Review paper

DROUGHT TOLERANCE IN RICE AND ROLE OF WRKY GENES

S. Kanwal*¹, S. Jamil¹, N. Afza², I. Ghafoor³, M. Shehzadi⁴, R. Kanwal² and M. Z. Iqbal¹

¹Agricultural Biotechnology Research Institute, ³Wheat Research Institute, AARI, Faisalabad

²University of Agriculture, Faisalabad; ⁴Agronomic Research Station, Farooqabad

^*Corresponding author’s email: kanwalshamsa32@yahoo.com

ABSTRACT

Anthropogenic alteration in climate has resulted in devastating global issue of drought for rice crop. Drought interferes with all the growth stages of crop by delaying its growth mechanisms, metabolic pathways and reducing the spikelet fertility. Rice plant has developed some morphological and physiological mechanisms to cope with the stress which include reduction in photosynthesis and transpiration, increased stomatal conductivity and density, root to shoot ratio, root length and carbon assimilation. Some biochemical modifications like biosynthesis of various hormones (ABA) and proteins (proline) also help in reducing the yield losses. Marker assisted selection and Quantitative Trait Loci (QTL) mapping are the advanced molecular techniques that played a vital role in developing the improved and stress tolerant rice cultivars. Identification of different stress responsive genes and transcriptional factors especially WRKY TFs have provided platform to obtain good crop stand and yield even under stress conditions. Rice possesses more WRKY genes (109 WRKY TFs in rice) as compared to Arabidopsis thaliana. An enormous variation in the expression patterns of WRKY genes and their contribution to the amplification of various signaling pathways and regulatory networks has been observed. These transcription factors work by regulating different mechanisms of drought tolerance and by releasing hormones, proteins, reducing-sugars, solutes and by affecting the stomatal conductance and root architecture. The WRKY genes work by showing up- and down- regulation to proteins, various protein-protein interactions and cross-regulation of WRKY TFs. Advance breeding methods; MAS, GWS, MABC, MARC and biotechnological tools along with different WRKY transcription factors have dynamically contributed in developing abiotic and biotic stress resistant rice varieties/cultivars on large scale.

Key words: Drought stress; WRKY TFs; QTLs; Biotechnological approaches; Rice breeding

http://doi.org/10.36899/JAPS.2022.3.0462
Published first online October 19. 2021

INTRODUCTION

Rice (Oryza sativa sp.) is the major staple food of about 3.2 billion population around the world and carries the smallest genome among all the cereal crops (Moin et al., 2017; Dien et al., 2019). Certain yield limiting factors in rice crop cause severe yield loss, for instance more than 50% reduction in yield occurs through abiotic stresses, majorly due to; drought, heat, cold, salinity and oxidative stress (Ijaz et al., 2021; Kanwal et al., 2021). About 23 million hectares area of cultivated rice under rain-fed regions suffers from drought (Serraj et al., 2011). Susceptibility to drught stress is mainly due to small root system, waxy cuticle and fast stomata closures (Ji et al., 2012; Pirasteh-Anosheh et al., 2016). Various growth stages of rice respond differently at different levels of drought (Sharifunnessa and Islam 2017; Kumara et al., 2020). Rice plant faces either terminal or intermittent drought stress. Terminal drought leads to water scarcity, causing severe stress to plant and ultimately kills the plant. Whereas, the intermittent drought stress occurs due to inadequate irrigations or rainfalls occurring either once or at intervals during the planting season causing distress in plant growth. Intermittent stress is less lethal as compared to the terminal stress (Polania et al., 2017). Figure 1 elaborates how of rice plans respond to drought stress at morpholological, biochemical and physiological levels.

