Transcriptome Analysis in Chinese Cabbage (Brassica rapa ssp. pekinensis) Provides the Role of Glucosinolate Metabolism in Response to Drought Stress

Abstract

Although drought stress is one of the most limiting factors in growth and production of Chinese cabbage (Brassica rapa L. ssp. pekinensis), the underlying biochemical and molecular causes are poorly understood. In the present study, to address the mechanisms underlying the drought responses, we analyzed the transcriptome profile of Chinese cabbage grown under drought conditions. Drought stress transcriptionally activated several transcription factor genes, including AP2/ERFs, bHLHs, NACs and bZIPs, and was found to possibly result in transcriptional variation in genes involved in organic substance metabolic processes. In addition, comparative expression analysis of selected BrbZIPs under different stress conditions suggested that drought-induced BrbZIPs are important for improving drought tolerance. Further, drought stress in Chinese cabbage caused differential acclimation responses in glucosinolate metabolism in leaves and roots. Analysis of stomatal aperture indicated that drought-induced accumulation of glucosinolates in leaves directly or indirectly controlled stomatal closure to prevent water loss, suggesting that organ-specific responses are essential for plant survival under drought stress condition. Taken together, our results provide information important for further studies on molecular mechanisms of drought tolerance in Chinese cabbage.

Keywords: Chinese cabbage; bZIP transcription factor; differentially expressed genes; drought stress; glucosinolate.

Figure 1

Physiological response to drought stress in Chinese cabbage. (A) Phenotypes of Chinese cabbage plants after exposure to drought stress for four days. S1 (Stage 1), normal water supply; S2 (Stage 2), water withholding for 2 d (soil water content at 20%); S3 (Stage 3), water withholding for 4 d (soil water content at 5%). Relative water content (B) and changes in levels of H₂O₂ (C), MDA (D), and protein carbonylation (E) after drought treatment were determined. Values are averages from three biological replicates consisting three plants per sample were used. The data are presented as mean ± standard error (SE). * p < 0.05, ** p < 0.01, and *** p < 0.001 represent the significant differences in comparison with Stage 1 (S1).

Figure 2

Analysis of differentially expressed genes (DEGs). (A) Distribution of up- and down-regulated DEGs in each comparison; (B) Venn diagram analysis of DEGs in different comparisons among groups; (C) Classification of DEGs based on metabolism categories. L1, L2, and L3, leaf samples obtained from Stage 1, Stage 2, and Stage 3 plants, respectively; R1, R2, and R3, root samples obtained from Stage 1, Stage 2, and Stage 3 plants, respectively.

Figure 3

Change in bZIP transcription factor (TF) expression during the response to abiotic stresses. (A) The heatmap shows the expression profiles of bZIP TFs in different comparisons among groups. The grey bar indicates bZIP TFs selected for qRT-PCR analysis; (B) The expression patterns of the selected bZIP TFs were analyzed using qRT-PCR. Transcript levels of the selected bZIP TFs were normalized to those of Chinese cabbage actin, and gene expression is relative to Stage 1 plants (L1 or R1) set to a value of 1 for each biological replicate. Values are averages from three independent biological experiments. Data are means ± SE. * p < 0.05, ** p < 0.01, and *** p < 0.001 represent the significant differences in comparison with L1 or R1; (C) Differential expression of the selected bZIP TFs under different stress treatments. Expression is indicated as a log2 ratio of experimental treatments relative to control samples and visualized in heatmaps showing hierarchical clustering of the selected bZIP TFs.

Figure 4

Mapman metabolism overview showing the differentially expressed genes (DEGs) in each comparison. The different colors represent the log2 values of the gene expression levels in response to drought stress.

Figure 5

Glucosinolate content in control (L1) and drought-treated (L3) Chinese cabbage leaves. Values are averages from three biological replicates. Error bars indicate S.E. * p < 0.05 and ** p < 0.01 indicate significantly different from L1.

Figure 6

Stomatal movement in Chinese cabbage in response to glucosinolate-derived products obtained from control (L1) and drought-treated (L3) Chinese cabbage leaves. Apertures were analyzed as width/length after 2 h of treatment. Values are means ± S.E., *** p < 0.001, n > 100.