12

Improving the Plant Root System Architecture to Combat

Abiotic Stresses Incurred as a Result of Global Climate Changes

Ananda K. Sarkar, Karthikeyan Mayandi, Vibhav Gautam, Suvakanta Barik,

and Shabari Sarkar Das

Abstract

Abiotic stresses incurred as a result of global climate changes are posing a threat toplant growth and productivity, and thereby leaving mankind in an upcoming foodcrisis. Although most plants are affected, domesticated crops are more vulnerableto damage by adverse environmental conditions or stresses. The root of a sessileland plant is a crucial organ for anchorage and uptake of nutrients and water. Thearchitecture of the root system, which is influenced both by the genetic makeup ofa plant and environmental factors, greatly impacts plant growth and productivity.Being less accessible and a complex underground organ, roots have beencomparatively ignored by plant biologists. Although realized late, significantprogress has beenmade in understanding the biology of the root and its contributionto the adaptability of plants to environmental stresses. In this chapter, we discuss therecent understanding of molecular regulation of the root architecture in relation toabiotic stress responses. We also highlight the future perspectives of this study inproducing abiotic stress-resistant plants with efficient root systems.

12.1

Introduction

To feed the ever-growing human population in the near future, we need to increasefood production by several fold. However, increasingly depleted agriculturalresources such as ground water, arable land, soil health, and deterioratingenvironmental conditions caused by highly hostile climate changes have chal-lenged the efforts of improving crop productivity. Global climate changes are notonly changing worldwide ecosystems, but also imposing several abiotic stresses,such as extreme low/high temperature, drought, salinity, flood, and so on. Variousfactors, including soil type, temperature, relative humidity, organic matter in thesoil, local vegetation, and precipitation, determine the severity and impact of abioticstresses [1–3]. Drought, one of the most severe stresses, is often accompanied by

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Climate Change and Plant Abiotic Stress Tolerance, First Edition. Edited by Narendra Tuteja and Sarvajeet S. Gill.� 2014 Wiley-VCH Verlag GmbH & Co. KGaA. Published 2014 by Wiley-VCH Verlag GmbH & Co. KGaA.

low water availability and subsoil mineral toxicity [1]. These abiotic stresses severelylimit the growth of plants and productivity of major crops [1,4,5]. Out of 215 644plant species cataloged, only four crops (rice, wheat, maize, and potato) contributeto more than 60% of energy intake by the world’ s human population ([6]; http://www.fao.org/biodiversity/components/plants/en/). Although the “Green Revolu-tion” in the middle of the last century boosted crop production through the use ofexcessive fertilizers, pesticides, high-yielding varieties, and improved irrigationsystems, the benefit was not equally accessible to poor people [5–8]. Moreover, thishas depleted ground water levels, and increased environmental pollution andhealth problems [7,8]. Additionally, it has been realized lately that the chosenvarieties of the aforesaid crops have attained their maximum productivity and theyare less responsive to excess supply of fertilizers or water [5,8,9].It has been predicted that plants attain only 25% of their yield potential due to

negative impacts of environmental factors [9], which implies that with propermanagement of abiotic stresses a considerable amount of the 75% that goes towaste due to environmental factors can be added to the overall production.Drastically reducing ground water levels (because of agricultural/human activityand global warming) and abiotic stresses have only exacerbated the situation andposted a challenge to plant biologists to consider an alternative approach for cropimprovement. The strategies used in the first “Green Revolution” mainly focusedon aerial biomass and seed yield, while overlooking the importance of the rootsystem, which is a crucial organ of a plant required for the uptake of nutrients andwater, anchorage, and optimal growth and development of the plant body [7,8].Since plant root system responds very fast to the environmental changes,understanding the biology of root system architecture (RSA) is a prerequisite fordeveloping plants with efficient root systems that can better tolerate abiotic stressesimposed by global climate changes [7,8]. Therefore, improving the root systemthrough understanding the molecular regulation of the root architecture and itsadaptive responses to environmental stresses is “the key to the second greenrevolution” [5,7,8]. Being a sessile organism, plants have evolved adaptivemechanisms to cope with environmental stresses, although the productivitysuffers. A balance between water uptake capacity by roots, transpiration throughleaf stomata, and water-holding capacity of plant tissue plays an important role inproviding tolerance to major abiotic stresses such as drought and extremetemperatures [2]. Although previously neglected, plant scientists have now turnedtheir attention to the study of the root system [7,8,10]. Here, we discuss how theRSA is likely to play an important role in abiotic stress tolerance, especially in cropplants.

12.2

RSA and its Basic Determinants

In contrast to animals, during postembryonic development, higher plantscontinuously grow and produce organs through the activity of stem cell populations

306 12 Improving the Plant Root System Architecture to Combat Abiotic Stresses

residing in the shoot, root, and cambiummeristem [11]. This helps sessile plants toadapt to the changing environment by favorably altering their morphological andorgan developmental pattern [3]. The active root meristem is maintained at the roottip by the activity of the root stem cell niche comprising of a quiescent center andsurrounding layer of “stem cells.” The combinatorial activity of many transcriptionfactors (e.g., SCARECROW (SCR), SHORT-ROOT (SHR, PLETHORA1 (PLT1),PLT2, andWUSCHEL RELATEDHOMEOBOX5 (WOX5) and phytohormones (e.g.,auxin homeostasis) maintains the root “stem cell niche” [11,12]. The shape andstructure of the root system, which includes the main root and root branches,constitute the RSA [3]. The complexity of the root system varies among as well aswithin species [3]. According to developmental ontogeny, three major root types –

primary, nodal, and lateral roots – have been categorized [3]. The RSA varies betweenmonocot and dicot plant species. The main root and its branches constitute the dicotRSA, whereas in monocots, RSA consists of the primary root (that gets depleted at alater stage), nodal roots, and the branches derived from them [3,13,14]. In contrast todicots, monocot roots normally do not undergo secondary growth [3]. Plantsoptimize their RSA by producing root branches or lateral roots from thedifferentiated distal region of the root (primary/nodal) and by regulating the growthof different root types [3]. Lateral root development and RSA are controlled by theintrinsic genetic makeup of a species, and modulated by environmental factors suchas soil composition, availability of nutrients and water, and so on [3,15].Since the root is one of the first plant organs to encounter the impact of

environmental factors, its growth and development is highly influenced by them[15]. Studies on the root system of the model plant Arabidopsis thaliana haveenriched our knowledge about root system development, while our knowledge isstill at the elementary stage in the case of major crops like rice, wheat, and maize[1,3,8]. Now, it is necessary to put more emphasis on using diverse model crops(such as rice, wheat, maize, etc.) to uncover the molecular basis of their rootarchitecture and stress responses using the knowledge already gained inArabidopsis [1]. Additionally, because of the fundamental differences between theroot system of monocot and dicot plants, the molecular mechanism of regulation ofthe root architecture as well as its response to abiotic stresses might often vary [1].As an example, no obvious root phenotype has been reported for mutations in theAtWOX11 transcription factor of dicotyledonous Arabidopsis (that lacks nodalroots); however, mutation in OsWOX11 of monocotyledonous rice drasticallyimpaired nodal crown root formation [16,17]. Different root types (crown root,seminal root, lateral roots; as specified above) of monocot maize have been shownto be genetically regulated by different genes [13], suggesting the variation in andcomplex regulation of root architecture among different species.The tremendous genetic diversity trapped in the root system of different varieties

of a species provides huge potential to improve crop yield and stress tolerance byimproving the RSA through exploring its molecular and physiological basis [4,7].Several strategies have been taken by plant scientists to tailor the RSA for optimumplant growth, productivity, and stress tolerance. Both conventional and molecularbreeding approaches that exploit quantitative trait loci (QTLs) and marker-assisted

12.2 RSA and its Basic Determinants 307

selection (MAS) are being used to incorporate better RSA traits into high-yieldingvarieties and to improve stress tolerance. Using the information derived fromgenomics studies, transgenic plants with improved stress tolerance and RSA traits arebeing developed for selected candidate genes. Additionally, information derived fromstudies on polyamines, osmolytes, hormones, and small RNAs has been helpful.Increasing use of phenomics in the study of RSA traits and stress tolerance is likely tobridge the gap between genomics and phenotype. Here, we discuss the aforesaidapproaches for improvement of stress tolerance by producing better RSA traits.