The provision of information about genome sequences, molecular markers and the inexpensive genotyping has played important role in the practical application of marker-assisted breeding technology on a routine basis (Swamy and Kumar 2013; Chung et al., 2017). In the recent past, extensive study has been done in proteomics, transcriptomics and metabolomics approaches of functional genomics which revealed that the rice plant has evolved a wide range of strategies to cope with drought stress (Shahzad et al., 2021b). Moreover, the drought stress responses can be improved by employing different techniques such as marker assisted selection along with conventional breeding methods and by application of different hormones, osmolytes & enzymes and generating transgenic rice varieties (Upadhayaya and Panda 2019).

Transcription factors are classes of genes which play important role in managing the responses to different abiotic and biotic stresses. Among all, the important TFs are NAM, ATAF₁/₂, dehydration responsive element binding protein (DREB), myeloblastosis (MYB), ethylene responsive element binding factor (AP2/ERF) and WRKY (He et al., 2016). The WRKY TFs family has expanded greatly in evolutionary process starting from algae to Arabidopsis and flowering plants through whole genome, segmental and tandem duplications. The WRKY transcription factors domain enhances the plant growth and development and majorly involved in biotic and abiotic stress tolerance processes (Chen et al., 2019). WRKY family has widely been identified in various plant species including large number of WRKY genes in Arabidopsis, wheat, rice, sorghum, soybean, barley, pinus, papaya, poplar (Chen et al., 2012), Brachypodium distachyon, Citrus sinenses, Citrus clementine, Jatropha curcas, Manihot esculenta, Malus domestica, Oryza sativa. ssp. Japonica, in Oryza sativa ssp. Indica, Zea mays, Solanum lycopersicum and Solanum tuberosum (Liu et al., 2017). The current review focuses on the effects of drought stress on rice crop, and the mechanisms which rice plant has developed for combating stress. Moreover, the major focus is on advance breeding techniques and the working of novel WRKY gene family for drought tolerance in rice.

Drought stress in Rice: During the vegetative phase, the drought stress causes delay in leaf and tiller establishment, reduces the number of panicles per plant and result in low grain production (Singh et al., 2017; Swain et al., 2017). A water tension of -20 kPa in soil at vegetative growth phase results in 50% grain yield reduction in comparison with normal water supply (Swain et al., 2017). Drought suppresses the tillering and expansion of leaf and causes reduction in the leaf area and rate of photosynthesis (Bunnag and Pongthai 2013; Zhao et al., 2020).

The reproductive stage of rice is more affected by drought than the vegetative stage (Shrestha 2019). The most affected stage under drought stress is grain filling (anthesis to maturity) stage which is followed by panicle initiation stage (Sharifunnessa and Islam 2017; Melandri et al., 2020). At flowering stage, drought stress harms the physiological parameters as well as the rice yield (Yang et al., 2019b). It affects spikelet fertility, viable pollen production, panicle exertion, pollen-shedding, germination and embryo development (Shrestha 2019) ultimately reducing final grain yield (Liu et al., 2006; Rang et al., 2011; Zhao et al., 2020).

Effect of drought stress on morphology of rice: Drought stress impairs the germination of the seedling (Padmini et al., 2014), decreases the number of tillers per plant (Ashfaq et al., 2012; Bunnag and Pongthai 2013), plant height (Bunnag and Pongthai 2013; Sokoto 2014), fresh and dry root and shoot weight (Centritto et al., 2009; Ji et al., 2012). Root is the first organ exposed to water stress due to an insufficient or excessive level of water in the soil. The root surface area depends on the root hair development and root diameter and these traits can be improved under water stress in order to increase the improved uptake of soil moisture and to maintain productivity of plant (Koevoets et al., 2016; Kim et al, 2020). Coarse deep roots having increased ability of penetration and branching and high root to shoot ratio are found to be significantly contributing to drought avoidance in rice plants (Gowda et al., 2011; Pandey and Shukla, 2015). Rice plant has three adaptive mechanisms for root system under drought stress (i) osmotic adjustment in roots, during relatively small soil water reservoir, (ii) increase in soil root penetration, and (iii) increase in root density, root depth, and the root-to-shoot ratio during large soil water reservoir (Kim et al., 2012; Kim et al., 2020). 

