Characterization of stress response at the transcriptome and proteome level in rice under a range of abiotic stress conditions

Introduction: The productivity of rice crops is threatened by multiple environmental stresses. In dryland crops, deep roots are the critical factor enabling extraction of soil water and thus, drought tolerance. Therefore, we investigated the proteomic and microRNA response of the root tips of rice varieties varying in drought tolerance following extended soil drying. We then tested the hypothesis that abscisic acid (ABA) pretreatment mimics drought responses at the proteome and transcriptome level. Investigation of direct drought and ABA-induced molecular responses produced a deeper understanding of stress response mechanisms in plants.

Methods: Proteins from root tips of young rice plants were extracted, peptides were TMT labelled and identified and quantified using high resolution orbitrap mass spectrometry, followed by peptide-to-spectrum matching. Total RNA was extracted and a small RNA library was created for micro-RNA profiling, with Illumina HiSeq2000 used to generate 50bp single-end reads.

In a second experiment, Nipponbare rice plants were grown and exposed to a suite of multiple abiotic stresses, either with or without pretreatment with the plant hormone ABA (abscisic acid). A total RNA library was prepared and sequenced using Illumina NovaSeq 6000 system to produce 75bp single-end reads. Protein was extracted and data independent acquisition (DIA) shotgun proteomics was performed using nanoLC and a triple-Tof 6600 mass spectrometer.

Preliminary data: In the primary study, root tips of plants from two different Oryza sativa varieties, IR64 and Azucena, were grown in root boxes and subjected to drought stress and recovery. Physiological variables measured included leaf water potential, photosynthetic and respiratory performance, growth rates, osmotic potential, electrolytic leakage and various enzyme assays. In a second experiment, plants were subjected to multiple abiotic stresses simultaneously, with or without prior stress-conditioning with ABA, and characterised in detail at both the proteomic and transcriptomic level.

Two miRNA modules found only in stress response in Azucena (miR164-MYB and miR164-NAC) are known to be regulated by ABA, while the targets of novel miRNAS identified include auxin responsive factors and NAC domain containing proteins. Several novel proteins involved in root elongation were also identified in Azucena, along with an antioxidant network and a metallo-sulfur cluster assembly linked to iron homoeostasis.

We found that pre-treatment with ABA significantly mitigated the leaf damage from combined abiotic stresses, and this was reflected at at the proteome level. The application of exogenous ABA did not greatly affect the proteome profile of the control plants, while in stress-exposed plants, a greater number of proteins changed in abundance, most of which were increased. Similarly, we also found that application of exogenous ABA to control plants under normal conditions did not alter the transcriptome profile significantly. The changes observed in the transcriptome of rice plants in response to multiple abiotic stress were made more pronounced by ABA pre-treatment, which induced the upregulation of a significant number of additional genes. Taken together, these results suggest that exogenous ABA has a potential priming effect for enhancing the rice tolerance of rice seedlings against combined abiotic stress, mainly by affecting stress-responsive mechanisms dependent on ABA signalling pathways in rice plants.


This allowed us to tease apart the tightly integrated networks of genes and proteins involved, highlighting the role of the TCA cycle and photosynthesis related proteins in complex networks acting in both stress response and ABA signalling.


Novel aspect: Integration of data from proteome and transcriptome analysis of abiotic stress in rice, producing novel insights into stress-response mechanisms.