12.3

Breeding Approaches to Improve RSA and Abiotic Stress Tolerance

12.3.1

Conventional Breeding Approach

Using conventional breeding approaches, RSA traits of crop plants can beimproved, which would lead to the better utilization of nutrient resources andwater sequestration, leading to better production and yield [18]. Some of theimportant criteria used for studying RSA traits are the length, thickness, anatomy,branch number, distribution in soil, and penetrating ability [19]. During thedomestication process, people selected plants mainly based on their aerial characterand yield, ignoring the root system. Recently, plant breeders have realized theimportance of RSA in crop productivity and turned their attention to breed varietiesconsidering their RSA traits [7,8]. Improved RSA traits in wheat, using conven-tional breeding, have provided tolerance to abiotic stresses [20]. Several stresstolerance and RSA-related traits of interest from selected wheat cultivars are beingintrogressed into high-yielding cultivars to develop cultivars with tolerance toabiotic stresses [7,8]. Through improvement of stem, leaf patterning, and rootarchitecture, an increase in yield of 68% has been achieved in foxtail millet [1]. Ithas been shown in barley (Hordeum vulgare) that the root system is significantlyimproved using a breeding approach [21]. Similar to other crops, huge geneticdiversity in RSA traits exists among different rice cultivars that also vary in terms ofyield and stress tolerance [22]. Drought-resistant rice cultivars with better RSAtraits have been developed using conventional breeding [23]. The RSA traits (suchas deeper and thicker roots with more xylem vessels, increased root length anddensity, etc.) have been shown to play very important roles in providing droughttolerance in rice [24,25]. IET1444 and GZ5121-5-2 rice cultivars with improved RSAtraits have been shown to produce high yields under stress condition [24].Although conventional breeding based on RSA traits has made some progress,

its efficiency is limited by the lack of huge breeding fields, trained breeders, and theextremely laborious process of studying less-accessible roots systems of a largenumber of plants. Moreover, it is time-consuming and the phenotypic observationsare often not reproducible due the variation of RSA traits caused by the complexinteraction between RSA and environmental factors as well as the soil condition

308 12 Improving the Plant Root System Architecture to Combat Abiotic Stresses

[21,25]. In an effort to overcome these limitations, the conventional breedingapproach has gradually been replaced by the modern QTL-based approach, whichincludes identification of genetic loci (QTLs) related to RSA traits and the use ofMAS-based breeding [25].

12.3.2

Identification of QTLs Associated with Specific RSA Traits and Stress Tolerance

A QTL is a region or portion of the genome responsible for a particular trait orphenotype. Several studies have identified QTLs associated with the root traits thatenable the plants to sustain abiotic stresses like drought, nutrient deficiency,extreme heat/cold, flooding, and so on [26]. QTLs have been identified for severaltraits like deep-root systems, root lodging, stele, and xylem structure in rice, maize,and wheat [27–29].It has been reported in rice that the genotypes classified as drought resistant

showed consistently higher cell membrane stability, more stable hydraulicconductivity, more responsiveness of root anatomy to drought, and higher levels oftemporal expression of aquaporin genes [30]. This range of traits allows efficientcontrol of the plant water status under drought [30]. Water uptake by roots isinfluenced by the size and number of xylem vessels; roots with thicker xylemvessels have a higher potential to uptake water than thinner ones [31,32]. Uga et al.have identified QTLs for rice root thickness, root stele transversal area and numberof late meta xylem vessels using a recombinant inbred line population derived fromthe cross between a lowland cultivar IR64 (with shallow RSA) and upland cultivarKinandang Patong (with deep RSA) [28]. Using the same population they furtheridentified a QTL Dro1 (deeper rooting 1) that is linked with deep RSA traits andinvolved in drought avoidance under natural conditions with limited water supply[27]. Recently, Dro1 has been shown to improve RSA and yield under droughtcondition, when introduced in a shallow-rooting rice [33]. Steele et al. have reporteda QTL for root length (between RN242 and RM201), which significantly increasedthe root length under stressed (and non-stressed) conditions and improved thepenetration ability of a rice cultivar [25,34]. By introgressing this QTL to Kalinga IIIlandrace, they developed a drought-tolerant rice cultivar that was released as “BirsaVikas Dhan 111” [25,35]. Using the double haploid population derived from a crossbetween IR64 and Azucena (upland rice cultivar), 15 QTLs for RSA and other traitshave been identified [36]. The chromosomal segment between RG171 and RG157markers contained QTLs that control tiller number and RSA traits of rice underdrought stress [36]. Genome-wide analysis of root development-related QTLs hasalso been initiated in rice [37].Sharp et al. have shown that maize roots growing under low water potential were

thinner and the radial growth rate was decreased throughout the elongation zone ofthe root, resulting in a greatly reduced rate of volume expansion [38]. Underdrought stress, plants try to avoid low water potential by balancing the water uptake(by root) and water loss (through transpiration) [39]. The avoidance of low waterpotential by developing deeper RSA has been related to an increase in grain yield

12.3 Breeding Approaches to Improve RSA and Abiotic Stress Tolerance 309

(in wheat and maize) and better adaption to drought conditions [39]. Ruta et al. haveidentified 13 QTLs for the elongation of axile and lateral roots of maize in responseto low water potential, and have identified the relationship between axile and lateralroots as a trait for improving the drought tolerance of maize [39]. Maize is highlysensitive to frost and moderately sensitive to chilling. Extreme cool temperatureaffects maize growth in a number of ways right from emergence through toflowering and seed setting. The severity of damage depends on temperature and itsduration, developmental stage, and genotype [40]. Early flowering with a long grain-filling duration and extended-stay green character provided the basis for hybridswith a high yield potential under low temperature stress [40]. QTLs controlling rootand shoot traits of maize seedlings under cold stress have been identified [40], andhave been shown for a large number of independently inherited loci suitable for theimprovement of early seedling growth through better seed vigor and/or a higherrate of photosynthesis [40]. An increase in the concentration of the phytohormoneabscisic acid (ABA) is an important adaptive response of plants to drought andtherefore “leaf ABA concentration” is a physiological trait considered by breeders[41]. In maize, a major QTL that affects leaf ABA concentration and RSA traits hasbeen identified as root-ABA1 (originally called L-ABA), which has been shown toaffect root lodging, grain yield, and other agronomic traits under water stress[29,41]. QTLs that affect lateral root development and length under low-phosphorous stress have been mapped in maize [42]. In pearl millet, the tolerantparent (PRLT 2/89-33) and QTL-NIL ICMR 01029 have been shown to have deepand profuse root systems, and are tolerant to drought [43]. Water stress-tolerantwheat cultivar was developed by crossing a susceptible cultivar with anothercultivar having the osmoregulation (OR) gene [44].Since the QTL is a large genomic region that may consist of multiple genes, it is

necessary to identify individual genes responsible for quantitative traits. Genes areidentified mainly by two approaches: positional cloning and candidate geneapproach. In positional cloning, QTLs are linked to a physical map, and furthermarkers are developed between the original markers used to tag QTL to narrowdown the target sequences and finally identify the target gene (may be bysequencing) [45]. Positional cloning requires a sufficient number of markers andgeneration of near-isogenic lines, which is time-consuming [34]. In the candidategene approach, sequence information (generated by reference genome sequencing)facilitates (with the help of bioinformatic tools) the listing of genes underlyingspecific loci. A reverse genetics approach can be followed to further pinpoint thegene function [45]. The biparental linkage-based mapping of QTLs to identify genesis mainly limited by the lack of sufficient genetic variation, insufficient recombina-tion frequency, and so on. These limitations can be overcome by population-based“association mapping,” also known as “linkage disequilibrium mapping,” whichhas recently been very popular for dissecting the molecular basis of complex traitsin plants. Association mapping is normally performed by correlating thephenotypic and genotypic data derived from larger natural populations with widegenetic diversity [46]. Marker tags are identified and placed to close proximity tothe target trait using efficient statistical methods [46]. It mainly depends on

310 12 Improving the Plant Root System Architecture to Combat Abiotic Stresses

molecular profiling and identifying the phenotypic details for the unrelatedaccessions [47]. Success of association mapping strongly depends on the level oflinkage disequilibrium present in the target population. A population that has ahigher value for linkage disequilibrium is considered as most suitable for QTLdiscovery [48]. Although association mapping is being used to indentify novelgenes and uncover their function-related traits of interest in different species suchas maize, wheat, oat, rice, and so on [46,47], it is yet to be exclusively used forimproving RSA traits in crops.