Number of leaves (Sokoto 2014), leaf area and index (Kumar et al., 2014; Cal et al., 2019), leaf angle and elasticity in leaf rolling are also affected by the prevalence of drought stress (Pandey and Shukla 2015). Water deficit conditions negatively affects the mineral uptake and metabolism thereby decreasing leaf area and change partitioning of assimilates among different pant organs. Under severe water stress, increased degree of leaf rolling observed in rice leaves (Cal et al., 2019). It is the acclimation response of rice plant, and is used as a principle tool for assessment of drought tolerance (Pandey and Shukla 2015). It results in the reduction of CO₂ which diffuses in leaf and decreases the transpiration rate minimizing leaf dehydration, (Kadioglu and Terzi 2007) and maintains the normal plant water status (Gana 2011). Leaf rolling delays when the cell turgor is maintained, however, increase in leaf rolling under severe drought stress helps in preventing water loss and damage from radiations (Pandey and Shukla 2015). It varies in different rice genotypes showing that it has genetic basis. A large number of QTLs linked to this mechanism are identified in rice (Salunkhe et al., 2011; Grondin et al., 2018).

Effect of drought stress on physiological characters of rice: Physiological characters affected by drought in rice including rate of photosynthesis, conductivity of stomata and water use efficiency (WUE). Moreover, the relativity of water, chlorophyll content, photosystem ll activity, stability of membranes, carbon isotopes distinction and abscisic acid content are also affected by drought (Pandey and Shukla 2015). Drought stress causes the closing of stomata which leads to decreasing chlorophyll content and transpiration rate (Li et al., 2015; Caine et al, 2019). Loss of chlorophyll is a negative consequence of abiotic stress and is an important adaptive character in proper growth of plants under drought.

Effect of drought stress on biochemical parameters of rice: Numerous biochemical changes occur in response to drought stress in rice including the accumulation of osmo-protectants like proline, sugar, polyamines and anti-oxidants (Joseph et al., 2015, Pandey and Shukla 2015). Soluble sugars play an important role as an osmo-protectant during water stress (Artadana et al., 2019). The high level of proline in plants enhances the drought tolerance, stimulates the oxidative stress tolerance by antioxidant enzymatic activities (Sharma and Dubey 2005; Dien et al., 2019) and regulates the plant growth during drought (Zandalinas et al., 2018). It also helps plant cell to stabilize the structure of cell membranes and proteins. Accumulated proline plays a significant role to reduce and regulate water loss under water deficit conditions by acting as compatible solute and helps in adjusting osmotic balance. Proline accumulates contribute to stress tolerance in plants by supplying energy for growth and survivor. It is a significant indicator to screen drought tolerant lines in water deficit conditions (Pandey and Shukla, 2015). Rice varieties manifesting enhanced tolerance to abiotic stress were found to have elevated rate of proline biosynthesis. (Dien et al., 2019). Drought stress causes increased accumulation of various reactive oxygen species (ROS), most of which are produced in chloroplast and in mitochondria also, resulting in oxidative stress (Impa et al., 2012). Superoxide anion radicals, singlet oxygen, hydrogen peroxide and hydroxyl radicals are the main reactive oxygen species. Plants exhibit a defense mechanism under drought stress for protection from the detrimental effect of oxidative stress. Plants having increased level of induced antioxidant exhibit improved resistance and tolerance to oxidative damage (Impa et al., 2012, Lum et al., 2014). The significant role of abscisic acid (ABA) in regulating drought stress and mediating plant stress response by activating stomatal movement and stress related genes is described in Fig.2 (Nadarajah and Kumar 2019). The release of ABA significantly increases during drought, salinity, chilling and freezing stress and enhances the plant tolerance to these stresses (Kanwal et al., 2019; Liu et al., 2019). ARAG₁ an ABA-responsive DREB gene, is hypersensitive to ABA signaling and the transcript level of ARAG₁increases under drought and ABA treatment and increases tolerance to drought.

