Strigolactones interact with other phytohormones to modulate plant root growth and development

Huwi Sun,Wiqiang Li,Davi J.Burritt,Hongtao Tian,Hng Zhang,Xiaohan Liang,Yuhn Miao,Mohamma Golam Mostofa,Lam-Son Phan Tran,f,*

a Laboratory of Rice Biology in Henan Province,Collaborative Innovation Center of Henan Grain Crops,College of Agronomy,Henan Agricultural University,Zhengzhou 450046,Henan,China

b Jilin Da’an Agro-ecosystem National Observation Research Station,Changchun Jingyuetan Remote Sensing Experiment Station,Key Laboratory of Mollisols Agroecology,Northeast Institute of Geography and Agroecology,Chinese Academy of Sciences,Changchun 130102,Jilin,China

c State Key Laboratory of Cotton Biology,Henan Joint International Laboratory for Crop Multi-Omics Research,School of Life Sciences,Henan University,Kaifeng 475001,Henan,China

d Department of Botany,University of Otago,Dunedin 9054,New Zealand

e Institute of Genomics for Crop Abiotic Stress Tolerance,Department of Plant and Soil Science,Texas Tech University,Lubbock,TX 79409,USA

f Institute of Research and Development,Duy Tan University,03 Quang Trung,Da Nang 550000,Vietnam

Keywords:Cross-regulation Development Phytohormones Roots Signaling Strigolactones

ABSTRACT Strigolactones(SLs),which are biosynthesized mainly in roots,modulate various aspects of plant growth and development.Here,we review recent research on the role of SLs and their cross-regulation with auxin,cytokinin,and ethylene in the modulation of root growth and development.Under nutrientsufficient conditions,SLs regulate the elongation of primary roots and inhibit adventitious root formation in eudicot plants.SLs promote the elongation of seminal roots and increase the number of adventitious roots in grass plants in the short term,while inhibiting lateral root development in both grass and eudicot plants.The effects of SLs on the elongation of root hairs are variable and depend on plant species,growth conditions,and SL concentration.Nitrogen or phosphate deficiency induces the accumulation of endogenous SLs,modulates root growth and development.Genetic analyses indicate cross-regulation of SLs with auxin,cytokinin,and ethylene in regulation of root growth and development.We discuss the implications of these studies and consider their potential for exploiting the components of SL signaling for the design of crop plants with more efficient soil-resource utilization.

Contents

1.Introduction........................................................................................................1518

2.SL biosynthesis and signal transduction..................................................................................1518

3.Karrikin(KAR)signal transduction......................................................................................1519

4.The diverse roles of SLs in regulating plant root architecture.................................................................1520

4.1.SLs modulate PR and SR elongation in plants........................................................................1520

4.2.SLs modulate AR formation and elongation..........................................................................1521

4.3.SLs modulate LR formation in both grass and eudicot plant species......................................................1521

4.4.Potential effects of SLs on RH elongation and density in eudicot plant species under various growth conditions.................1522

5.Cross-regulation between SLs and other signaling pathways in root growth and development.....................................1522

5.1.Cross-regulation between SL signaling and auxin signaling.............................................................1522

5.2.Cross-regulation between SL and CK signaling.......................................................................1523

5.3.Cross-regulation of SL and/or KAR signaling with ethylene signaling.....................................................1523

5.4.Cross-regulation between SL signaling and KAR signaling..............................................................1524

6.Manipulation of SL signaling to design root architecture and improve soil nutrient utilization in crop plants.........................1524

7.Concluding and perspective remarks....................................................................................1524

CRediT authorship contribution statement.................................................................................1525

Declaration of competing interest......................................................................................1525

Acknowledgments...................................................................................................1525

References.........................................................................................................1525

1.Introduction

Plants sense the availability of water and nutrients in the soil through their root systems,and root morphology influences plant growth and development.At the early seedling stage,root systems can be classified as taproot and fibrous types for eudicot and monocot(grass)plant species,respectively.Most eudicots are characterized by a typical allorhizic or taproot system that includes the embryonic primary roots(PRs),adventitious roots(ARs)(from any non-root tissues),and lateral roots(LRs)that originate from PRs(Fig.1A)[1,2].The fibrous or secondary homorhizic root system of grasses consists of three types of roots:embryonic seminal roots(SRs),ARs(also called crown roots,CRs),and LRs originating from SRs or ARs(Fig.1B)[1].On the micro-scale,root hairs(RHs)develop and extend from epidermal root cells,and are common to both grasses and eudicots(Fig.1C,D).RHs effectively expand the surface area of the root system and increase the capacity of plants to obtain nutrients and water from the soil[3].