12.4

Genomic Approaches to Identify Regulators of RSA Associated

with Abiotic Stress Tolerance

Genomics plays an important role in genome-wide identification of genes that areresponsible for conferring variation in traits (such as RSA) and tolerance to abioticstresses between two or more species. Genome-wide analysis of the availablegenome sequence data has helped in locating root development-related QTLs inrice [37]. Suryapriya et al. have developed a Web tool “rootbrowse” that has beenused to locate 861 QTLs related to root development using information gatheredfrom several QTL studies on populations derived from crosses between variousupland and lowland rice cultivars (indica and japonica) [37]. Using this tool,valuable information such as simple sequence repeat markers, protein-codinggenes, and functional annotation of the genes are also displayed along with theQTLs, and thus it would be helpful in predicting the genes related to RSA traits[37]. Both genetic and genomic approaches have been used to understand andimprove drought tolerance-related RSA traits in pearl millet [49]. Recently, NGS-based whole genome resequencing of DNA (QTL-seq) has been used to identifyQTL in rice [50]. It is worth mentioning that the use of association mapping touncover complex traits has been boosted by advance genomics tools that help in therapid identification and scoring of genetic markers (using sequence informationwhen comparing plants) [46]. Thus, the use of association mapping to study RSAtraits and stress tolerance offers tremendous opportunities. However, combiningrecently developed high-throughput phenotyping technology with associationmapping studies would be of great advantage to score huge number of RSA traitsand correlate them with genetic markers or genes.Recent advances in high-throughput sequencing or “next-generation sequen-

cing” (NGS) are capable of producing huge amounts of sequence information in avery short period of time. This has accelerated genome-wide identification ofmarkers, differentially expressed transcriptomes, and small RNAs betweendifferent genotypes with contrasting traits. Microarray analysis is another popularapproach for genome-wide comparative transcriptome analysis. Arsenic toxicitystress has been shown to affect RSA traits by altering the expression of key signalingcomponents such as receptor-like cytoplasmic protein kinase, AP2 (APETALA2)/ERF (ethylene response factor), heat shock factor, MYB (myeloblastosis), zinc finger

12.4 Genomic Approaches to Identify Regulators of RSA Associated with Abiotic Stress Tolerance 311

proteins, and so on, as observed by root-specific transcriptional profiling [51]. Usinga microarray-based functional genomics approach, the rice JAmyb was found tobe differentially expressed in roots under salinity stress, and its functionalanalysis has revealed the role in seed germination, seedling growth, and rootelongation [52]. In wheat, expression of TdAtg8 was highly upregulated in rootunder drought stress. Through functional analysis, it has been shown to affectroot architecture and regulate drought and osmotic stress responses in bothArabidopsis and wheat [53]. Using SAGE (serial analysis of gene expression)-based analysis, root-specific transcriptome data of maize has been shown tohave multiple stress-responsive factors, which might regulate RSA traits [54].ZmMKK4, a novel group C mitogen-activated protein kinase kinase (MAPKK)of maize, was shown to be involved in providing salt and cold tolerance. Usinga functional genomics approach, this gene was shown to affect the develop-mental process of the plant, including the root architecture [55]. Microarrayand subtractive hybridization approaches have identified and validated theexpression of WRKY transcription factors, zinc finger proteins, and NACdomain protein in tomato root under salinity stress [56,57]. Nutrient stress(e.g., excess nitrogen)-responsive genes have been identified and validated intomato roots using a microarray-based functional genomics approach [58]. Ironand potassium deficiency and salinity stresses have been shown to alter theexpression pattern of the 14-3-3 gene family in tomato root, indicating a cross-talk between these stresses to regulate RSA and stress tolerance [59]. Salinitystress was shown to affect RSA traits and root proteome of the tomato plant byaltering the fate of the various genes necessary for regulating the physiologicalphenomenon of tomato plant [60]. It has been shown that salt stress leads tothe accumulation of ABA, which influences the physiological response of theplant and therefore affects its proteome [61].Using a deep sequencing approach, transcripts differentially expressed in

maize root under low and high water potential have been identified, and theirpotential role in regulating the RSA has been indicated [62]. In addition toprotein-coding genes, many microRNAs (miRNAs) have been shown to bedifferentially expressed under abiotic stresses [63]. Salinity stress-responsivemiRNAs have been identified from maize root using miRNA microarrayanalysis [64]. Using NGS of small RNA, it has been shown in maize thatmiRNAs such as miR159, miR164, miR167, miR393, miR408, and miR528were upregulated under salinity stress, and were involved in the adaptivestress response of root [65]. Functional analysis of these miRNAs and theirtarget genes would uncover their role in root development and stresstolerance. Thus, increasingly available whole-genome sequences of variouscrops and use of microarray or NGS-based comparative genomics, along withimproving bioinformatic approaches, are likely to provide information on thestructural and functional aspects of genes involved in regulating RSA andstress tolerance. With such information and tools at hand, it would be easierto do translational genomic research aiming at the development of high-yielding and stress-tolerant crops with efficient root systems.

312 12 Improving the Plant Root System Architecture to Combat Abiotic Stresses

12.5

Transgenic Approaches to Improve RSA for Abiotic Stress Tolerance

It is well documented in Arabidopsis that NAC genes are involved in lateral rootformation. In Arabidopsis, the NAC1 (NAM/ATAF/CUC1) functions downstream ofTIR1 (TRANSPORT INHIBITOR RESPONSE1) to mediate lateral root formationthrough auxin signaling [66]. AtNAC1 is not regulated by any environmentalfactors like salt stress, ABA, and ethylene [67]. However, AtNAC2, which functionsdownstream of the ethylene and auxin signaling pathway, is induced by salt stressand also promotes lateral root formation [67]. A similar kind of NAC gene, whichfunctions both in lateral root development as well as in salt tolerance, was reportedin soybean [68]. Overexpression of soybean GmNAC20 in Arabidopsis conferredenhanced tolerance to salt and freezing stresses, and it also increased the numberand density of lateral roots both in normal as well as in salt stressed conditions [68].The expression of GmNAC20 was induced by drought, salinity, cold, andnaphthalene acetic acid (a synthetic auxin). Expression levels of AIR3, ARF7, ARF19,AXR3, and LBD12 were upregulated in transgenic plants, but the expression levels ofARF2 and AXR1 were downregulated [68]. These results indicate that GmNAC20promotes lateral root formation through auxin signaling [68].Rice is predicted to have 140 NAC genes. Jeong et al. have identified a NAC gene,

OsNAC10, which is predominantly expressed in roots, and regulated by droughtand salinity [69]. They overexpressed the OsNAC10, both under constitutivepromoter (GOS2) and root-specific promoter (RCc3). Both the transgenic linesperformed better under drought than the non-transgenic line in laboratoryconditions [69]. However, under drought, in field conditions, only the root-specificoverexpression line performed better and resulted in a 25–42% increase in totalgrain weight in comparison to the non-transgenic line [69]. It was found that theroot diameters of the pRCc3:OsNAC10 lines were thicker by 1.25-fold than those ofthe pGOS2:OsNAC10 and non-transgenic lines [69]. Thus, this study shows directevidence for the link between root architecture and crop yield.Arabidopsis plants overexpressing HARDY, an AP2/ERF-like transcription factor,

were found to have increased secondary and tertiary roots, which resulted in20–50%more pulling force required to pull out the mature plants. Pulling force is aparameter normally used to evaluate root penetration in field conditions [70].Transgenic rice plants overexpressing AtHARDY showed increased drought andsalt tolerance through increased water-use efficiency (WUE), increased photosyn-thetic efficiency, and reduced mean transpiration rate [70]. Although this study didnot report on yield, higher WUE means higher biomass [70].It has been proposed that root hydraulic pressure is a positive regulator of water

uptake by roots. In general, rice has lower hydraulic pressure than that of othercrops like maize. Moreover, there is genetic variation in hydraulic pressure amongdifferent rice cultivars [71]. Generally, drought-tolerant upland cultivars likeAzucena have higher hydraulic pressure than lowland cultivars like IR64 [72].Aquaporins are channel proteins present in the plasma and intracellularmembranes of plant cells, where they facilitate the transport of water, small neutral