This gene is overexpressed in roots, inflorescences, immature embryos and germinating seeds (Zhao et al., 2010). Rice OsbZIP42 is a member of the group E bZIP and is positive regulator of ABA signaling conferring drought tolerance (Joo et al., 2019). PYL10 is one of the ABA receptors (ABARs) and the over-expressed PYL10 transgenic rice shows high RWC, membrane stability index, chlorophyll content, and lower amount of H₂O₂ manifesting both drought and cold tolerance (Verma et al., 2019).

Breeding for Drought Tolerance: In modern breeding, general terms such as marker-assisted selection (MAS), genome-wide selection (GWS), marker-assisted pedigree selection (MAPS), marker-assisted recurrent selection (MARS) and marker-assisted back crossing (MABC) are used. Recurrent selection is a type of varietal improvement method that involves a number of crosses for gathering multiple genes, controlling favorable characters and maintain varietal diversity. This method has been used in rice, maize (Bolaños and Edmeades 1993; Pang et al., 2017) and soybean (Posadas et al., 2014). A male sterile line “Jiabuyu” has been developed which has a dominant male sterility gene (Pang et al., 2017) and is mapped on the chromosome number 8 (Yang et al., 2012).This method is superior to pedigree selection as it gives both improved agronomic traits and enhanced drought tolerance (Pang et al., 2017).

Marker-assisted back crossing method is most efficient and widely used for the phenotypic characters that are under control of single gene (Miah et al., 2017; Chukwu et al., 2019). Backcrossing is mostly used in rice breeding for the introgression of targeted genes controlling the particular desirable trait from donor parent to recipient parent along with the high recovery of the recipient parent genome and eliminating the donor parent genome. Drought tolerant rice varieties have been developed by backcrossing the donor lines Basmati, Bg 300, Binam, BR24, Cheng-Hui 448, FR13A, OM1706, OM1723, Tarom molaii and Type3 (having high drought tolerance) with the high yielding (IR64 and Teqing) rice cultivars (Lafitte et al., 2006).

Induced mutation breeding is another approach which plays an important role in creating new genetic sequences of alleles that are not present in nature and these mutants can be introduced as a new variety or may be added in breeding programs (Oladosu et al., 2016). Mutagenic treatments with strong genetic effects and less physiological changes are desirable. Mutation breeding has evolved many mutant rice varieties including Nagina-22 (Poli et al., 2013), NERICA mutant (Sharifunnessa and Islam 2017), DSS1 rice mutant, 2 OsCDPK7 rice mutant (Ansari et al., 2015) and Manawthukha rice mutant that are tolerant to drought stress. In Malaysia, MR219-9 and MR219-4 are drought tolerant high yielding rice varieties derived from MR219 through mutation breeding (Rahim et al., 2012)

QTLs and marker assisted selection: Drought related traits are controlled by several quantitative trait loci (QTLs) (Fleury et al., 2010; Liu et al., 2012) and tolerance against drought cannot be achieved unless the molecular mechanisms involved in grain production are focused while using the techniques of traditional breeding or modern genetics. Thousands of the QTLs have been identified and cloned by using genetic engineering tools (Francia et al., 2005). For the production of new transgenic lines, candidate genes are major targets and molecular approaches use these QTLs for crop improvement (Varshney et al., 2011). Linkage-based QTL mapping involves those mapping populations (which have the traits correlated to drought tolerance) that segregate and identify polymorphic markers, construct the genetic map, and show accurate phenotyping and QTL mapping according to the phenotypic and genotypic data (Fleury et al., 2010).