Root growth and development are regulated by environmental factors such as drought,salinity,light and nutrient conditions,and by intrinsic factors such as phytohormones[4-6].Recently,a class of phytohormones known as strigolactones(SLs)and their derivatives have been reported[7-10]to contribute to a variety of processes of plant growth and development,including shoot branching,root growth,and plant responses to a wide range of abiotic and biotic stresses.

SLs or their derivatives have been found in plant roots and root exudates[11-14],and can be transported from roots to shoots,where they can inhibit shoot branching[15-17].Mutations in genes participating in SL biosynthesis and signaling have been reported in several plant species,supporting genetic studies[18-22]of SL biosynthesis and signal transduction.Several recent reviews have focused on the roles of SLs and their crossregulation with other phytohormones to inhibit shoot branching[10,17-18]and leaf senescence[23].However,a comprehensive review of the roles of SLs and their cross-regulation with other phytohormones in modulating root growth to regulate the absorption of nutrients is lacking.In this review,we fill this gap and discuss recently published literature[24]associated with these findings.

2.SL biosynthesis and signal transduction

Various genes involved in SL biosynthesis and signaling in rice(Oryza sativa),Arabidopsis thaliana,pea(Pisum sativum)and petunia(Petunia hybrida)have been identified,shedding light on these pathways[7-10].Two structurally distinct groups of SLs,canonical and non-canonical,have been identified in plants.Canonical SLs contain the A,B,C and D(butanolide)ring system,and noncanonical SLs lack the A,B,or C ring,but contain the enol ether-D ring moiety,which is necessary for biological activities[7,25].SLs are biosynthesized from all-trans-β-carotene by the sequential action of several enzymes,encoded by the DWARF 27(AtD27),MORE AXILLARY GROWTH 3(MAX3)and MAX4 genes in Arabidopsis and the corresponding D27,D17 and D10 genes in rice,to produce carlactone in plastids(Fig.2A).The MAX3/D17 and MAX4/D10 genes encode respectively CAROTENOID CLEAVAGE DIOXYGENASE 7(CCD7)and CCD8,the key enzymes in SL biosynthesis[26].Carlactone is then converted to carlactonoic acid by MAX1(a cytochrome P450 enzyme)in Arabidopsis or to other noncanonical SLs(hydroxyl carlactone derivatives)[9,12,27,28]such as 18-hydroxycarlactonoic acid in Lotus japonicus by the MAX1 homolog LjMAX1[14],whereas it is converted to canonical SL(orobanchol)by two P450 enzymes,Os1g0700900/CYP711A2 and Os1g0701400/CYP711A3,in rice,in the cytosol(Fig.2A)[29].LATERAL BRANCHING OXIDOREDUCTASE(LBO)acts downstream of MAX1 to produce a non-canonical SL compound in Arabidopsis[8,30].A recent study[31]showed that anα/β-hydrolase enzyme,CARBOXYLESTERASE 15(CXE15),functions in SLs catabolism and is evolutionarily conserved in seed plants.These findings collectively imply that although SL compounds have diverse structures,their core biosynthetic pathway[9]is conserved in many plant species.

Fig.1.Morphology of the roots of Arabidopsis thaliana(a eudicot)and rice(Oryza sativa,a grass).(A)The root of a 9-day-old Arabidopsis seedling.Scale bar,0.5 cm.(B)The root of a 9-day-old rice seedling.Scale bar,1 cm.The red boxes indicate the root hair region.(C)Root hairs of Arabidopsis.Scale bar,0.5 mm.(D)Root hairs of rice.Scale bar,1 mm.AR,adventitious root;CR,crown root;LR,lateral root;SR,seminal root;PR,primary root.RH,root hair.

Fig.2.A model of the biosynthesis and catabolism and signaling pathway of strigolactones(SLs)in the modulation of root growth and development(A)SLbiosynthetic components in Arabidopsis thaliana and rice(Oryza sativa).All-trans-βcarotene is converted to carlactone by Arabidopsis thaliana DWARF27(AtD27)/D27,MORE AXILLARY GROWTH 3(MAX3)/D17 and MAX4/D10 in Arabidopsis and rice,respectively.Then carlactone is converted to noncanonical SLs through MAX1 and LATERAL BRANCHING OXIDOREDUCTASE(LBO)in Arabidopsis,whereas it is converted to canonical SLs by the MAX1 homologs Cytochrome P450 family 711A2(CYP711A2)and CYP711A3 in rice.SLs are catabolized to a butenolide and a residual moiety by CARBOXYLESTERASE 15(CXE15)in Arabidopsis.(B)SL signal transduction components in Arabidopsis and rice.When SLs bind to their receptors AtD14/D14(Arabidopsis/rice),the receptors conformationally change their structure and become active forms.The activated receptors then recruit MAX2/D3(Arabidopsis/rice),an F-box protein,and form the S-phase kinase-associated protein 1(SKP1)-Cullin 1(CUL1)-F-box protein(SCF)-MAX2/D3 E3 complex,which is involved in degradation of the suppressor proteins of SL signaling,namely the SUPPRESSOR of MAX2 1 LIKE 2,6,7 and 8(SMXL2,6,7 and 8)/D53 proteins,through polyubiquitination in Arabidopsis/rice.This leads to the release of transcriptional repression of unknown transcription factors(TFs),inducing the expression of SLresponsive genes.