12.5 Transgenic Approaches to Improve RSA for Abiotic Stress Tolerance 313

solutes (urea, boric acid, silicic acid), and gases (ammonia, carbon dioxide), andtheir expression is positively correlated with higher water uptake and hydraulicwater pressure [73]; most of them are expressed in roots [74]. In rice, it has beenshown that drought affects the expression of aquaporins in root, but not in leaf [30].In Arabidopsis, simultaneous downregulation of PIP1 (PLASMA MEMBRANEINTRINSCI PROTEIN1) and PIP2 aquaporins using an antisense techniquereduced hydraulic conductivity by 5- to 30-fold. When control and antisensetransgenic lines were subjected to water stress and rewatered, the leaf waterpotential was higher in control plants, which proved that PIP1 and PIP2 playimportant roles in water uptake [75]. Yu et al. proposed a possible role foraquaporins in plant chilling stress [76]. When they subjected two rice cultivars tochilling stress and after subsequent recovery, they found higher expression of someaquaporins in the shoot and root of the chilling-tolerant Somewake cultivar than inthe chilling-sensitive Wasetoitsu cultivar [76]. The expression levels of OsPIP1;1and OsPIP2;1 were significantly higher in roots of Somewake [76]. Lian et al. founddifferential expression of OsPIP genes between upland and lowland rice cultivarsunder drought and ABA treatment [77]. They found OsPIP1;2, Os-PIP1;3,OsPIP2;1, and OsPIP2;5 were more upregulated in the upland cultivar than thelowland cultivar, where they were unchanged or downregulated [77]. Transgenicrice plants overexpressing OsPIP1 were shown to have increased rice seed yield,salt resistance, root hydraulic conductivity, and seed germination rate [65]. RWC3,an aquaporin that was upregulated in an upland cultivar under stress conditions,was introduced into a lowland cultivar and the transgenic lowland cultivar hadbetter water potential than non-transgenic plants [77]. It was reported thataquaporins HvPIP2;2,HvPIP2;5, and HvTIP1;1 contribute mostly for water uptakeby roots in barley [78]. Transgenic tobacco plants, overexpressing TdPIP1;1 andTdPIP;2 genes from durum wheat, were resistant to drought and salinity stresses[79]. All this evidence suggests that aquaporins are a major component of the stressavoidance mechanism that primarily involves the root system. Lorenzo et al. haveidentified a salt stress-induced leucine-rich repeat receptor-like kinase (LLR-RLK)gene Srlk in Medicago truncatula, a legume model crop. Downregulation of Srlkusing antisense technology resulted in transgenics with better root growth undersalt stress [80]. Regulation of lateral root emergence under stress conditions is oneof the important adaptation mechanisms for plants. In M. truncatula, salt-inducedHD-ZIP1 transcription factor HB1 was shown to regulate root architecture understress conditions via LBD1 [81]. de Z�elicourt et al. reported a NAC transcriptionfactor, MtNAC969, which is involved in root development and salt stress tolerance[82].

12.6

Use of Polyamines and Osmotic Regulators in Stress-Induced Modulation of RSA

Polyamines are small ubiquitous polycations that have been suggested to beinvolved in plant responses to various abiotic stresses [83]. It has been reported that

314 12 Improving the Plant Root System Architecture to Combat Abiotic Stresses

the roots of the salt-tolerant cultivar Pokkali accumulated more polyamines such asputrescine and tyramine than the salt-sensitive cultivar “I Kang Pao” upon exposureto salt stress [83]. Boron-deficient tobacco plants had higher amounts of freeputrescine, which led to inhibition of root growth [84]. Putrescine accumulation inrice roots under aluminum toxicity led to growth inhibition of roots; at the sametime, some putrescines are required for the normal growth and development of riceroots [85]. Treatment with S-methylmethionine, an intermediate compound in sulfurmetabolism, led to the accumulation of polyamines in roots and subsequently led tochilling tolerance through increased cell membrane integrity [86].Proline accumulates in the primary root tip of maize in low water potential

conditions, which is mediated by ABA [87]. Proline has been shown to accumulatemore in roots of water stress-resistant wild cultivars of barley than the susceptiblemodern cultivar [88]. Lin et al. reported that NaCl-mediated inhibition of rootgrowth of rice seedlings is associated with increasing levels of proline accumula-tion in roots upon exposure to NaCl [89]. AtP5CR, an important enzyme involved inproline biosynthesis, is expressed at high levels in the apical meristems, includingroot meristems and lateral root primordial [90]. In rice, overexpression ofD0-pyrroline-5-carboxylate synthetase (P5CS) was shown to increase root growthand biomass production under salt treatment [91].

12.7

Hormonal Regulation of Root Architecture and Abiotic Stress Response

Exposure to environmental stresses alters the production and accumulation ofsome plant hormones (e.g., ABA, ethylene, gibberellic acid, and salicylic acid) inplant roots. Signaling through these hormones plays an important role inmodulating the root architecture in response to abiotic stresses [2].It is known that various abiotic stresses (e.g., drought, extreme temperature, flood,

etc.) lead to the increased biosynthesis and accumulation of the stress hormone ABAin plants cells (including root cells), which is accompanied by stress/ABA-inducedexpression of genes [2,92,93]. Functional analysis of both ABA biosynthetic mutants(e.g., aba1, aba2) as well as ABA response mutants (e.g., abi1, abi2) has suggestedthe role of ABA in root development in Arabidopsis and drought-induced inhibitionof lateral roots is partly mediated by ABA [93–95]. Other groups have independentlyshown that both osmotic stress and ABA affect root architecture and lateral rootdevelopment [96,97]. External application of ABA inhibited the negative effect ofNaCl (salt) on roots, suggesting the role of ABA in cellular osmotic adjustmentunder stress [2]. A genetic screen has identified DIG3 (DROUGHT INHIBITIONOF LATERAL ROOTGROWTH3), which is required for ABA-mediated inhibition oflateral root growth and responses to drought stress [93] in Arabidopsis. Plantsectopically expressing ABA-inducible and root-enriched NAC genes (e.g., OsNAC10and 45) and MAPK (OsMAPK5) genes exhibit tolerance to abiotic stress [98].Biosynthesis and accumulation of another stress related phytohormone, ethy-

lene, varies under different stress condition (drought, salinity, flooding, etc.) [2].

12.7 Hormonal Regulation of Root Architecture and Abiotic Stress Response 315

Ethylene has been shown to affect division of root stem cells, root architecture, androot hair formation in Arabidopsis [99,100]. Tomato ER5, a cDNA encoding anethylene-responsive LEA-like protein, is highly induced under drought stress (andwounding) and regulates the RSA of the tomato plant [101]. ABA has been shownto limit ethylene production and thereby control root growth in maize under waterstress [102]. It has been hypothesized to affect water uptake through roots understress conditions, probably by modulating root PIP aquaporin expression [2].Expression of some PIP aquaporin genes in ABA-deficient tomato plants wasupregulated by ABA depletion [2]. Although auxin plays a major role in root growthand branching, its interaction with cytokinin and ethylene is also necessary forproper patterning of roots [103,104]. In Arabidopsis, low boron supply caused aninhibition of primary root growth and increased root hair formation withoutaffecting lateral root growth and number. It was found that boron deficiency affectsthe root growth by controlling auxin and ethylene signaling through AUX1/IAA1(AUXIN 1/ Indole-3-Acetic Acid1)) and EIN1 (ETHYLENE INSENSITIVE1) genes[105]. Exogenous application of salicylic acid has been shown to inhibit the growthof Arabidopsis (probably affecting the PIP aquaporins) and inhibit symplastic watertransport in maize root [2]. Reduction of the root-specific cytokinin (a negativeregulator of root growth) content in transgenic Arabidopsis and tobacco promotedthe elongation of both primary root and lateral roots, and increased the rootbiomass by 60%, while growth and development of the shoot remained unaltered.These transgenic plants with robust root systems also demonstrated highertolerance to severe drought [106].DELLA, a nuclear growth repressor protein, becomes destabilized in the

presence of gibberellic acid, which leads to normal plant growth. It has beenshown that under environmental stresses, stress hormones like ABA and ethylenelead to the accumulation of DELLA protein and subsequent reduction of gibberellicacid through a novel unknown mechanism, which resulted in growth retardation[107]. Arabidopsis mutants of DELLA did not show any growth restraint underenvironmental stresses, which made the mutants more susceptible to stresses thanthe wild-type [107]. Gibberellic acid-mediated hormone response through DELLAwas shown to be controlled by another phytohormone, auxin, which promotes rootdevelopment [108,109]. Phosphate starvation induced modification of the rootarchitecture and root hair induction is modulated by the gibberellic acid–DELLAsignaling pathway in Arabidopsis [110]. The evidence suggests that DELLA acts as alink between different the intrinsic hormone response and stress responsepathways in root system.In rice, only a few F-box proteins have been characterized, which were shown to

be involved in the gibberellic acid signaling pathway [111]. F-box proteins are one ofthe largest protein families with nearly 700 members in Arabidopsis and rice [112].They are an integral component of the SCF (Skp, Cullin, F-box) ligase complex,which provides substrate specificity for the 26S proteasome that degrades theproteins marked by the SCF E3 complex [111]. They have been shown to have animportant role in many developmental pathways mediated by hormones [113].Overexpressed MAIF1, an F-box protein that was induced by auxin, cytokinin, and