QTL mapping is not that efficient in providing complete data about the QTLs as many of the inheritance related limitations are associated with mapping-populations. This is because the QTLs are mostly linked with large chromosomal segments having insufficient time for recombination, also the QTLs segregation is linked with the same traits and have inadequate phenotypic variation in diverse mapping-populations (Liu et al., 2012). Many of the QTLs that are related to drought tolerance, show responses to stress at root and shoot level, osmotic adjustment, photosynthesis, stomatal adjustment and hormonal responses. A major QTL qDTY1.1, has additive effect on the grain quality at reproductive stage under drought and is present on chromosome 1, flanked by RM431 and RM11943 in the populations of N22/Swarna, N22/MTU1010 and N22/IR64 (Vikram et al., 2011).

Marker assisted selection is not efficient as it functions only with the major QTLs, the minor QTLs are under-represented in the selection process resulting in missed genetic gains. Genomic selection (GS) is highly recommended and is more advantageous as it also deals with the minor genes than the traditional MAS (Hayes et al., 2009). The GS technique shows a combined effect of all the alleles on polygenic traits and is a type of MAS revealing the genetic variation in almost every individual and reduces the time of selection for most traits (Lorenzana and Bernardo 2009). Genotyping-based sequencing (GBS) is a novel approach and tool of NGS having its applications in discovering and genotyping of SNPs in crop plants. This GBS approach works by using restriction enzymes to digest the genomic DNA, ligation of barcode adapter, PCR and then sequence the amplified DNA on a lane of flow cells. (He et al., 2014; Chung et al., 2017).

Role of biotechnological approaches in mitigating drought stress: Genetic engineering has played a key role in developing drought tolerant transgenic rice varieties. Different genes encoding proteins such as kinases, transcription factors, enzymes and some regulatory proteins are involved in developing transgenics with enhanced drought tolerance (Todaka et al., 2015). The transcription factor OsbZIP23 is the leading regulator of ABA and its over-expression in rice is highly sensitive to ABA at germination and post-germination stages (Xiang et al., 2008; Yang et al., 2019a). Another TF OsbZIP46 in its active form OsbZIP46CAI shows drought tolerance in transgenic rice. AtCYP78A7 protein encodes cytochrome P450 in transgenic brown rice and improves the tolerance to water deficit by keeping the quality and composition of the key nutrients same as present in non-transgenic rice (Nam et al., 2013).

Response of rice plant to drought stress is found to be a multigenic trait at molecular level. Candidate genes showing drought tolerance have been identified in different plants by using genome annotation, functional genomics and molecular physiology (Feuillet et al., 2011). Different proteins related to the drought-responsive candidate genes directly take part in (1) cell protection like osmotic adjustment, repairing, degradation, structural changes, detoxification and (2) the positive correlations with the protein transcription factors DREB, MYB, bZIP and protein kinases to fight with drought (Hadiarto and Tran 2011). Some of the drought tolerant candidate genes, CDPK7 gene accession number LOC-Os03g03660 (Saijo et al., 2000), CIPK03 gene accession number LOC-Os07g48760 and CIPK12 gene accession number LOC-Os01g55450 have been identified in Oryza sativa for drought tolerance; showing signal transduction, have protein kinases and other regulatory proteins (Xiang et al., 2007). There are two main groups of the products of stress-inducible genes. The first group involves proteins which protect the plants directly under stress conditions such as protein conferring late embryogenesis, chaperones, antifreeze proteins and enzymes involved in biosynthesis of osmoprotectants. The second group includes those proteins which are involved in signal transduction and regulation of gene expression which are protein kinases and transcription factors.

Thousands of regulatory elements and proteins have been identified in plants and many of them incorporated in the rice genome in order to study their response to drought stress either by showing suppression or over-expression (Huang et al., 2009; Usman et al., 2017). In rice, WRKY genes encoded transcriptional factors control different biological processes, particularly zinc-finger proteins are widely distributed in rice for regulating stress responses. A zinc-finger “DST mutant” shows enhanced drought and salinity tolerance by decreasing the stomatal density (Huang et al., 2009) and another zinc-finger “OsZFP252” shows 74-79% high survival rate of rice plant in drought stress (Xu et al., 2008; Yang et al., 2010). Abscisic acid accumulates in guard cells and helps in closing of stomata which reduces water loss during drought (Lim et al., 2015). A major enzyme LOS5/ABA3 that appears in the end stage of ABA biosynthesis was over-expressed in transgenic rice and the grain yield and grain filling traits were improved during drought stress (Hu and Xiong 2014).