In both Arabidopsis and rice,the SL signaling transduction processes are initiated by the binding and activation of AtD14/D14(Arabidopsis/rice),anα/β-hydrolase receptor that possesses both receptor and hydrolase functions(Fig.2B)[32,33].The interaction of AtD14/D14 with SLs triggers a conformational change in AtD14/D14,which is suitable for AtD14/D14 interaction with MAX2/D3(Arabidopsis/rice),an F-box protein,forming the Skp1-Cullin-Fbox(SCF)MAX2-E3 ubiquitin ligase complex in both Arabidopsis and rice.This D14-SCFMAX2-E3 complex then tags the class I Clp-ATPase suppressor,SUPPRESSOR OF MAX2-LIKE 2,6,7 and 8(SMXL2,6,7 and 8)in Arabidopsis and D53 in rice,resulting in degradation via the polyubiquitin/proteosome pathway[34-38].D53 of rice,and SMXL6,7,and 8 of Arabidopsis have been proposed[34-37]to be transcriptional repressors that function by interacting with TOPLESS(TPL)and TOPLESS-related(TPR)proteins that are transcriptional co-repressors,and the interaction of SMXL6,7,and 8 with TPR2 increased their transcriptional repression activity and also supported their repressor function[37].A recent investigation[38]revealed that SMXL6 functions as a transcription factor(TF),with transcriptional repression activity,by directly repressing the transcription of the SMXL6,7,and 8 genes.SMXL6 may also form other complexes with unknown TFs to repress downstream genes[BRANCHED 1(BRC1)and PRODUCTION OF ANTHOCYANIN PIGMENT 1(PAP1)][22].Degradation of SMXL6,7,8/D53 removes the suppression of downstream genes,including BRC1 and PAP1,that are involved in modulating plant growth and development and in stress responses[22,34,35,37-41].

3.Karrikin(KAR)signal transduction

KARs are smoke-derived butanolide compounds,containing the same butanolide ring as the D ring of SLs,that have been shown to promote seed germination and inhibit hypocotyl elongation[42].The commercially available racemic mixture of GR24 contains two enantiomers with the stereochemical configurations GR245DSand GR24ent-5DS,which mimic the functions of SL and KAR respectively[43].The components of KAR signaling are similar to those of SL signaling[42,44].KAR INSENSITIVE 2(KAI2),a homolog of D14 in Arabidopsis,is thought to be a receptor of KARs and a stillunknown KAI2 ligand(KL),and can interact with MAX2 to form a KAI2-SCFMAX2complex after receiving KARs/KL(Fig.3)[35,42,44,45].In Arabidopsis,similar to the D14-SCFMAX2complex,the KAI2-SCFMAX2complex targets the degradation of SMAX1 and SMXL2 proteins(Fig.3)[45].This degradation is required for the inhibition of hypocotyl elongation,while the D14-SCFMAX2complex-mediated degradation of SMXL6,7,and 8 is required for inhibition of shoot branching[36,37,41,46,47].In Arabidopsis recent studies[43,47]have investigated the role of SMXL2 in inhibition of hypocotyl elongation using GR244DOand GR24ent-5DSseparated from rac-GR24 as SL and KAR mimics,respectively,and these studies showed that SMXL2 could also be degraded through the D14-SCFMAX2complex.These lines of genetic and evolutionary evidence suggest that KAR signaling and SL signaling share the same(e.g.,MAX2 and SMXL2)or at least similar perception and signaling components,and that there is cross-regulation between KAR and SL signaling.