316 12 Improving the Plant Root System Architecture to Combat Abiotic Stresses

ABA in rice, led to increased root growth through enhanced cell division underabiotic stress; however, it increased the susceptibility of the plants to stress [112].This shows the importance of growth restraint as an adaptive mechanism understress conditions. The pathway through which MAIF1 regulates growth restraint isunclear; one interesting fact is that MAIF1 is a predicted target of OsmiR446,(a small regulatory RNA; see next paragraph for details) indicating its post-transcriptional regulation [112].Mounting evidence suggests the possible role of plant hormones and stress-

induced reactive oxygen species (ROS) as the main components of stress signalperception and subsequent downstream developmental or morphological adaptiveresponses. It is known that interaction between auxin and cytokinin maintains rootgrowth and development in normal conditions [2,114]. It has been proposed that,under stress conditions, ROS acts as a signal molecule that interacts with the auxinsignaling pathway to control the various plant developmental programs, such as thecell cycle, cell wall plasticity, abiotic stress adaptation, and programmed cell death[114]. Under abiotic stresses, ROS generated in roots have been related to wateruptake properties [2]. Exogenous application of hydrogen peroxide (H2O2), one ofthe most studied ROS, affects water uptake by roots, probably by changingaquaporin pores [2]. The integration of auxin signaling with other hormonesignaling pathways like cytokinin and gibberellic acid, and stress hormones likeABA and ethylene, is well documented [114]. It implies that ROS act as importantintermediate components in stress-mediated signaling pathways that result in thestress-induced adaptive responses in plant.

12.8

Small RNA-Mediated Regulation of RSA and Abiotic Stress Response

Although stress-induced transcriptional gene regulation is well known, theimportance of post-transcriptional gene regulation by small RNAs under abioticstress has recently been evident [63]. Small non-protein coding RNAs (20–30nucleotides long) have been implicated in root growth and development both inmonocot and dicot plants [115,116]. Two major classes of small RNAs, miRNAmicroRNA (miRNA) and trans-acting short-interfering RNA (tasiRNA) and short-interfering RNA (siRNA), negatively regulate the expression of their target genes,mostly by cleaving the mRNA at complementary sequences [116,117]. Recentstudies have shown that the expression of many miRNAs is altered in response tovarious abiotic stresses [63,118–120]. Drought stress-responsive miRNAs havebeen reported in many plant species, such as Arabidopsis [121], rice [122], cowpea[123], tobacco [124], Triticum dicoccoides [125], soybean [126], Phaseolus vulgaris [127],and so on. miR165, miR167, miR159, miR158, miR169, miR171, miR156, miR168,miR393, and miR396 have been shown to be drought responsive in Arabidopsis[63,120,128]. Expression of miR474, miR398, miR156, miR894, and miR1432 wasinduced, whereas miR166 and miR171 were downregulated, in T. dicoccoides rootunder drought stress [120,125]. Submergence-responsive miRNAs potentially

12.8 Small RNA-Mediated Regulation of RSA and Abiotic Stress Response 317

involved in root adaptation have been identified in maize [129]. Hypoxia-inducedmodification of the root architecture was shown to be accompanied by alteredexpression of many miRNA and siRNAs in Arabidopsis [130], indicating a potentialrole of these small RNAs in hypoxia stress-responsive adaptation of the root. Abioticstresses created by altered nutrient homeostasis also impact the expression of manysmall RNAs [63,120]. Here, a few important small RNAs are mentioned that arepredominantly present in roots and impact the tolerance of plants to abiotic stresses.MiR164, which targets NAC1 transcription factor, has been shown to regulate

root development through auxin homeostasis. Since the induction of miR164 leadsto reduced NAC1 expression and reduced lateral root emergence [131], down-regulation of miR164 under drought stress has been hypothesized to increase theroot/shoot ratio by inducing the expression of NAC1 in M. truncatula [132]. InArabidopsis, auxin response factors ARF6 and ARF8, which are targets of miR167,regulate adventitious root formation [133], whereas miR160 targets ARF10, ARF16,and ARF17, which are required for proper root development and adventitious rootformation [131,133]. Various abiotic stress responses (drought, salinity, hypoxia,etc.) have been shown to either induce or downregulate the expression of miR167in different species, such as Arabidopsis, maize, and rice [63,120]. This indicates thepotential role of miR167 in abiotic stress-responsive root growth, which could beused for root stress tolerance. Similarly, the expression pattern of miR160 changesunder low sulfate and heavy metals (in Brassica), heat stress (in wheat), and ABAstress and UV irradiation (in Arabidopsis), indicating its potential role in adaptiveroot morphogenesis under various stresses [63,120]. However, further work wouldbe necessary to confirm this hypothesis. miR171 expression is also influenced byvarious abiotic stresses (such as hypoxia, drought, and salinity) in different species[63,120]. miR171 has been suggested to regulate radial patterning of root bytargeting SCL (SCARECROW LIKE) genes [63,120]. Similarly, differential expres-sion of miR156/miR157 (target: SPL genes) or miR166/miR165 (target: HD-ZIPIIIgenes) genes under various stresses [63,120] is likely to affect root development, astheir target genes have been implicated in root development [116]. The expressionof miR393, which targets F-box auxin receptor gene (AFB) transcripts (includingTIR1), has been shown to be induced by various abiotic stresses (such as drought,cold, salinity, and ABA stress) in different species [120,134]. Ectopic expression ofmiR393-resistant TIR1 (that is involved in auxin signaling) led to robust rootbranching and altered RSA in Arabidopsis [135]. Additionally, the nitrate-responsivemiR393/AFB3 (target of miR393) module has been shown to regulate rootarchitecture in Arabidopsis [136]. Thus, miR393 plays a crucial role in stress/nutrient-induced adaptive root morphogenesis through auxin signaling.In Arabidopsis, miR399, miR395, and miR398 are expressed in response to

phosphate, sulfate, and copper starvation conditions (stress), respectively [63,120].Experimental evidence indicates that under phosphate starvation, PHR (PHOS-PHATE STARVATION1)-mediated induction of miR399 downregulates the targettranscript of phosphate transporter 2 PHO2 (PHOSPHATE TRANSPORTER2),UBC24 (UBIQUITIN E2 CONJUGASE24) in root [63,120], and maintainsphosphate transport and homeostasis through the root [137,138]. miR395 has been

318 12 Improving the Plant Root System Architecture to Combat Abiotic Stresses

shown to regulate sulfate homeostasis (in sulfate-deprived condition) as well ashypoxia response of roots [63,120]. The expression of miR399, which targets twoZn/Cu-SUPEROXIDE DISMUTASE (CSD1 and CSD2) genes in Arabidopsis, isinduced by copper starvation and other stresses, and is required for copperhomeostasis [63,120]. Many miRNAs are involved in ABA-mediated stressresponses that affect root architecture [120]. Endogenous siRNAs derived from apair of natural cis-antisense transcripts were shown to regulate salt tolerance(accompanying changes in root system) in Arabidopsis [139]. It is obvious from theaforesaid examples that the expression of many miRNAs responds to the variousabiotic stresses in root. An increasing body of evidence indicates their potential rolein stress-responsive adaptive morphogenesis of the root architecture, which islikely to provide stress tolerance to the plant. However, more functional analysis isrequired to shed light on this aspect.