The late embryogenesis abundant (LEA) proteins ranging from 10-30 kDa are found in many plants (Olvera-Carrillo et al., 2011). Over expression of these proteins like OsLEA3 in rice enhance drought tolerance (Xiao et al., 2007) and HVA1 (an encoded LEA protein gene) from barley, wheat and rice shows increase in WUE and growth performance under drought (Babu et al., 2004; Chen et al., 2015). The encoding LEA genes OsLEA3-1 & OsLEA3-2 also show tolerance to water deficit conditions and minimum yield losses as compared to controlled treatments in severe droughts (Xiao et al., 2007; Duan and Cai 2012).

Overview of WRKY Transcription Factors: WRKY transcriptional factors are responsible for genes regulation that show response at different growth stages of a plant. The gene expression of WRKY TFs occurs mainly by binding to the DNA at specific cis-regulatory W-box elements, which are short sequences (T)TGAC(C/T) located at the promoter site of gene sequence in DNA (Finatto et al., 2018; Jimmy and Babu 2019). They have an invariant amino acid sequence at the N-terminus which is about 60 residues called the WRKY signatures, while the C-terminus of WRKY TFs contains a Zinc-finger structure of Cx4-5Cx22-23HxH or Cx7Cx23HxC (Shahzad et al., 2021a; Jimmy and Babu 2019). WRKYTFs show expression during various diseases, environmental stresses, seed germination, seed dormancy and other growth related events in higher plants (Rushton et al., 2010; Li etal., 2020). The WRKY proteins also function in immune response mechanisms of a plant such as the mechanical damages and wounding (Hara et al., 2000) and stresses like highly saline soils (Niu et al., 2012; Wang et al., 2012), drought stress (Zhang et al., 2008b), heat shock stress (Li et al., 2010), damages by ultra-violet radiations (Izaguirre et al., 2003), sugar signaling by sugar-responsive elements (Sun et al., 2003), gibberellin signaling in aleurone cells (Zhang et al., 2004) and ABA stimulation (Chen et al., 2010).

• The WRKY domains contain a zinc-finger at the N-terminus and is supported by the evidence that 2-phenanthroline chelates the zinc ions when added to the gel retardation assays, resulting in the non-binding with W-box target sequence in E.coli. An in-variant motif of 60 amino acids sequence has a well-conserved WRKY signature WRKYGQK on N-terminus (Rushton et al., 1995; Xu et al., 2020).
• The amino acid sequence on N-terminus binds with the W-box on the targeted DNA sequences. They are highly specific to their promoters in the target DNA. Along with the W-box core, the nearby sequences also play an important role in binding the W-box with the WRKY TFs (Eulgem et al., 2000; Yamasaki et al., 2005).
• Yamasaki determined the structure of C-terminal WRKY domain forming a complex with W-box in the DNA of Arabidopsis (WRKY4). He found that a four-stranded β-sheet perpendicular to the helical axis enters the major groove of DNA in the β-wedge (Yamasaki et al., 2005; Duan et al., 2007; Yamasaki et al., 2012).

The WRKY Gene Family in Rice: Rice (Oryza sativa.sp) contains higher number of WRKY TFs than in Arabidopsisthaliana (Goff et al., 2002; Yuan et al., 2005). Many WRKY TFs have identified in Oryza sativa L. ssb indica and L. ssb japonica. The plant transcription factor databases have different number of WRKYTFs in indica and japonica by having 111 and 129 WRKY TFs respectively whereas Indica has 107 and japonica has 126 coding regions out of these gene models. Only 101 WRKY TFs in indica and japonica were classified in groups and the rest are putative gene sequences. These genes mainly responsive to the abiotic and biotic stresses (Jimmy and Babu 2019). Indica rice contains 102 copies of WRKY genes out of which 92 are unique. Two identical copies of OsWRKY46 are present on chromosome 11 and 12. OsWRKY55 and -89 are doubled in indica rice showing the tandem repeats on chromosome 3 and 5 respectively (Ross et al., 2007).