Fig.3.Schematic model for the crosstalk between strigolactone(SL)signaling and karrikin(KAR)signaling in regulation of root growth and development in Arabidopsis thaliana under normal growth conditions.In the presence of KARs and/or SLs,the perception of KARs and SLs is achieved via KAR INSENSITIVITY 2(KAI2)and D14 receptors,respectively.The activated receptor then recruits MORE AXILLARY GROWTH 2(MAX2)and forms the S-phase kinase-associated protein 1(SKP1)-Cullin 1(CUL1)-F-box protein(SCF)-MAX2 E3 complex,which is involved in degradation of the common suppressor proteins of KAR signaling and SL signaling,namely the SUPPRESSOR of MAX2 1(SMAX1)and SMAX1-LIKE 2(SMXL2)proteins,via polyubiquitination in Arabidopsis,resulting in expression of target genes.Activation of KAI2 and/or SL signaling results in promotion of root hair(RH)growth,primary root(PR)elongation,and root diameter thickening,but inhibition of lateral root(LR)density,and root skewing and waving.Black arrows indicate positive regulation and cross bars indicate negative regulation.Thin arrow from SL signaling to SMAX1 indicates a weaker relationship.Dotted lines indicate possible regulatory relationships.

4.The diverse roles of SLs in regulating plant root architectur e

SLs are biosynthesized mainly in roots,and their biosynthesis is induced when plants are grown under the conditions of either nitrogen(N)or phosphate(Pi)deficiency[11].Increased SL levels in roots and root exudates promote arbuscular mycorrhizal fungal hyphal branching and root colonization,allowing plants to obtain more nutrients and water from the surrounding environment[11,48,49].SLs and subsequent signaling events also affect root growth and development,including PR elongation,AR development,LR initiation and growth,and RH formation and elongation in various plant species(Tables 1,2),which together affect nutrient and water uptake from the soil[24,50-58].For this reason,identification and dissection of mechanisms used by SLs to modulate root growth and development via nutrient and water uptake have received much attention from the research community.We will first focus on the function of SLs and their signaling in modulating root architecture(Tables 1,2).The roles of SLs in regulation of root growth and development are inconsistent,and are dependent upon the growth conditions(such sugar,N and Pi levels in growth media)and plant species(Tables 1,2).Regulatory mechanisms appear to be complex and affected by interactions with the levels,transport,and signal transduction pathways of plant hormones such as auxin,cytokinin(CK)and ethylene.The interactions of SLs with these plant hormones also appear to be species-specific.Below,we discuss possible mechanisms associated with the interactions between SLs and other plant hormones.

4.1.SLs modulate PR and SR elongation in plants

SL promote the elongation of PRs and SRs in Arabidopsis and rice,respectively,under various growth conditions(Fig.4;Tables 1,2)[52,56,59-62].The PRs of Arabidopsis max1,max3,max4 and d14 mutants are shorter than those found in wild-type(WT)plants[62],and the short PR phenotype can be rescued in the SL-deficient max4 mutant,by application of GR24,a synthetic analog of SLs[59].Caution should be taken when GR24 is used in exogenous applications,because the commonly used commercial rac-GR24 is a mixture of two enantiomers,GR245DSand GR24ent-5DS,which mimic canonical SL and KAR signaling activities,via AtD14 and KAI2 in Arabidopsis,respectively[43].Combining the results of SL-specific mutant plant phenotypes and GR24 application should be more reliable for interpreting the functions of SLs in relation to a given phenotype.In rice,GR24 positively regulates SR elongation of WT and SL-deficient d10 mutant genotypes,but not in SL-signaling d14 mutants[51,56].The opposite trend was found in L.japonicus plants,where silencing of the MAX3 ortholog induced longer PRs than found in WT plants[24].These results imply that SLs modulate PR and SR elongation differently in different plant species.The reason may be the homeostasis of auxin metabolism and signal transduction[52,56,59-61].

Table 1Functions of strigolactones in regulation of root growth and development in various plant species.

Table 2Root phenotypes of SL-deficient and SL-insensitive mutants.

Fig.4.Schematic model and function of strigolactones(SLs)in regulating taproot and fibrous root systems in Arabidopsis thaliana and rice(Oryza sativa),as representative examples of dicot and grasses plant species,respectively,under normal conditions.SLs promote primary elongation(PR)and root hair(RH)growth,while inhibiting adventitious root(AR)and lateral root(LR)development in dicot plant species such as Arabidopsis.In grass species,such as rice,SLs may promote RH growth,while inhibiting LR and secondary LR formation.Unlike in Arabidopsis,SLs may promote AR(also called crown root,CR)formation and elongation in rice.Black arrows indicate positive regulation,while crossed bars indicate negative modulation of root growth and development.Red arrows indicate the names of root types.Seminal root(SR)is the root originating from germinated seeds of grass species.CR in rice is similar to AR in Arabidopsis,which is postembryonic shoot borne-roots(initiated from shoot-root joint position).The question mark indicates an unknown pathway.