12.9

Application of Phenomics in Understanding Stress-Associated RSA

Characterization of the RSA is one of the important prerequisites to understand thedevelopment and function of vascular plants. Plants suffer from various kinds ofabiotic stresses that severely affect their RSA as well as overall plant growth [140].The huge variation in constitutive RSA traits exhibited by different genotypes isfurther complicated by an additional layer of stress-responsive variation underadverse environmental conditions. This complex nature of variation in RSA traitsand less accessibility of roots makes it difficult to characterize the root phenotype oflarge populations under different conditions [140]. Recent technological advanceshave developed a high-throughput phenotypic tool along with highly efficientcomputational models/software for scoring the root phenotype and its relation toother traits or stresses [140]. Various software and algorithms have been developedto study the different RSA traits. Software designed to explore kinematic ormorphometric root growth and gravitropism includes “RootTrace” [141], “relativeelemental growth rate (REGR)” analysis [142], “Kine-Root” [143], and “Root Flow RT”[144]. These programs focus on analyzing root growth from a time series of imagestaken by a high-resolution camera. By applying the above tools and algorithms, rootphenotypic study can be easily performed. These tools provide informationregarding the gravitropism response, relative growth rate, kinematics details, andreal-time root growth. Using these tools it is also possible to understand how the rootgrowth parameters change in response to various stresses [145].Additional phenomics tools related to RSA in plants include the “GiA Roots

software” (General Image Analysis of Roots), a semi-automated software tooldesigned specifically for the high-throughput analysis of root system images [146].“GiA Roots” includes user-assisted algorithms to distinguish roots from thebackground and a fully automated pipeline that extracts dozens of root systemphenotypes [146]. Another tool to study RSA in plants is “Shovelomics,” which is ahigh-throughput phenotyping tool for field plants and is used to study the

12.9 Application of Phenomics in Understanding Stress-Associated RSA 319

phenotypic profiling of the maize roots [147]. “Minirhizotrons” is another tool forthe non-invasive study of RSA of plants having fine roots. In this method, aminirhizotron tube is kept below the root of the plant and images are captured,which are further processed digitally to produce detailed information regarding theRSA [148]. Use of “RootReader3D” is another approach to study the RSA of plantsunder laboratory conditions; it gives information about the gravitropic response,root circumnutation, average length, and so on [149]. Another approach is the useof “RooTrak software,” which is a X-ray micro-computed tomography (mCT) tool forvisualizing plant root systems within their natural soil environment non-invasively[150]. Use of these advanced technologies is very helpful to study multiplecharacters at a time for huge numbers of plants in a less-laborious and time-savingmanner. This would be instrumental in exploring the huge diversity amongcultivars/species as well performing genetic screening to understand the rootarchitecture and its adaptability to the environment. High-throughput NGStechnology is being steadily adopted by plant biologists to discover genesresponsible for the variation in phenotype or stress responses among differentplants. However, correlating the expression and function of thousands of geneswith specific traits has been difficult. Large-scale analysis of different phenotypic orphysiological traits using the aforesaid phenomics technology can bridge the gapbetween genomics and phenotype, and thus help us to reach our goal faster.

12.10

Conclusion and Future Perspectives

Decreasing agricultural resources and increasing abiotic stress conditions (mainlydue to geo climatic changes) are major factors that adversely limit crop productivityin agriculture. The strategy used in the first “Green Revolution” has reached its limitand additionally created many negative effects. Root architecture, which plays acrucial role in the optimum growth and productivity of a plant, had been overlookedby plant scientists. Sensing its importance, people have started to investigate rootarchitecture and its adaptive responses to stresses. In an effort to improve rootsystems, both breeding and transgenic approaches are being followed. Additionally,the combinatorial use of today’s advanced NGS and phenomics technologies ishelping to speed up the characterization of root architectural variation and its geneticbasis, which in turn could assist us to generate stress-tolerant crops without any yieldpenalty. The huge dataset obtained from stress studies, hormonal regulation, andsmall RNA regulation studies through NGS and phenomics approaches can be usedin MAS-based breeding as well as in the production of transgenic crops with thedesired root characteristics. Developing plants with robust RSA will make themmore efficient in exploiting underground water and nutrients, and tolerant toenvironmental stresses. This would also help in the absorption of more atmosphericCO2 (a greenhouse gas), fixing the CO2 into underground root systems and therebycontributing to the effort to reduce global warming by greenhouse gases.Both in breeding and transgenic approaches, while improving one character,

we often end up downgrading others, probably because of the very complex

320 12 Improving the Plant Root System Architecture to Combat Abiotic Stresses

and multigenic nature of the regulation of stress tolerance and adaptiveresponses. For example, a plant with an improved root system may havereduced quality of leaf or grains. Using advanced phenomic and modelingtechnology, it is possible to consider multiple desired characters (such as rootlength, branches, angles, biomass, penetrance, moisture holding capacity, root/shoot ratio, etc.) and exclude undesired characters in a time-saving and less-laborious manner [8]. The influence of soil microbes on the rhizosphericgrowth and distribution of roots (in relation to stress) is an interesting aspect.Den Herder et al. have highlighted a few challenges that plant scientistsshould tackle while trying to improve root architecture and crop yield: (i) abetter understanding of root development and its interaction with biotic andabiotic factors, (ii) detailed analysis of root–shoot relationships and the impactof root development on plant fitness, and (iii) integration of all aspects of rootbiology (root structure, environmental impact, resources, etc.) in a simpleexisting model (related to phenomics) and performing combinatorial studieson root functioning [8]. Epigenetic processes (such as DNA methylation andhistone modifications), small RNAs, and transposable elements play essentialroles in modulating gene activity in response to environmental stimuli [151].Abiotic stresses can induce changes in gene expression through epigeneticregulation, which can be an important aspect to consider for improvingtolerance to abiotic stresses in crops. Mutation in the epigenetic regulatorSWP1 has been shown to significantly increase root length and branching inArabidopsis [152]. The functional analysis of this kind of epigenetic regulator aswell as root epigenomics studies in crop system can provide novel moleculartools for developing stress tolerance through improved RSA. The combinationof genomics, proteomics, and phenomics approaches should speed up theidentification and characterization of regulators related to RSA traits and stresstolerance. Thus, although challenging, the possibility of increasing crop yieldand tolerance to stresses through the improvement of root system is immense.

Acknowledgments

A.K.S. acknowledges a Ramalingaswami Fellowship by the Department of Biotech-nology (DBT), India (BT/HRD/35/02/06/2008). Fellowships to K.M. by DBT, India(BT/PR3292/AGR/2/811/2011), to V.G. by Center for Scientific and IndustrialResearch (CSIR, India), and to S.B. (NIPGR) are sincerely acknowledged.

References

1 Tester, M. and Langridge, P. (2010) Science,327, 818–822.

2 Aroca, R. Porcel, R., and Ruiz-Lozano, J.M.(2011) J. Exp. Bot., 63, 43–57.

3 Hodge, A., Berta, G., Doussan, C. et al.(2009) Plant Soil, 21, 153–187.

4 Lynch, J.P. (1995) Plant Physiol., 109, 7–13.5 Lynch, J. (2007) Aust. J. Bot., 55, 493–512.

References 321

6 Mora, C., Tittensor, D.P., Adl, S. et al.(2011) PLoS Biol., 9, e1001127.

7 Gewin, V. (2010) Nature, 466, 552–553.8 Den Herder, G., Van Isterdael, G.,Beeckman, T. et al. (2010) Trends Plant Sci.,15, 600–607.

9 Boyer, J.S. (1982) Science, 218, 443–448.10 Pennisi, E. (2008) Science, 320,

171–173.11 Aichinger, E., Kornet, N., Friedrich, T. et al.

(2012) Annu. Rev. Plant Biol., 63,615–636.

12 Sarkar, A.K., Luijten, M., Miyashima, S.et al. (2007) Nature, 446, 811–814.

13 Hochholdinger, F. and Tuberosa, R. (2009)Curr. Opin. Plant Biol., 12, 172–177.

14 Hochholdinger, F., Park, W.J., Sauer, M.et al. (2004) Trends Plant Sci., 9, 42–48.

15 Malamy, J.E. (2005) Plant Cell Environ.,28, 67–77.

16 Haecker, A., Gross-Hardt, R., Geiges, B.et al. (2004) Development, 131, 657–668.

17 Zhao, Y., Hu, Y., Dai, M. et al. (2009) PlantCell, 21, 736–748.

18 Kell, D.B. (2012) Philos. Trans. R. Soc. Lond.B Biol. Sci., 367, 1589–1597.

19 Kell, D.B. (2011) Ann. Bot., 108, 407–418.20 Donald, C.M. (1968) Euphytica, 17,

385–403.21 de Dorlodot, S., Forster, B., Pages, L. et al.

