Morphological, transcriptomic and proteomic responses of contrasting rice genotypes towards drought stress
Introduction
Based on the current scenario, human population is expected to reach up to 10 billion in next 30 years as predicted by U.S. Census Bureau, 2012 (Bureau, 2012). There is more emphasis on growing globally consumed crops to balance the food consumption and production rate. Rice is a dietary staple crop supporting more than 3 billion people globally (Khush, 2005). India, one of the leading producers of rice, contributes 158.7 million tons per year which is approximately 27% of total rice production in the world (FAOSTAT, 2018). The challenges to be overcome by India are to enhance rice production which comprise decrease rate of cultivation land, water deficiency, cultivation cost and storage loss due to climate change.
In comparison to various other grains such as maize, wheat, and sorghum, rice gets easily affected by drought due to the high requirement of water throughout its lifecycle. Drought is one of the major imminent threats for the plant growth, development, and yield. In the year 2015, in India, eleven states were affected by drought that constrained the nation’s food and water security (Sevekari, 2015). Effective abiotic stress management of rice crop can offer significant food security. Plants are often exposed to drought that affects them at various levels such as morphological (root and shoot length), anatomical (xylem and stomata), physiological (RWC), biochemical (proline accumulation) and molecular (protein and gene expression) (Pandey and Shukla, 2015). Plants are known to adopt four strategies to encounter drought - drought tolerance, drought avoidance, drought escape and drought recovery (Yambao et al., 1992). Therefore, the major focus of our study is to gain insight of the various traits that can play role in drought tolerance strategies that differentiate a drought stress tolerant rice variety from stress sensitive variety.
Conduction of water and nutrients affects the growth and development of plants significantly. Water uptake in plants is carried out by roots involving xylem mediated transportation as well as stomata mediated transpiration pull. Both, root and shoot system of a plant are interconnected and are affected by each other’s anatomy and physiology (Sinclair, 2011). Further, in case of rice roots, focus can be on two aspects: root architecture that is responsible for anchorage and root anatomy that plays role in water transport. Some of the root indices for describing root morphology such as length, average diameter, volume, and number contribute to some extent in the enrichment of drought management (Gray and Brady, 2016). Furthermore, it has been suggested that under drought condition, water retention capacity is improved by limiting the root water uptake (Henry et al., 2012). Therefore, quantitative analysis of root morphological characteristics is important to understand the connection between root morphology and its function. WinRhizo (Regent Instruments Inc., Quebec, Canada), an advanced root image analysis software that can be used for washed roots has been employed to determine root morphology such as length, area, volume and diameter in the present study.
Xylem plays an important role in water uptake and mechanical support. The alteration in xylem number and diameter in legume crops has been associated with adaptation mechanisms under drought conditions (Prince et al., 2017). Reduction in the development of xylem vessels under osmotic stress may also reduce embolism (Lovisolo et al., 1998). Therefore, the anatomical traits such as xylem number and area have been measured to investigate the genotypic variability under soil water regimes (Pouzoulet et al., 2014). An efficient allocation of resources in plants is maintained through water loss by stomata, the pores present in epidermis of the leaf for two major processes: influx of carbon dioxide needed in photosynthesis and maintain leaf temperature through water loss in transpiration. Plant leaves usually optimize the adaption against abiotic stress by stomatal pore opening or closing, aperture size, stomatal density and distribution pattern (Hetherington and Woodward, 2003). It has been also reported that morphological and physiological responses against stress are associated with stomatal characteristics. The plant species with crassulacean acid metabolism (CAM) have been shown to possess a lower stomatal number under water deficient condition (Silva et al., 2001). Relative water content (RWC) has been also measured for better characterization of plant root and shoot physiology under drought stress. It has been reported that the drought-tolerant varieties maintain a higher RWC during drought stress as compared to drought-sensitive varieties. Proline accumulation acts as one of the prominent metabolic response of plants against drought stress. Additionally, proline is an osmoprotectant that serves as a biochemical marker/indicator for drought tolerance (Ashraf and Foolad, 2007). In order to gain better understanding of drought tolerance in rice, proline content has been also assessed in different genotypes. It is known that free proline accumulation in plants under drought stress can enhance damage repair ability by increasing antioxidant activity and reducing ROS level (Aleksza et al., 2017).