SLs accumulated in the roots of several species under conditions of either N or Pi deficiency[11,56,63].In rice,application of GR24 to SL-biosynthetic mutant plants,but not d3 mutant plants,triggered complete recovery of root length,compared with WT plants,grown under either N-or Pi-deficient conditions[56],suggesting that SL signaling is involved as a positive regulator of root elongation under N and Pi deficiencies.The diverse roles of SLs in root development may be the results of interactions between SL signaling and nutrient responses,and other phytohormones and will be discussed later.

4.2.SLs modulate AR formation and elongation

In Arabidopsis and pea plants,both SL-deficient and SLsignaling mutants have more ARs than do WT plants,showing that endogenous SLs inhibit AR formation[4,55].In rice,Arite et al.[52]did not observe differences in AR numbers when 2-week-old WT was compared with d10 and d14 mutant seedlings;however,the authors found that ARs were shorter in d10 and d14 mutants than in WT plants,suggesting that SLs promote AR elongation[52].The numbers of ARs in 2-to-4-week-old rice d10 mutant plants were lower than in WT plants,and application of GR24 rescued the AR number of d10 seedlings to WT levels,suggesting that SLs promote AR formation at an earlier growth stage[57].Similarly,SL-deficient maize(Zea mays)zmccd8 mutants showed fewer and shorter nodal ARs than WT[53],indicating the positive role of SLs in nodal AR development.These results suggest that SLs inhibit AR formation in eudicot plants,such as Arabidopsis,while increasing both the elongation and the number of ARs in grasses such as rice and maize(Fig.4;Tables 1,2).The opposite biological functions of SLs in AR formation in eudicots versus grasses suggests a complex regulation of AR development by SLs in different plant species that invites further investigation.One possible explanation might be that eudicots and grasses have different root systems,and another could be differences between eudicots and grasses in interactions among the signaling pathways of SLs,auxin and CKs[53].Given that both Pi and N deficiencies induce SL biosynthesis,it would also be interesting to investigate the effects of SLs on AR development under Pi or N deficiency in multiple plant species.

4.3.SLs modulate LR formation in both grass and eudicot plant species

In comparison with Arabidopsis WT plants,higher LR numbers and longer LR lengths were observed in SL-deficient(max1,max3,max4,and d14)mutant plants under normal growth conditions.Mutant smxl6,7,8 plants showed fewer LRs and shorter LR length,suggesting a negative role for SLs in LR formation and growth(Fig.4;Tables 1,2)[36,50,59,62,64,65].Secondary LRs[roots developed from primary LRs(referred as‘‘LR/LRs”in this review)]were also induced in d10 mutant plants,but not in WT rice plants,suggesting that SL signaling might also negatively modulate secondary LR development in rice[66].These findings indicate that SLs negatively regulate LR development in both grasses and eudicot plants under normal growth conditions[36,50,59,62,64-66].

In Arabidopsis,GR24 treatment reduced WT,max1 and max4 LR density under Pi-sufficient conditions[50].However,under Pideficient conditions,GR24 treatment promoted LR development in Arabidopsis WT plants as compared with SL-deficient max1 and max4 mutant plants[50].These results suggest that the dual effects of SLs in the regulation of LR formation in Arabidopsis plants are dependent upon environmental Pi levels,and may depend upon the Pi-dependent auxin status.A later study[56]in rice revealed that although there were no differences between WT and SL-related d10 and d27 mutants in LR density under normal conditions,WT rice plants displayed much lower LR density than d10 and d27 mutant plants under Pi-deficient conditions,and exogenous application of GR24 decreased LR density of SLdeficient d10 and d27 mutants to the level of GR24-untreated WT.Under N-deficient conditions,WT plants showed slightly lower LR densities than d10 mutant plants,and exogenous application of GR24 to the SL-synthetic d10 mutants resulted in a decrease in LR density,similar to that found with WT plants.These results revealed a partially negative role of SLs in LR root development in rice,under Pi-or N-deficient conditions.Thus,in LR development under Pi-deficient conditions,SLs play a positive regulatory role in Arabidopsis,but a negative regulatory role in rice,suggesting a dependency on nutrient condition and plant species.

4.4.Potential effects of SLs on RH elongation and density in eudicot plant species under various growth conditions

Several early studies[36,51,58,59]using exogenous application of GR24,suggested that SLs might be a positive modulator of RH formation and growth in several eudicot plant species,under both nutrient-sufficient and nutrient-deficient conditions.However,a more recent study[62]of mutants revealed no significant differences in RH length and density between WT and Arabidopsis SLspecific d14,max1,max3,and max4 mutants,but lower RH density and shorter RH length in max2 and KAR-specific kai2 mutants under normal conditions.As MAX2 is also involved in KAR signaling[42,44,45],it was proposed that RH formation and elongation are regulated mainly by KAR signaling and that GR24 induces RH formation and elongation through the KAR rather than SL signaling in Arabidopsis under normal growth conditions.Interestingly,in tomato,SL-depleted plants showed slightly shorter RH than WT under normal sufficient-Pi conditions[58].These findings suggest that SL signaling may still play a lesser role than KAR signaling in RH development,perhaps in a species-dependent manner.More studies in plant species,including both grasses and eudicots,are indicated.