(2007) Trends Plant Sci., 12, 474–481.22 Uga, Y., Ebana, K., Abe, J. et al. (2009)

Breed. Sci., 59, 87–93.23 Nguyen, H.T., Babu, R.C., and Blum, A.

(1997) Crop Sci., 37, 1426–1434.24 Abd Allah, A.A., Badawy, S.A., Zayed, B.A.

et al. (2010) Int. J. Agric. Biol. Sci., 1, 83–87.25 Babu, R.C. (2010) Electron. J. Plant Breed.,

1, 1133–1141.26 Collins, N.C., Tardieu, F., and Tuberosa, R.

(2008) Plant Physiol., 147, 469–486.27 Uga, Y., Okuno, K., and Yano, M. (2011) J.

Exp. Bot., 62, 2485–2494.28 Uga, Y., Okuno, K. and Yano, M. (2008)

Breed. Sci., 58, 7–14.29 Landi, P., Sanguineti, M.C., Liu, C. et al.

(2007) J. Exp. Bot., 58, 319–326.30 Henry, A., Cal, A.J., Batoto, T.C. et al.

(2012) J. Exp. Bot., 63, 4751–4763.31 Kondo, M., Murty, M.R., and Aragones, D.

V. (2000) Soil Sci. Plant Nutr., 46, 721–732.32 Fukai, S. and Cooper, M. (1995) Field Crops

Res., 40, 67–86.

33 Uga, Y. et al. (2013) Nat. Genet., 45 (9),1097–1102.

34 Steele, K.A., Price, A.H., Shashidhar,H.E. et al. (2006) Theor. Appl. Genet.,112, 208–221.

35 Steele, K.A., Virk, D.S., Kumar, R. et al.(2007) Field Crops Res., 101, 180–186.

36 Hemamalini, G.S., Shashidhar, H.E.,and Hittalmani, S. (2000) Euphytica,112, 69–78.

37 Suryapriya, P., Snehalatha, A., Kayalvili, U.et al. (2009) Bioinformation, 3, 279–281.

38 Sharp, R.E., Silk, W.K., and Hsiao, T.C.(1988) Plant Physiol., 87, 50–57.

39 Ruta, N., Liedgens, M., Fracheboud, Y.et al. (2010) Theor. Appl. Genet., 120,621–631.

40 Hund, A., Fracheboud, Y., Soldati, A. et al.(2004) Theor. Appl. Genet., 109, 618–629.

41 Tuberosa, R., Sanguineti, M.C., Landi, P.et al. (1998) Theor. Appl. Genet., 97,744–755.

42 Zhu, J., Kaeppler, S., and Lynch, J. (2005)Theor. Appl. Genet., 111, 688–695.

43 Vadez, V., Krishnamurthy, L., Kashiwagi,J.W. et al. (2007) J. Semi-Arid Trop. Agric.Res., 4, 1–61.

44 Morgan, J.M. (1997) Aust. J. Agri. Res.,51, 971–978.

45 Price, A.H. (2006) Trends Plant Sci.,11, 213–216.

46 Abdurakhmonov, I.Y. and Abdukarimov, A.(2008) Int. J. Plant Genomics, 2008, 574927.

47 Yu, J., Holland, J.B., McMullen, M.D. et al.(2008) Genetics, 178, 539–551.

48 Rosyara, U.R., Gonzalez-Hernandez, J.L.,Glover, K.D. et al. (2009) Theor. Appl.Genet., 118, 1617–1631.

49 Yadav, R.S., Sehgal, D., and Vadez, V.(2011) J. Exp. Bot., 62, 397–408.

50 Huang, T.L., Nguyen, Q.T., Fu, S.F. et al.(2012) Plant Mol. Biol., 80, 587–608.

51 Takagi, H. et al. (2013) Plant J., 74 (1),174–183.

52 Yokotani, N., Ichikawa, T., Kondou, Y. et al.(2013) J. Plant Res., 126, 131–139.

53 Kuzuoglu-Ozturk, D., Cebeci Yalcinkaya,O., Akpinar, B.A. et al. (2012) Planta,236, 1081–1092.

54 Poroyko, V., Hejlek, L.G., Spollen, W.G.et al. (2005) Plant Physiol., 138, 1700–1710.

55 Kong, X., Pan, J., Zhang, M. et al. (2011)Plant Cell Environ., 34, 1291–1303.

322 12 Improving the Plant Root System Architecture to Combat Abiotic Stresses

56 Ouyang, B., Yang, T., Li, H. et al. (2007)J. Exp. Bot., 58, 507–520.

57 Wei, J.-Z., Tirajoh, A., Effendy, J. et al.(2000) Plant Sci., 159, 135–148.

58 Ruzicka, D., Barrios-Masias, F.,Hausmann, N. et al. (2010) BMC PlantBiol., 10, 75.

59 Xu, W.F. and Shi, W.M. (2006) Ann. Bot.,98, 965–974.

60 Manaa, A., Ben Ahmed, H., Valot, B. et al.(2011) J. Exp. Bot., 62, 2797–2813.

61 Chen, C.C.S. and Plant, A.L. (1999) J. Exp.Bot., 50, 677–687.

62 Poroyko, V., Spollen, W.G., Hejlek, L.G.et al. (2007) J. Exp. Bot., 58, 279–289.

63 Shukla, L.I., Chinnusamy, V., and Sunkar,R. (2008) Biochim. Biophys. Acta,1779, 743–748.

64 Ding, D., Zhang, L., Wang, H. et al. (2009)Ann. Bot., 103, 29–38.

65 Liu, Z., Kumari, S., Zhang, L. et al. (2012)PLoS ONE, 7, e39786.

66 Xie, Q., Frugis, G., Colgan, D. et al. (2000)Genes Dev., 14, 3024–3036.

67 He, X.J., Mu, R.L., Cao, W.H. et al. (2005)Plant J., 44, 903–916.

68 Hao, Y.J., Wei, W., Song, Q.X. et al. (2011)Plant J., 68, 302–313.

69 Jeong, J.S., Kim, Y.S., Baek, K.H. et al.(2010) Plant Physiol., 153, 185–197.

70 Karaba, A., Dixit, S., Greco, R. et al.(2007) Proc. Natl. Acad. Sci. USA,104, 15270–15275.

71 Matsuo, N., Ozawa, K., and Mochizuki, T.(2009) Plant Soil, 316, 25–34.

72 Gowda, V.R.P., Henry, A., Yamauchi, A.et al. (2011) Field Crops Res., 122, 1–13.

73 Maurel, C., Verdoucq, L., Luu, D.T. et al.(2008) Annu. Rev. Plant Biol., 59, 595–624.

74 Bramley, H., Turner, N.C., Turner, D.W.et al. (2009) Plant Physiol., 150, 348–364.

75 Martre, P., Morillon, R., Barrieu, F. et al.(2002) Plant Physiol., 130, 2101–2110.

76 Yu, X., Peng, Y.H., Zhang, M.H. et al.(2006) Cell Res., 16, 599–608.

77 Lian, H.L., Yu, X., Ye, Q. et al. (2004) PlantCell Physiol., 45, 481–489.

78 Knipfer, T., Besse, M., Verdeil, J.L. et al.(2011) J. Exp. Bot., 62, 4115–4126.

79 Ayadi, M., Cavez, D., Miled, N. et al.(2011) Plant Physiol. Biochem., 49,1029–1039.

80 de Lorenzo, L., Merchan, F., Laporte, P.et al. (2009) Plant Cell, 21, 668–680.

81 Ariel, F.D., Diet, A., Crespi, M. et al. (2010)Plant Signal. Behav., 5, 1666–1668.

82 de Z�elicourt, A., Diet, A., Marion, J. et al.(2012) Plant J., 70, 220–230.

