (c, d), and 48 h (e, f) of cold stress. ‘Not DETs’ indicates ‘not detected expression tags’ and ‘Not DEGs’ indicates ‘not detected expression genes’. The horizontal line indicates the log₁₀ of transcripts per million of the control (A) and the vertical line indicates the log₁₀ of transcripts per million of 12 (B), 24 (C), and 48 h (D) of treatment at 12°C. The criteria for screening differentially expressed individuals are based on FDR ≤ 0.001 and the absolute value of log₂Ratio ≥ 1.</p

Venn diagrams showing differentially expressed <i>Jatropha curcas</i> genes after cold exposure for three time points.

<p>A: upregulated genes. B: downregulated genes.</p

Expressions of enzymes relevant to starch metabolism after 12, 24, and 48 h at 12°C.

<p>Expressions of enzymes relevant to starch metabolism after 12, 24, and 48 h at 12°C.</p

Clustering analysis of differential gene expression pattern relevant to ABA-independent signal transduction pathway.

<p>Clustering analysis of differential gene expression pattern relevant to ABA-independent signal transduction pathway.</p

Number and expression model of novel <i>Jatropha curcas</i> genes after cold exposure for schemed periods.

<p>A: Venn diagram indicating the total number of novel, expressed genes after 12, 24, and 48 h of 12°C. B: Expression model of common, novel genes at three time points. The vertical line indicates the raw expression of novel genes in each sample.</p

Simplified model of starch catabolism pathway.

<p>Simplified model of starch catabolism pathway.</p

Putative ABA-independent signal transduction pathway.

<p>Putative ABA-independent signal transduction pathway was constructed based on reference 31. HPK: two-component histidine kinase; RLK: receptor protein kinase; TRP: transmembrane responsive protein; PLC: phospholipase C; PLD: phospholipase D; CK I: casein kinase I; PIP₂: phosphatidylinositol 4,5-biphosphate; IP₃: inositol 1,4,5-trisphosphate; CDPK: calcium dependent protein kinase; PP2A: protein phosphatase 2A; PP2B: protein phosphatase 2B; PKC: protein kinase C; MAPK: mitogen-activated protein kinase; MAPKK: MAPK-kinase; MAPKKK: MAPKK kinase; ICEr: ICE recognition motifs; MYBRS: MYB recognition sequence; Hos9r1: Hos9 recognition motif 1; U: ubiquitin protein; S: SUMO protein; P: phosphoryl group; HOS1: RING E3 ligase; HOS10: R2R3-type MYB transcription factor; HOS15: WD40-repeat protein; FRY1/2: inositol polyphosphate 1-phosphatase; SIZ1: SUMO E3 ligase; SFR6: sensitive to freezing 6; Gcn5: histone acetyltransferase enzyme; ADA2/3: transcriptional adaptor protein 2/3.</p