Under Pi-deficient conditions,the RH density was lower in max2 and max4 than in WT plants,and exogenous application of 50μmol/L GR24 increased RH density in max4 mutant plants,but not in max2 mutant plants[54,67].Consistently,application of 5μmol L-1GR24 to tomato promoted RH elongation,and SLdepleted tomato plants displayed shorter RH than WT under Pideficient conditions.These results indicated a partially positive role for SLs in regulating RH density in eudicot plant species such as Arabidopsis and tomato under Pi-deficient conditions[54].The effects of SLs on RH development were observed not under normal but under Pi-deficient conditions,suggesting that the SL signaling may be activated by Pi deficiency and the enhanced SL signaling induce RH development under normal growth conditions.Further investigations on RH development are needed using multiple SLdepleted Arabidopsis and tomato mutants,as well as those from other eudicot species,under multiple environmental stress(Pior N-deficient)conditions.Given that SLs differentially regulate the development of other parts of roots such as ARs in eudicots and grasses,it would also be interesting to investigate the roles of SLs in the regulation of RH formation and elongation in rice and other grasses in future studies.

5.Cross-regulation between SLs and other signaling pathways in root growth and development

Root growth and development are also modulated by a variety of signaling pathways,such as auxin[68],CKs[65],ethylene[69],and KAR pathways[62].Some studies[59,65,70-76]supported both direct and indirect cross-regulation between SLs and other phytohormones in the regulation of root development in rice,Arabidopsis,and Lotus japonicus(Fig.5).In the sections below,we discuss the interactions between SLs and some other hormones in the modulation of root growth and development.

5.1.Cross-regulation between SL signaling and auxin signaling

Several studies have supported the interaction between auxin and SLs in the modulation of root growth and development.First,in Arabidopsis,auxin induces SL biosynthesis in the roots by upregulating the expression of SL-biosynthetic genes,such as MAX3 and MAX4(Fig.5A)[77].Second,the effects of SLs in modulating root growth and development in Arabidopsis and rice plants might be explained by their role in inhibiting PIN-mediated polar auxin transport from aerial tissues to roots(Fig.5A)[49-51,54,56,59,74,78,79].As Arabidopsis and rice have taproot and fibrous root systems,respectively(Fig.1),we will discuss these two root systems separately.

In Arabidopsis,SLs may modulate LR formation via regulation of auxin levels(Fig.5A).In the presence of a normal auxin level,SLs repress LR development and auxin-induced expression of the DR5::GUS construct by regulating PIN protein levels and localization[59],such as PIN trafficking and clathrin-mediated PIN endocytosis in Arabidopsis[79].However,in the presence of high levels of auxin(such as exogenously applied),GR24 promotes LR formation and growth[54,59].These results suggested a complex function between SLs(or KARs)and auxin fluxes and homeostasis with respect to LR formation[80].Interestingly,another study[54]showed that Pi deficiency promoted the accumulation of SLs,which induced the expression of the auxin receptor gene TRANSPORT INHIBITOR RESPONSE 1(TIR1)in WT but not in max4 mutant plants,suggesting that Pi deficiency affects RH density via the action of both SL and auxin pathways,with the auxin pathway being downstream of the SL pathway.For AR development,exogenous application of auxin further increased AR numbers in Arabidopsis SL-deficient mutants,and GR24 partially inhibited the promoting effect of auxin on AR development[55],suggesting a cross-regulation between auxin and SLs in AR development in Arabidopsis.These results reveal complex interactions between SL and auxin signals in the modulation of LR[59,80],RH[54,70,80]and AR[55,80]development in Arabidopsis(Fig.5A).

In rice,higher auxin concentrations are found in the roots of d10 mutant plants than in WT plants[56,57].GR24 application promotes SR elongation,and inhibits primary LR formation in WT rice plants by reducing auxin transport from shoots to roots by inhibiting the transcription of PIN1a-b,PIN5a,PIN9 and PIN10a genes[52,56],suggesting the involvement of SLs or KARs in reducing auxin levels in the roots to regulate SR and LR development.Application of endogenous SLs and SL-specific mutant plants(such as d10 and d14 plants)could be used to confirm those findings.However,considering the growth and development of AR,increased auxin concentrations may not be responsible for the increased AR growth in d10 mutants in the short term[57],suggesting that the interactions between auxin and SLs in regulating AR formation differ from those in SR elongation and LR formation(Fig.5A).The interaction between SLs and auxin in RH development in rice invites further study.

