Comparative Transcriptome Profiling of Chilling Stress Responsiveness in Two Contrasting Rice Genotypes

<div><p>Rice is sensitive to chilling stress, especially at the seedling stage. To elucidate the molecular genetic mechanisms of chilling tolerance in rice, comprehensive gene expressions of two rice genotypes (chilling-tolerant LTH and chilling-sensitive IR29) with contrasting responses to chilling stress were comparatively analyzed. Results revealed a differential constitutive gene expression prior to stress and distinct global transcription reprogramming between the two rice genotypes under time-series chilling stress and subsequent recovery conditions. A set of genes with higher basal expression were identified in chilling-tolerant LTH compared with chilling-sensitive IR29, indicating their possible role in intrinsic tolerance to chilling stress. Under chilling stress, the major effect on gene expression was up-regulation in the chilling- tolerant genotype and strong repression in chilling-sensitive genotype. Early responses to chilling stress in both genotypes featured commonly up-regulated genes related to transcription regulation and signal transduction, while functional categories for late phase chilling regulated genes were diverse with a wide range of functional adaptations to continuous stress. Following the cessation of chilling treatments, there was quick and efficient reversion of gene expression in the chilling-tolerant genotype, while the chilling-sensitive genotype displayed considerably slower recovering capacity at the transcriptional level. In addition, the detection of differentially-regulated TF genes and enriched <em>cis</em>-elements demonstrated that multiple regulatory pathways, including CBF and MYBS3 regulons, were involved in chilling stress tolerance. A number of the chilling-regulated genes identified in this study were co-localized onto previously fine-mapped cold-tolerance-related QTLs, providing candidates for gene cloning and elucidation of molecular mechanisms responsible for chilling tolerance in rice.</p> </div

Phenotypes of two rice genotypes under chilling stress and subsequent recovery.

<p>(A) Comparison of seedlings of LTH and IR29 in control, treated at 4°C for 8 h, 48 h and recovery for 24 h after 48 h treatment. (B) Cell membrane injury of LTH and IR29 treated at 4°C for 8 h and recovery for 24 h after 48 h treatment. (C) and (D) MDA and proline content of LTH and IR29 treated at 4°C for 8 h and recovery for 24 h after 48 h treatment. LTH cultivar (white) and IR29 cultivar (black). Values are means of 3 replicates. Vertical bars indicate standard error.</p

GO slims of functional categorization of the genes during the late response phase.

<p>The genes found to be commonly- or genotype-specifically-induced in LTH and IR29 during chilling stress of 8, 24, and 48 h. Bars show numbers of common (blue), LTH-specific (purple) and IR29-specific (yellow) induced genes. GO slim categories significantly overrepresented are calculated by a hypergeometric distribution and indicated by ***for P≤0.05.</p

Hierarchical cluster analysis of all DEGs in LTH and IR29.

<p>These include 8484 DEGs at 2, 8, 24, and 48 h time points during chilling-stress treatments and subsequent 24 h recovery. The median ratio (stressed/control) was log (base 2)-transformed and subjected to linkage hierarchical clustering. 1, 2, 3 indicate LTH at 8, 24, and 48 h chilling-stress time points, respectively; 4 and 5 indicate LTH and IR29 at 2 h chilling stress, respectively; 6 and 7 indicate LTH and IR29 during 24 h recovery, respectively; 8, 9, and 10 indicate IR29 at 8, 24, and 48 h chilling-stress time points, respectively.</p

Venn diagram of up- and down-regulated genes in LTH and IR29 under chilling stress conditions.

<p>(A) Up-regulated genes; (B) down-regulated genes.</p

GO slims of functional categorization of genes at the 2 h chilling-stress time point.

<p>The genes found to be commonly- or genotype-specifically-induced in LTH and IR29. Genes with unknown function are not included. Bars show numbers of common (blue), LTH-specific (purple) and IR29-specific (yellow) induced genes. GO slim categories significantly overrepresented are calculated by a hypergeometric distribution and indicated by ***for P≤0.05.</p