Japonica has two identical copies of OsWRKY46 at chromosome 11 and 12 but unlike indica, OsWRKY55 and OsWRKY89 has only one copy in Japonica. WRKY genes are unevenly distributed in the rice and chromosome 1 has maximum number of WRKY TFs (Xie et al., 2005). The densities of WRKY TFs on all the 12 chromosomes appear to be same in both japonica and indica. Equal number of WRKY TFs are present on the chromosome (1, 2, 5, 6, 10 and 11) while other chromosomes have difference of one or two WRKY TFs. Genes on the chromosomes 1, 3, 5, 8, 11, and 12 are present in clustered form (Ross et al., 2007). From the evolutionary analysis of rice WRKY TFs, the group 1 WRKY genes are classified into G1a and G1b.

Two differences have found in group 1a and group1b, out of which first difference is, the zinc-finger motif on N-terminus and C-terminus of group 1a is of C₂H₂ type, while the zinc-finger motif on N-terminus and C-terminus of group 1b is of C₂HC, while the second difference is that the group 1a genes has a conserved intron only on the C-terminal of the WRKY domain encoded region. However, the group 1b contains conserved intron on both C-terminal and N-terminal. The group 1 WRKY TFs of slime mold and Arabidopsis has highest similarity with the rice group 1a and not the group 1b. So, the only group 1a genes of rice are part of ancestral history of WRKY genes in plants (Jimmy and Babu 2019). The evolutionary study of rice suggests the group1 of WRKY TFs as the oldest one because it contains 2 WRKY domains and that the group 2 and 3 have originated from group1 (Ulker and Somssich 2004). A fourth group has also emerged and classified as a set of WRKY genes that has a complete WRKY motif but the Zinc-finger is absent (Ross et al., 2007). Table 1 describes the various groups of WRKY Transcriptional factors traits being controlled.

Table 1 Classification and Role of Different WRKY Transcriptional Factors

Role of WRKY TFs for Drought Tolerance in Rice: 103 WRKY TFs of WRKY family have identified in rice. Four genes out of these 103, are up-regulated by drought while 13 by both salt and drought stress. Two WRKY TFs are up-regulated by drought but down-regulated by cold stress. One gene is regulated by drought but suppressed by cold and salinity (Ramamoorthy et al., 2008).

OsWRKY72 shows tolerance against drought in rice and is induced by NaCl, PEG, ABA and high temperature. It shows high sensitivity to mannitol, NaCl and ABA (Song et al., 2010a). This TF is an up-regulatory gene and is stimulated by abscisic acid released in aleuron cells (Li et al., 2015). OsWRKY72 imparts tolerance to salinity in rice as it shows higher expression during salts stress in different rice varieties like Rasi & Tellahamsa (Ashwini et al., 2016). Another gene OsWRKY11 is induced by drought in rice seedlings. The over-expression of OsWRKY11 along with HSP101 promoter which shows the drought and heat tolerance and results in slower leaf-wilting (Wu et al., 2009). Research shows that OsWRKY11 binds to RAB21 promoter, enhances the heat and drought tolerance in transgenic rice (Shi et al., 2018). A WRKYTF OsWRKY30 has identified in rice which is activated by MAP Kinases (Shen et al., 2012). OsWRKY30 has two WRKY domains and belongs to group la. It is induced by M. grisea infection and phytohormones SA (salicylic acid) and JA (jasmonic acid) is discussed in Table. 1. The SA antagonizes JA by showing response against disease resistance (Ryu et al., 2006). The over-expression of OsWRKY30 shows a dramatical increase in drought tolerance in rice. (Shen et al., 2012).