Perception of SLs by AtD14/D14,which triggers the degradation of the SMXL2,6,7,and 8/D53 transcriptional repressors,could result in the release of the repression of some of the interacting TFs that participate in modulation of root growth in Arabidopsis and rice[17,22,24,38].In rice,in the absence of SLs,D53 interacts with IDEAL PLANT ARCHITECTURE 1/SQUAMOSA PROMOTER BINDING PROTEIN LIKE 14(IPA1/SPL14,a plant-specific TF)and suppresses its transcriptional activation activity and thereby its downstream genes(Fig.5B)[40,81,82].In a recent study[82],in the presence of SLs,the degradation of the D53 protein released the repression of both SPL14-mediated and its homolog SPL17-mediated up-regulation of OsPIN1b,promoting SR elongation(Fig.5C).These biochemical and genetic results reveal a relationship between SLs and auxin in rice,which affects auxin transport and SR elongation.It will be interesting to determine whether there are such protein interactions between the homologs of D53 and SPL14 in other species,such as SMXL2,6,7,and 8 and AtSPLs in Arabidopsis,and their functions in auxin transport,ultimately in modulation of root growth and development.In summary,increasing genetic,biochemical and physiological lines of evidence support roles for SLs in the regulation of auxin perception and transport(for example,via modulating the expression of PIN genes and PIN polarity and trafficking at the subcellular level),in turn affecting root morphology.

Fig.5.Schematic model for crosstalk of strigolactone(SL)signaling with auxin,cytokinin(CK)and ethylene signaling pathways in regulation of root growth and development in Arabidopsis thaliana or rice(Oryza sativa).(A)Crosstalk between SL and auxin signaling in regulating lateral root(LR),adventitious root(AR)and RH development in Arabidopsis.SLs inhibit the polarization and localization of auxin transporter PINs,affecting auxin levels in various tissues via efflux(reducing auxin levels in some tissues)and influx(increasing auxin levels in other tissues)processes.The local accumulation of auxin in those root tissue areas promotes LR,AR and RH development.SLs also promote the expression of the TRANSPORT INHIBITOR RESPONSE 1(TIR1)gene,which encodes an auxin receptor,leading to increased auxin responsiveness,and subsequent RH development.Accumulation of auxin also promotes SL biosynthesis through up-regulation of MORE AXILLARY GROWTH 3(MAX3)and MAX4 genes.GR24 partially inhibits the promoting effect of auxin on AR development.(B,C)Roles of SQUAMOSA PROMOTER BINDING PROTEIN LIKE 14(SPL14)and SPL17 transcription factors(TFs)in regulating the expression of PIN1b and subsequently seminal root(SR)growth in rice.In the absence of SLs,DWARF53(D53)protein binds to SPL14 and 17 and represses their transcriptional activity,resulting in repression of PIN1b expression,subsequently inhibiting SR root growth in rice(B).In the presence of SLs,D53 is degraded by the proteasome system,which releases the repression of SPL14 and 17,thereby promoting the transcription of PIN1b,subsequently improving SR root growth in rice(C).(D)Crosstalk between SL and CK signaling pathways in rice.The perception of SLs leads to degradation of D53 by the proteasome system,resulting in induced expression of CYTOKININ(CK)OXIDASE/DEHYDROGENASE 9(CKX9)via unknown TFs and subsequently reduced CK levels,which might affect root growth.(E)Crosstalk between SL or KAR signaling and ethylene signaling in regulating RH growth in Arabidopsis.GR24(a synthetic analog of SLs)induces ethylene biosynthesisbiosynthesis by up-regulating the expression of 1-AMINOCYCLOPROPANE-1-CARBOXYLIC ACID(ACC)SYNTHASE(ACS)and ACC OXIDASE(ACO)genes,the key genes in ethylene biosynthesis.Accumulation of ethylene promotes root hair(RH)growth in Arabidopsis.Question marks indicate unknown pathways.Black arrows indicate positive regulation,while crossed bars indicate negative regulation of root growth and development.Dotted arrow indicates possible regulatory relationships in roots.

