Chung Wen Lin

Comparison of early transcriptome responses to copper and cadmium in rice roots

The phytotoxic effects of copper (Cu) and cadmium (Cd) on plant growth are well documented. However, Cu and Cd toxicity targets and the cellular systems contributing to acquisition of tolerance are not fully understood at the molecular level. We aimed to identify genes and pathways that discriminate the actions of Cu and Cd in rice roots (Oryza sativa L. cv. TN67). The transcripts of 1,450 and 1,172 genes were regulated after Cu and Cd treatments, respectively. We identified 882 genes specifically respond to Cu treatment, and 604 unique genes as Cd-responsive by comparison of expression profiles of these two regulated gene groups. Gene ontology analysis for 538 genes involved in primary metabolism, oxidation reduction and response to stimulus was changed in response to both metals. In the individual aspect, Cu specifically altered levels of genes involved in vesicle trafficking transport, fatty acid metabolism and cellular component biogenesis. Cd-regulated genes related to unfolded protein binding and sulfate assimilation. To further characterize the functions of vesicle trafficking transport under Cu stress, interference of excytosis in root tissues was conducted by inhibitors and silencing of Exo70 genes. It was demonstrated that vesicle-trafficking is required for mediation of Cu-induced reactive oxygen species (ROS) production in root tissues. These results may provide new insights into understanding the molecular basis of the early metal stress response in plants.

Early signalling pathways in rice roots under vanadate stress.

Vanadate is beneficial to plant growth at low concentration. However, plant exposure to high concentrations of vanadate has been shown to arrest cell growth and lead to cell death. We are interested in understanding the signalling pathways of rice roots in response to vanadate stress. In this study, we demonstrated that vanadate induced rice root cell death and suppressed root growth. In addition, we found that vanadate induced ROS accumulation, increased lipid peroxidation and elicited a remarkable increase of MAPKs and CDPKs activities in rice roots. In contrast, pre-treatment of rice roots with ROS scavenger (sodium benzoate), serine/threonine protein phosphatase inhibitor (endothall), and CDPK antagonist (W7), reduced the vanadate-induced MAPKs activation. Furthermore, the expression of a MAPK gene (OsMPK3) and four tyrosine phosphatase genes (OsDSP3, OsDSP5, OsDSP6, and OsDSP10) were regulated by vanadate in rice roots. Collectively, these results strongly suggest that ROS, protein phosphatase, and CDPK may function in the vanadate-triggered MAPK signalling pathway cause cell death and retarded growth in rice roots.

PaPTP1, a Gene Encoding Protein Tyrosine Phosphatase from Orchid, Phalaenopsis amabilis, is Regulated During Floral Development and Induced by Wounding

Protein tyrosine phosphorylation is crucial in many cellular regulatory mechanisms, including development, stress management, and hormonal responses. However, the precise function of plant protein tyrosine phosphatase (PTP) has not been well elucidated. In this study, we report on the isolation and characterization of an orchid (Phalaenopsis amabilis) PTP gene, referred to as PaPTP1. The cloned PaPTP1 cDNA is 1,588 nucleotides long and the deduced protein sequence composed of 346 amino acid residues. The PaPTP1 gene encodes a classical plant PTP containing a catalytically active cysteine residue within the signature motif. Sequence homology analysis indicated that the predicted amino acid sequence is 57% identical to that of Arabidopsis AtPTP1. To our knowledge, the PaPTP1 is the first classical PTP identified in monocotyledon plant species. Southern blot analysis of genomic DNA revealed the presence of a single PaPTP1 gene in P. amabilis. Expression of PaPTP1 gene was examined in different organs of P. amabilis plants. The result showed that the PaPTP1 gene was abundantly expressed in labella and column of 24-month-old plants, but poorly in leaves. In addition, the PaPTP1 gene expression in response to wounding was investigated. The steady-state level of the PaPTP1 transcript increased significantly in wound-treated leaves. The identity as phosphatase was also demonstrated by characterizing the PaPTP1 expressed as a recombinant glutathione-S-transferase-fusion protein. The enzyme exhibited phosphatase activity towards a synthetic substrate, p-nitrophenolphosphate. Taken together, these results suggest that this PaPTP1 gene encodes a functional protein phosphatase, and its expression patterns are associated with the development of the orchid flowers and in response to wounding.

Common stress transcriptome analysis reveals functional and genomic architecture differences between early and delayed response genes

To identify the similarities among responses to diverse environmental stresses, we analyzed the transcriptome response of rice roots to three rhizotoxic perturbations (chromium, ferulic acid and mercury) and identified common early-transient, early-constant and delayed gene inductions. Common early response genes were mostly associated with signal transduction and hormones, and delayed response genes with lipid metabolism. Network component analysis revealed complicated interactions among common genes, the most highly connected signaling hubs being PP2C68, MPK5, LRR-RLK and NPR1. Gene architecture studies revealed different conserved promoter motifs and a different ratio of CpG island distribution between early and delayed genes. In addition, early-transient genes had more exons and a shorter first exon. IMEter was used to calculate the transcription regulation effects of introns, with greater effects for the first introns of early-transient than delayed genes. The higher Ka/Ks (non-synonymous/synonymous mutation) ratio of early-constant genes than early-transient, delayed and the genome median demonstrates the rapid evolution of early-constant genes. Our results suggest that finely tuned transcriptional control in response to environmental stress in rice depends on genomic architecture and signal intensity and duration.

Integrating Early Transcriptomic Responses to Rhizotoxins in Rice (Oryza sativa. L.) Reveals Key Regulators and a Potential Early Biomarker of Cadmium Toxicity

As sessile organisms, plants were constantly challenged with biotic and abiotic stresses. Transcriptional activation of stress-responsive genes is a crucial part of the plant adaptation to environmental changes. Here, early response of rice root to eight rhizotoxic stressors: arsenate, copper, cadmium, mercury, chromate, vanadate, ferulic acid and juglone, was analyzed using published microarray data. There were 539 general stress response (GSR) genes up-regulated under all eight treatments, including genes related to carbohydrate metabolism, phytohormone balance, and cell wall structure. Genes related to transcriptional coactivation showed higher Ka/Ks ratio compared to the other GSR genes. Network analysis discovered complicated interaction within GSR genes and the most connected signaling hubs were WRKY53, WRKY71, and MAPK5. Promoter analysis discovers enriched SCGCGCS cis-element in GSR genes. Moreover, GSR genes tend to be intronless and genes with shorter total intron length were induced in a higher level. Among genes uniquely up-regulated by a single stress, a phosphoenolpyruvate carboxylase kinase (PPCK) was identified as a candidate biomarker for detecting cadmium contamination. Our findings provide insights into the transcriptome dynamics of molecular response of rice to different rhizotoxic stress and also demonstrate potential use of comparative transcriptome analysis in identifying a novel potential early biomarker.