Comparative Transcriptomics of Multi-Stress Responses in Pachycladon cheesemanii and Arabidopsis thaliana

Abstract

The environment is seldom optimal for plant growth and changes in abiotic and biotic signals, including temperature, water availability, radiation and pests, induce plant responses to optimise survival. The New Zealand native plant species and close relative to Arabidopsis thaliana, Pachycladon cheesemanii, grows under environmental conditions that are unsustainable for many plant species. Here, we compare the responses of both species to different stressors (low temperature, salt and UV-B radiation) to help understand how P. cheesemanii can grow in such harsh environments. The stress transcriptomes were determined and comparative transcriptome and network analyses discovered similar and unique responses within species, and between the two plant species. A number of widely studied plant stress processes were highly conserved in A. thaliana and P. cheesemanii. However, in response to cold stress, Gene Ontology terms related to glycosinolate metabolism were only enriched in P. cheesemanii. Salt stress was associated with alteration of the cuticle and proline biosynthesis in A. thaliana and P. cheesemanii, respectively. Anthocyanin production may be a more important strategy to contribute to the UV-B radiation tolerance in P. cheesemanii. These results allowed us to define broad stress response pathways in A. thaliana and P. cheesemanii and suggested that regulation of glycosinolate, proline and anthocyanin metabolism are strategies that help mitigate environmental stress.

Keywords: Arabidopsis; Pachycladon; comparative transcriptomics; cross-species comparison; multi-stress responses; network analysis.

Figure 1

Differential gene expression of P. cheesemanii and A. thaliana caused by different stresses. Volcano plots, showing fold change in abundance (logFC) against significance, indicate DE genes in response to cold, salt and UV-B radiation stress. FC: fold change; FDR: false discovery rate. Upper row: A. thaliana; lower row: P. cheesemanii. The differentially expressed genes were grouped into 4 levels with different colours: Level I (orange), with 2 ≤ |FC| < 4; Level II (blue), with 4 ≤ |FC| < 32; Level III (green), with 32 ≤ |FC| < 1024 and Level IV (purple), with 1024 ≤ |FC|. FC, fold change.

Figure 2

Venn diagrams of up- and downregulated genes in response to stress in A. thaliana and P. cheesemanii. The diagrams show the number of up- and downregulated genes and overlap in response to cold, salt, and UV-B radiation. Upper row: A. thaliana; lower row: P. cheesemanii.

Figure 3

Venn diagram of the numbers of GO terms of biological processes between stresses in A. thaliana and P. cheesemanii. AT: A. thaliana; PC: P. cheesemanii. UV circles: the number of overrepresented GO terms in UV-B radiation response; Cold circles: the number of overrepresented GO terms in cold response; Salt circles: the number of overrepresented GO terms in salt response.

Figure 4

Network analysis of upregulated biological processes of A. thaliana multiple stress-responsive transcriptomes. Purple, red and blue circles: overrepresented GO terms in response to UV-B radiation, salt and cold stress, respectively. Boxes with dashed lines: clusters of overrepresented GO terms involved in similar biological processes.

Figure 5

Network analysis of biological processes of P. cheesemanii multiple stress-responsive transcriptomes. Purple, red and blue circles: overrepresented GO terms in response to UV-B radiation, salt and cold stress, respectively. Boxes with dashed lines: clusters of overrepresented GO terms involved in similar biological processes.

Figure 6

Venn diagrams of the numbers of overrepresented GO terms of A. thaliana and P. cheesemanii in response to different stresses. Species-specific and overlapping numbers of overrepresented GO terms in response to cold, salt and UV-B radiation stress. AT: A. thaliana; PC: P. cheesemanii; UV circles: the number of overrepresented GO terms in UV-B radiation (UV) response; Cold circles: the number of overrepresented GO terms in cold stress response; salt circles: the number of overrepresented GO terms in salt stress response.

