Water deficit as a regulatory switch for legume root responses

Abstract

Plant roots perceive declining soil water potential as an initial signal which further triggers an array of physiological, morphological and molecular responses in the whole plant. Understanding the root responses with parallel insights on protein level changes has always been an area of interest for stress biologists. In a recent study, we reported drought stress-induced changes among certain structural and functional root proteins involved in reactive oxygen species (ROS) detoxification, primary and secondary metabolite biosynthetic pathways as well as proteins associated with cell signaling in an economically important legume crop Vigna radiata (L.) Wilczek. We also demonstrated photosynthetic gas exchange characteristics and root physiology under varying levels of water-deficit and recovery. In this report, we depict a closer analysis of the expression patterns of the identified proteins which were categorized into five major functional groups. These proteins represent a unique coherence and networking with each other as well as with the overall physiological and metabolic machinery in the plant cell.

Critical Role of Legume Root Responses Under Drought Stress

Apart from being the structural support, roots also serve an array of diverse functions which are crucial for the plant's growth and survival under adverse conditions.¹ Most of the abiotic stress factors including drought, salinity, temperature and heavy metals interact with the plant through soil. However, roots are the first organs to encounter these environmental stress factors. The perception of the initial stress signal and its subsequent transduction to the other plant organs is extremely important for triggering a specific stress response in providing either tolerance or susceptibility to the plant.² Among the various abiotic stresses, drought is the major concern for stress biologists owing to its complex response mechanisms which lead to deleterious effects on crop growth and productivity.^3–5

Legumes form a group of economically as well as agriculturally important crop plants, being the major source of proteins in our diet and also having a unique ability to fix atmospheric nitrogen through root nodules.⁶ The presence of these specialized structures (nodules) in the roots further complicates the signaling mechanism under declining soil water potential. Various in-depth analyses were carried out on both legume root and nodule responses under different abiotic stress conditions.^7–9 However, the acquisition of new insights in stress signaling pathways is still at its juvenile phase which requires further complements with advanced technology and innovative concepts.

We have recently shown, using a comparative root proteomics approach, that major and functionally essential root proteins of an important food legume Vigna radiata, were differentially expressed in response to medium and high water-deficit as well as during recovery phase. Progressive increase in water deficit manifested a simultaneous effect on the photosynthetic gas exchange characteristics, root architecture and physiology.¹⁰ In this report, we have categorized the identified proteins into five major groups: (1) ROS-detoxification, (2) Sulphur-metabolism, (3) Root-morphology, (4) Protein synthesis/energy metabolism and (5) Cell signaling Figure 1. The analysis of the expression patterns of these proteins highlight their possible role in the plant's overall drought-response mechanisms which could be used as a crucial piece of information for further interpretations of plant responses under progressive water-deficit.

Figure 1.

Categorization of differentially regulated protein spots, identified through MALDI-TOF-TOF, into five major groups based on their functional similarity. The up or downregulation of individual proteins in each category have also been depicted.

Inter-Relationship and Coherence among the Structural and Functional Proteins

Root proteome analysis at different stages of drought stress revealed up and downregulation of various root proteins, which at a glance appears to be random and un-related. However upon a closer analysis, we observed that there exists an inter-relationship and coherence among the identified groups of proteins. Major regulatory switch for these complex networks is the soil water status either in the form of medium or high stress as depicted in Figure 2. We analyzed the effects of medium and high water-deficit on each of the five groups of proteins for a better understanding of the regulatory mechanisms which can be summarized as:

1. ROS detoxifying proteins (Cu/Zn SOD and aldehyde reductase) were primarily upregulated in response to medium stress while high stress caused no significant changes when compared to the control plants. The significance of ROS detoxification in plants is primarily to provide protection to the essential biomolecules against oxidative damage.¹¹ Moreover, activity of these antioxidative enzymes plays an important role in maintenance of redox homeostasis inside the cell and also mediates a redox signaling for induction of specific stress responses.¹²

2. Key enzymes of sulphur metabolism (methionine synthase and cobalamine independent methionine synthase) were downregulated both by medium and high stresses when compared to the control plants. However, the intensity of downregulation was more during medium stress. It is known that root nodules play an important role in sulphur metabolism, especially in the biosynthesis of sulphur containing amino acids and regulatory compounds such as S-adenosyl L-Methionine (SAM).⁹ This correlation is evident in our study too, as we observed a gradual decline in the root nodule number and dry weight/plant in response to progressive water-deficit.

3. Medium stress highly enhanced the expression of enzymes associated with root morphology (Xyloglucan endotransglucosylase; XET), while the other structural proteins (actin and tubulin) were down-regulated. However, during high stress, these structural proteins were highly upregulated. We believe that alterations in the expression of these proteins should have a positive correlation with the root architectural modifications which in-turn have an indirect effect on the overall plant photosynthetic process, due to the alterations in the net water conductance.¹³ Dynamic modifications in the actin networks were also known to be associated with cell-signaling mechanisms.¹⁴

4. Medium stress significantly down-regulated the key components involved in protein synthesis (translational initiation factor; tIF), folding (heat shock protein 90-1, HSP 90) and carbohydrate metabolism (enolase) but enhanced the expression of a membrane transport protein involved in energy metabolism (V-type proton ATPase subunit E1; ATPase E). However, high stress significantly enhanced the expression of HSP and partially downregulated the other proteins. Photosynthetic efficiency in plants should directly or indirectly be dependent on such alterations in protein synthesis and energy metabolism.^15–17

5. We demonstrate that proteins associated with cell signaling (lectins and oxidoreductases) as well as plant's normal metabolism were significantly upregulated during both medium and high stress. However, another key enzyme in flavonoid biosynthesis and also known to influence the process of nodulation (Chalcone isomerase; CHI) was significantly downregulated by both medium and high stresses. The expression patterns of this enzyme were positively correlated with the concomitant changes in the root nodule number and dry weight. All the proteins mentioned above are known to be involved in cellular signaling which play the most crucial role in any stress response mechanisms.^18–21

Figure 2.

Schematic representation of the inter-relationship and networking between the identified groups of proteins and overall plant metabolism in response to medium or high water-deficit. Solid arrows indicate upregulation and broken arrows indicate downregulation. Solid lines signify inter-relationship between two different functional activities in the plant cell.

Considering the above interpretations and correlations, we have shown a schematic network of the possible inter-relationships between the identified proteins and subsequent physiological effects in plant cell during medium and high water-deficit regimes (Fig. 2). Further studies on individual networking pathways can be carried out to complement the present understanding of drought-induced cell signaling and whole plant responses.

Addendum to: Sengupta D, Kannan M, Reddy AR. A root proteomics-based insight reveals dynamic regulation of root proteins under progressive drought stress and recovery in Vigna radiata (L.) Wilczek. Planta. doi: 10.1007/s004.25-011-1365-4. In press.