A rice zinc-finger protein “DST mutant” shows drought tolerance by decreasing the distribution of stomata and enhancing the closing of stomata in rice. Similarly, over-expression of the zinc-finger protein OsZFP252 increases the drought tolerance by accumulation of soluble sugars and proline and increase the plant survival upto 74-79% (Oladosu et al., 2019). The SbWRKY30 works as a transcriptional factor (localizes in the nucleus) and is positively regulated by drought stress. The heterologous expression of SbWRKY30 shows enhanced drought tolerance in rice by affecting the root architecture. Moreover, SbWRKY30 transgenic rice has higher proline content, SOD (superoxide dismutase), POD (peroxidase) and CAT (catalase) activities. In rice SbWRKY30 shows tolerance to environmental stresses by up-regulating some stress-related genes (Yang et al., 2020).

The heterologous expression of AtWRKY57 induces drought tolerance in transgenic rice. This over-expressing of AtWRKY57 also contains salt tolerance. Firstly it showed drought tolerance in Arabidopsis thaliana and then introduced in rice to check whether it shows the same expression or not (Jiang et al., 2016). Similarly, the WRKY gene ZmWRKY58 when over-expressed, enhances the drought tolerance in rice by increasing the RWC (relative water content) and hence plant survival. The ZmWRKY58 has isolated from maize and its expression pattern indicates that it is induced by drought, salt and abscisic acid. Moreover, the ZmWRKY58 is also present in onion and yeast (ZmWRKY58 can interact with the Z2mCaM, suggesting its function as calmodulin binding proteins and promotes abiotic stress tolerance). In rice, ZmWRKY58 causes delay in germination and inhibits the post-germination (Cai et al., 2014). Ectopic expression of ZmWRKY33 and ZmWRKY58 in Arabidopsis thaliana and Oryza sativa improve the salinity and drought tolerance respectively (Shi et al., 2018).

OsWRKY13 is a transcriptional repressor of OsWRKY45-2 but suppresses the OsWRKY45-1 under drought stress and binds to the W-box sequence. It shows cross-talk between abiotic and biotic stresses in rice. This transcriptional factor is induced in the leaf vascular tissue, guard cells and bacterial infection, where another TF SNAC1 increases the tolerance to the drought stress (by inducing stomatal closure) and some bacterial infection. The OsWRKY13 makes antagonistic cross-talk between drought and disease resistance pathway It suppress the SNAC1 and WRKY45-1 (Xiao et al., 2013). OsWRKY47 belongs to the gourp-ll of WRKY gene family and is the positive regulator of drought stress. Overexpression of OsWRKY47 in transgenic PSARK::IPT plants shows enhanced tolerance to drought. OsWRKY47 has sequences similar to the regions of CRSSP and CaMBP promoters, its gene expression profile in the flag leaves of PSARK::IPT plants shows the differential expression of OsWRKY47 and reduction in chlorophyll content hence2, enhancing drought tolerance (Raineri et al., 2015). Another WRKY TF 0sWRKY80 is induced by different factors including Rhizoctonia solani infection in rice, exogenous jasmonic acid (JA) and ethylene (ET) (Peng et al., 2016). Its overexpression increases disease resistance and in rice it is up-regulated in roots, leaves and stems showing drought tolerance (Ricachenevsky et al., 2010). Figure 3 gives an overview of how WRKY TFs enhance the tolerance to drought stress.

Conclusion: To conclude, it is evident from aforementioned illustrations that rice plant is highly susceptible to drought. Drought negatively influences the rice plants at different growth stages. In order to overcome the damages from water deficiency, and cope with the stress this crop has adapted morphological, biochemical and physiology parameters accordingly. The advanced techniques have improved the understanding of rice genome QTLs and WRKYTFs resulting in development of more precise drought tolerant rice types focusing only on attributes and mechanisms which directly interfere with their development and production under drought stress conditions.

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