Figure 7

Common GO biological processes of A. thaliana and P. cheesemanii in response to cold, salt and UV-B radiation stress. Clustering of the common overrepresented terms of GO biological processes of A. thaliana and P. cheesemanii responding to cold, salt, and UV-B radiation stress. (a) Eighty-four common overrepresented GO terms in responding to cold stress were clustered with fourteen representatives of GO biological processes. (b) Twenty-six common overrepresented GO terms in responding to salt stress were clustered with four representatives of GO biological processes. (c) Sixty-five common overrepresented GO terms in responding to UV-B radiation were clustered with fourteen representatives of GO biological processes. Semantic space X and Y: no intrinsic meaning; uniqueness: measure whether the term is an outlier compared to the list, namely, the negative of average similarity of a term to all other terms. In REVIGO, multi-dimensional scaling was used to reduce the dimensionality of a matrix of the GO terms’ pairwise semantic similarities. First, the terms were placed by using an eigenvalue decomposition of the terms’ pairwise distance matrix. Then, a stress minimization step improved the agreement between the semantic similarities of the terms and their closeness in the two-dimensional space. Thus, the semantically similar GO terms should remain close together in the plot. Figure was generated from REVIGO web (http://revigo.irb.hr/, accessed on 20 October 2021) [30].

Figure 8

Unique overrepresented terms of GO biological process of A. thaliana and P. cheesemanii in response to cold, salt and UV-B radiation stress. Clustering of the unique overrepresented terms of GO biological process of A. thaliana and P. cheesemanii responding to cold, salt and UV-B radiation stress. (a) A. thaliana Cold vs. P. cheesemanii Cold: 32 and 150 unique overrepresented GO terms of A. thaliana and P. cheesemanii in responding to cold stress were clustered with 7 and 12 representatives of GO biological processes. (b) A. thaliana Salt vs. P. cheesemanii Salt: 90 and 52 unique overrepresented GO terms of A. thaliana and P. cheesemanii in responding to salt stress were clustered with 11 and 9 representatives of GO biological processes. (c) A. thaliana UV-B vs. P. cheesemanii UV-B: 114 and 57 unique overrepresented GO terms of A. thaliana and P. cheesemanii in responding to UV-B radiation were clustered with 13 and 5 representatives of GO biological processes. Semantic space X and Y: no intrinsic meaning; uniqueness: measure whether the term is an outlier compared to the list. Namely, the negative of average similarity of a term to all other terms. In REVIGO, multi-dimensional scaling was used to reduce the dimensionality of a matrix of the GO terms’ pairwise semantic similarities. First, the terms were placed by using an eigenvalue decomposition of the terms’ pairwise distance matrix. Then, a stress minimization step improved the agreement between the semantic similarities of the terms and their closeness in the two-dimensional space. Thus, the semantically similar GO terms should remain close together in the plot. Figure was generated from REVIGO web (http://revigo.irb.hr/) [30].

Figure 8

Unique overrepresented terms of GO biological process of A. thaliana and P. cheesemanii in response to cold, salt and UV-B radiation stress. Clustering of the unique overrepresented terms of GO biological process of A. thaliana and P. cheesemanii responding to cold, salt and UV-B radiation stress. (a) A. thaliana Cold vs. P. cheesemanii Cold: 32 and 150 unique overrepresented GO terms of A. thaliana and P. cheesemanii in responding to cold stress were clustered with 7 and 12 representatives of GO biological processes. (b) A. thaliana Salt vs. P. cheesemanii Salt: 90 and 52 unique overrepresented GO terms of A. thaliana and P. cheesemanii in responding to salt stress were clustered with 11 and 9 representatives of GO biological processes. (c) A. thaliana UV-B vs. P. cheesemanii UV-B: 114 and 57 unique overrepresented GO terms of A. thaliana and P. cheesemanii in responding to UV-B radiation were clustered with 13 and 5 representatives of GO biological processes. Semantic space X and Y: no intrinsic meaning; uniqueness: measure whether the term is an outlier compared to the list. Namely, the negative of average similarity of a term to all other terms. In REVIGO, multi-dimensional scaling was used to reduce the dimensionality of a matrix of the GO terms’ pairwise semantic similarities. First, the terms were placed by using an eigenvalue decomposition of the terms’ pairwise distance matrix. Then, a stress minimization step improved the agreement between the semantic similarities of the terms and their closeness in the two-dimensional space. Thus, the semantically similar GO terms should remain close together in the plot. Figure was generated from REVIGO web (http://revigo.irb.hr/) [30].

Figure 9

Summary of responses of A. thaliana and P. cheesemanii plants to cold, salt, and UV-B radiation stresses. Main boxes indicate the three different types of stresses. In each box, main common enrichments in GO terms are indicated in the center, while unique GO enrichments are indicated towards the left for A. thaliana and the right for P. cheesemanii. See text for details.