83 Gill, S.S. and Tuteja, N. (2010) Plant Signal.Behav., 5, 26–33.

84 Camacho-Cristobal, J.J., Maldonado, J.M.,and Gonzlaez-Fontes, A. (2005) J. PlantPhysiol., 162, 921–928.

85 Wang, J.W. and Kao, C.H. (2006) Plant Soil,288, 373–381.

86 Racz, I., Paldi, E., Szalai, G. et al. (2008)J. Plant Physiol., 165, 1483–1490.

87 Ober, E.S. and Sharp, R.E. (1994) PlantPhysiol., 105, 981–987.

88 Bandurska, H. and Stroi�nski, A. (2003)Acta Physiol. Plant., 25, 55–61.

89 Lin, C.C. and Kao, C.H. (1996) Plant Sci.,114, 121–128.

90 Hua, X.J., van de Cotte, B., Van Montagu,M. et al. (1997) Plant Physiol., 114,1215–1224.

91 Kavi Kishor, P.B., Sangam, S., Amrutha,R.N. et al. (2005) Curr. Sci., 88, 424–438.

92 Tuteja, N. (2007) Plant Signal. Behav.,2, 135–138.

93 Xiong, L., Wang, R.G., Mao, G. et al. (2006)Plant Physiol., 142, 1065–1074.

94 Brady, S.M., Sarkar, S.F., Bonetta, D. et al.(2003) Plant J., 34, 67–75.

95 Leon-Kloosterziel, K.M., Gil, M.A., Ruijs,G.J. et al. (1996) Plant J., 10, 655–661.

96 De Smet, I., Signora, L., Beeckman, T. et al.(2003) Plant J., 33, 543–555.

97 Deak, K.I. and Malamy, J. (2005) Plant J.,43, 17–28.

98 Hadiarto, T. and Tran, L.S. (2010) Plant CellRep., 30, 297–310.

99 Patrick, B., Antonin, L., Servane, L.L. et al.(2009) Plant Signal. Behav., 4, 44–46.

100 Ortega-Martinez, O., Pernas, M., Carol, R.J. et al. (2007) Science, 317, 507–510.

101 Zegzouti, H., Jones, B., Marty, C. et al.(1997) Plant Mol. Biol., 35, 847–854.

102 Sharp, R.E. and LeNoble, M.E. (2002)J. Exp. Bot., 53, 33–37.

103 Peret, B., De Rybel, B., Casimiro, I. et al.(2009) Trends Plant Sci., 14, 399–408.

104 Laplaze, L., Benkova, E., Casimiro, I. et al.(2007) Plant Cell, 19, 3889–3900.

References 323

105 Martin-Rejano, E.M., Camacho-Cristobal,J.J., Herrera-Rodriguez, M.B. et al. (2011)Physiol. Plant., 142, 170–178.

106 Werner, T., Nehnevajova, E., Kollmer, I.et al. (2010) Plant Cell, 22, 3905–3920.

107 Achard, P., Cheng, H., De Grauwe, L. et al.(2006) Science, 311, 91–94.

108 Fu, X. and Harberd, N.P. (2003) Nature,421, 740–743.

109 Gou, J., Strauss, S.H., Tsai, C.J. et al. (2010)Plant Cell, 22, 623–639.

110 Jiang, C., Gao, X., Liao, L. et al. (2007) PlantPhysiol., 145, 1460–1470.

111 Lechner, E., Achard, P., Vansiri, A. et al.(2006) Curr. Opin. Plant Biol., 9, 631–638.

112 Yan, Y.S., Chen, X.Y., Yang, K. et al. (2011)Mol. Plant Pathol., 4, 190–197.

113 Dreher, K. and Callis, J. (2007) Ann. Bot.,99, 787–822.

114 Tognetti, V.B., Muhlenbock, P., and VanBreusegem, F. (2012) Plant Cell Environ.,35, 321–333.

115 Meng, Y., Ma, X., Chen, D. et al. (2010)Biochem. Biophys. Res. Commun., 393,345–349.

116 Khan, G.A., Declerck, M., Sorin, C. et al.(2011) Plant Mol. Biol., 77, 47–58.

117 Allen, E., Xie, Z., Gustafson, A.M. et al.(2005) Cell, 121, 207–221.

118 Sunkar, R. and Zhu, J.K. (2004) Plant Cell,16, 2001–2019.

119 Phillips, J.R., Dalmay, T., and Bartels, D.(2007) FEBS Lett., 581, 3592–3597.

120 Khraiwesh, B., Zhu, J.K., and Zhu, J.(2011) Biochim. Biophys. Acta, 1819,137–148.

121 Sunkar, R., Chinnusamy, V., Zhu, J. et al.(2007) Trends Plant Sci., 12, 301–309.

122 Zhou, L., Liu, Y., Liu, Z. et al. (2010) J. Exp.Bot., 61, 4157–4168.

123 Barrera-Figueroa, B.E., Gao, L., Diop, N.N.et al. (2011) BMC Plant Biol., 11, 127.

124 Frazier, T.P., Sun, G., Burklew, C.E. et al.(2011)Mol. Biotechnol., 49, 159–165.

125 Kantar, M., Lucas, S.J., and Budak, H.(2011) Planta, 233, 471–484.

126 Kulcheski, F.R., de Oliveira, L.F., Molina,L.G. et al. (2011) BMC Genomics, 12, 307.

127 Arenas-Huertero, C., Perez, B., Rabanal, F.et al. (2009) Plant Mol. Biol., 70, 385–401.

128 Liu, H.H., Tian, X., Li, Y.J. et al. (2008)RNA, 14, 836–843.

129 Zhang, Z., Wei, L., Zou, X. et al. (2008)Ann. Bot., 102, 509–519.

130 Moldovan, D., Spriggs, A., Yang, J. et al.(2009) J. Exp. Bot., 61, 165–177.

131 Guo, H.S., Xie, Q., Fei, J.F. et al. (2005)Plant Cell, 17, 1376–1386.

132 Wang, T., Chen, L., Zhao, M. et al. (2011)BMC Genomics, 12, 367.

133 Gutierrez, L., Bussell, J.D., Pacurar,D.I. et al. (2009) Plant Cell, 21,3119–3132.

134 Chen, X. (2009) Annu. Rev. Cell Dev. Biol.,25, 21–44.

135 Chen, Z.H., Bao, M.L., Sun, Y.Z. et al.(2011) Plant Mol. Biol., 77, 619–629.

136 Vidal, E.A., Araus, V., Lu, C. et al. (2010)Proc. Natl. Acad. Sci. USA, 107,4477–4482.

137 Fujii, H., Chiou, T.J., Lin, S.I. et al. (2005)Curr. Biol., 15, 2038–2043.

138 Chiou, T.J., Aung, K., Lin, S.I. et al. (2006)Plant Cell, 18, 412–421.

139 Borsani, O., Zhu, J., Verslues, P.E. et al.(2005) Cell, 123, 1279–1291.

140 Zhu, J., Ingram, P.A., Benfey, P.N. et al.(2011) Curr. Opin. Plant Biol., 14,310–317.

141 French, A., Ubeda-Tomas, S., Holman,T.J. et al. (2009) Plant Physiol., 150,1784–1795.

142 Walter, A., Spies, H., Terjung, S. et al.(2002) J. Exp. Bot., 53, 689–698.

143 Basu, P., Pal, A., Lynch, J.P. et al. (2007)Plant Physiol., 145, 305–316.

144 van der Weele, C.M., Jiang, H.S.,Palaniappan, K.K. et al. (2003) PlantPhysiol., 132, 1138–1148.

145 Hodge, A. (2004) New Phytol., 162, 9–24.146 Galkovskyi, T., Mileyko, Y., Bucksch, A.

et al. (2012) BMC Plant Biol., 12, 116.147 Trachsel, S., Kaeppler, S.M., Brown, K.M.

et al. (2011) Plant Soil, 341, 75–87.148 Hendrick, R.L. and Pregitzer, S.K. (1992)

Plant Soil, 143, 283–288.149 Clark, R.T., MacCurdy, R.B., Jung, J.K. et al.

(2011) Plant Physiol., 156, 455–465.150 Mairhofer, S., Zappala, S., Tracy, S.R. et al.

(2011) Plant Physiol., 158, 561–569.151 Henderson, I.R. and Jacobsen, S.E. (2007)

Nature, 447, 418–424.152 Singh, S., Singh, A., Roy, S. et al. (2012)

Plant Signal. Behav., 7.

324 12 Improving the Plant Root System Architecture to Combat Abiotic Stresses