The expression and functional analysis of stress-inducible proteins can be paramount to understanding the molecular mechanism of drought stress as some of the proteins may either be up-regulated or down-regulated under drought conditions (Seki et al., 2003). In the present study, we have carried out comparative proteome analysis through two-dimensional gel electrophoresis and proteins associated with drought stress have been detected. The variation in gene expression of the various drought related proteins among the different rice varieties under stress has been also investigated in the present study using Real Time PCR.
The conventional approach of plant breeding program is based on ‘yield-based selection’ but it has limitations such as inherent variation in the field and low heritability (Ribaut et al., 1997). As an alternative, significant efforts have been made for a breeding program using ‘trait-based selection’ (Chimungu et al., 2014). For instance, root attributes that affect water uptake under stress can help breeders to elucidate genotype and environment interaction for yield improvement (Serraj et al., 2011). Breeding strategies on the basis of secondary traits have been effectively employed for maize (Chapman and Edmeades, 1999), wheat (Richards and Passioura, 1989) and sorghum (Tuinstra et al., 1998). Keeping this in sight we have hypothesized that there is an evolutionary advancement in the rice growth mechanism with the development of specific strategies to overcome drought condition. We have investigated the relative performance of various attributes such as root morphology, root xylem, stomata, RWC, proline content, proteins and gene under drought stress and non-stress environment that contribute towards drought tolerance in rice.
Section snippets
Plant growth condition and stress imposition
Sample collection was carried out at Indian Agricultural Research Institute (IARI), Delhi. Three cultivars including PB6 (drought-tolerant indica rice variety), Moroberakan (drought-tolerant japonica rice variety) and Way Rarem (drought-sensitive indica rice variety) were used for detailed characterization under drought stress condition. Pot experiments were carried out in greenhouse conditions as described in Table 1.
Stress was measured in the terms of SMP (Soil Matric Potential). Therefore,
Root morphology
In the present study, root length, area, average diameter and volume of roots of drought-tolerant and drought-sensitive varieties has been calculated under control and drought conditions (Fig. 4A–D). We observed that the ratio of root length (stress/control) increased significantly (P < 0.05) in drought-tolerant varieties (PB6) while in drought-sensitive variety (Way Rarem) there was a significant decrease (P < 0.05). Similar trend was observed for root area, average diameter and volume,
Discussion
Improvement of drought tolerance in rice is one of the most challenging tasks due to high complexity of the traits and poor understanding of plant response against drought (Sinclair, 2011). Yield stability under severe drought stress can be achieved by developing a wider knowledge of various morphological, physiological, biochemical and molecular attributes of rice during reproductive stage. Further, these phenes together have the potential to drastically improve the breeding of crop ideotype
Conclusion
Improving drought resistance in crop plants to prevent yield loss remains one of the major challenges in the field of agriculture. Rice plant has been observed to develop suitable strategies in defense against drought stress. The ultimate aim of our work is to elucidate the drought-related traits using “omics”-based technologies such as phenomics, genomics, proteomics and transcriptomics that participate in enhancement of drought-tolerance traits. Furthermore, the precise knowledge of the
Contributions
AC and AA conceived and designed the study. AA performed the experiments. AC supervised, contributed towards data analysis and writing of the manuscript. AA and SB collected the root samples. BL and SC supervised the image processing experiment. All authors have read and approved the final manuscript.
Acknowledgement
This work is funded by National Agricultural Science Fund (NASF) under Indian Council of Agricultural Research (ICAR), Delhi, India [Phenomics of moisture deficit and low-temperature stress tolerance in rice, Funding number: NFBSFARA/Phen-2015/2010- 11]. Anupama is thankful to Council of Scientific and Industrial Research (CSIR) for providing JRF and SRF. We are thankful to Prof. Viswanathan Chinnusamy, IARI for providing rice plant root and shoot samples, Dr. P.K.Mandal, IARI for providing
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