Transcriptomic footprints disclose specificity of reactive oxygen species signaling in Arabidopsis

Abstract

Reactive oxygen species (ROS) are key players in the regulation of plant development, stress responses, and programmed cell death. Previous studies indicated that depending on the type of ROS (hydrogen peroxide, superoxide, or singlet oxygen) or its subcellular production site (plastidic, cytosolic, peroxisomal, or apoplastic), a different physiological, biochemical, and molecular response is provoked. We used transcriptome data generated from ROS-related microarray experiments to assess the specificity of ROS-driven transcript expression. Data sets obtained by exogenous application of oxidative stress-causing agents (methyl viologen, Alternaria alternata toxin, 3-aminotriazole, and ozone) and from a mutant (fluorescent) and transgenic plants, in which the activity of an individual antioxidant enzyme was perturbed (catalase, cytosolic ascorbate peroxidase, and copper/zinc superoxide dismutase), were compared. In total, the abundance of nearly 26,000 transcripts of Arabidopsis (Arabidopsis thaliana) was monitored in response to different ROS. Overall, 8,056, 5,312, and 3,925 transcripts showed at least a 3-, 4-, or 5-fold change in expression, respectively. In addition to marker transcripts that were specifically regulated by hydrogen peroxide, superoxide, or singlet oxygen, several transcripts were identified as general oxidative stress response markers because their steady-state levels were at least 5-fold elevated in most experiments. We also assessed the expression characteristics of all annotated transcription factors and inferred new candidate regulatory transcripts that could be responsible for orchestrating the specific transcriptomic signatures triggered by different ROS. Our analysis provides a framework that will assist future efforts to address the impact of ROS signals within environmental stress conditions and elucidate the molecular mechanisms of the oxidative stress response in plants.

Figure 1.

Transcript level changes within the individual ROS experiments. The number of transcripts with a 3-, 4-, or 5-fold increase or decrease in expression in transgenic or treated plants compared to wild-type or control plants at different time points are represented by black, gray, and white bars, respectively. Data were processed as described in “Materials and Methods.” FC, Fold change.

Figure 2.

Hierarchical clustering of the 3,925 transcripts across the different ROS-generating conditions. Transcripts with at least a 5-fold difference in expression within one time point across all experiments were subjected to hierarchical average linkage clustering. Each horizontal line displays the expression data for one gene. Red and green indicate up- and down-regulation in transgenic or treated plants compared to wild-type or untreated plants, respectively. Intensity of the colors is proportional to the absolute value of the fold difference. Gray corresponds to missing expression values for transcripts that are not represented on either the Affymetrix ATH1 or Agilent Arabidopsis2 microarrays. The horizontal dendrogram (top) indicates the relationship among the experiments across all transcripts included in the cluster analysis. Three main clusters (I, II, and III) are indicated.

Figure 3.

Relative abundance of specific ROS in abiotic stresses. The relative contribution of specific ROS-induced transcripts that respond positively or negatively to different abiotic stresses is presented. For each abiotic stress, the total numbers of ROS-responsive transcripts are indicated. Black, gray, and white bars represent O₂^·−-, H₂O₂-, and ¹O₂-responsive transcripts, respectively. Data were processed as described in “Materials and Methods.”