5.2.Cross-regulation between SL and CK signaling

CKs are negative regulators of LR development in both Aradobipsis and rice,and of PR elongation in Arabidopsis,reducing root meristem size and activity[83].Studies[65,74]have suggested that CKs interact with SLs to modulate root growth and development in Arabidopsis and rice.Mutations in CK signaling components,such as ARABIDOPSIS HISTIDINE KINASE3(AHK3),and ARABIDOPSIS RESPONSE REGULATOR1(ARR1)and ARR12,in Arabidopsis plants rendered LR formation insensitive to GR24 treatments[65],suggesting that SL and/or KAR signaling negatively regulate(s)LR formation by affecting CK signaling.Recently[84],GR24 has been shown to promote CK degradation through its transcriptional activation of CK OXIDASE/DEHYDROGENASE 9(CKX9)in shoots of WT plants,but not in SL-specific d53 mutant shoots of rice,suggesting that SL and CK metabolisms are interlinked(Fig.5D).

5.3.Cross-regulation of SL and/or KAR signaling with ethylene signaling

SLs or KARs have been shown[70]to regulate RH growth via interactions with ethylene in Arabidopsis.Application of GR24 up-regulated the expression of 1-AMINO-CYCLOPROPANE-1-CAR BOXYLATE(ACC)SYNTHASE 2(ACS2)and a subset of ACC OXIDASE(ACO)genes,which are involved in ethylene biosynthesis in Arabidopsis[70,85].Mutant max2 plants showed a lower response to GR24,but their response to ACC(a direct precursor of ethylene biosynthesis),as measured by RH formation,was similar to that of WT plants[70].These results indicate that MAX2 is not required for ethylene response in RH formation.The ethylene signalingdeficient mutant ethylene receptor 1(etr1)did not respond to GR24 in RH development[70],suggesting that ethylene signaling is necessary for GR24-induced RH development.Recently[86],genetic evidence suggested that ethylene might act independently in regulation of AR formation in Arabidopsis.These results implied that the ethylene signaling pathway might act downstream of SL or KAR signaling,and that MAX2-mediated SL signaling or KAR signaling positively modulates ethylene biosynthesis via upregulation of ACS or ACO genes,thereby regulating at least RH development in Arabidopsis(Fig.5E),but did not provide strong or direct evidence for the cross-regulation of SL with ethylene signaling.

In recent studies[87,88],KAR-mediated signaling positively regulated ethylene biosynthesis by increasing the expression of the ACS7 gene in both Lotus japonicus and Arabidopsis,confirming the function of KARs in ethylene biosynthesis,and perhaps RH development through the ethylene pathway and local auxin accumulation[88].The results of an ethylene biosynthesis inhibitor(aminoethoxyvinylglycine)study[89],in which GR24-induced seed germination of parasitic Striga hermonthica plants was blocked,suggests that ethylene biosynthesis might be necessary and occur downstream of SL-induced seed germination,at least in S.hermonthica.

5.4.Cross-regulation between SL signaling and KAR signaling

Recent studies have suggested that KAR and SL signaling function in both common and specific aspects of root growth and development processes in Arabidopsis.Investigations of RH development using SL-specific(d14)and KAR-specific(kai2)signaling mutants revealed that only KAR-specific signaling components,such as KAI2,but not SL-specific signaling components,such as D14,are involved in regulation of RH formation and elongation in Arabidopsis under normal growth conditions(Fig.3).This finding suggests a major role for KAR signaling and that SL signaling plays a lesser role in RH development under Pi-deficient condition[54,67],as previously mentioned in this review.Using the SLspecific and KL-specific signaling mutants,it was found[44,50,55,62,87]that both the SL signaling and KAR signaling negatively regulate LR density,while positively modulating PR elongation.Detailed observation of the LR-related phenotype in Arabidopsis[62]implied that KAR signaling has control over LR development at slightly earlier stage than SL signaling does,and that D14-mediated and KAI2-mediated signaling modulate LR development in an additive manner.Recent reports[62,87]have revealed that KAR signaling negatively regulates root skewing and waving and positively modulates root diameter thickness in Arabidopsis(Fig.3),whereas SL signaling showed no effect on root skewing,straightness,or root diameter.A very recent report[90]indicated that D14-mediated SL signaling also modulated the degradation of SMAX1 and SMXL2 to different extents and affected hypocotyl growth.We hypothesize that SMXL2 is involved in PR elongation and LR development in both SL-signaling and KARsignaling pathway(Fig.3).The functions of SL and KAR signaling in LR development,and root skewing,waving,and diameter phenotypes in other plant species are still unavailable for crossspecies comparison,and would be an interesting topic for future studies.Available evidence[44,62,87,91]suggests that in Arabidopsis only KAR signaling plays a negative role in regulating root skewing and waving,and a positive role in root diameter and RH development,whereas both SL and KAR signaling have negative regulatory functions in PR elongation and positive regulatory functions in LR density(Fig.3).

6.Manipulation of SL signaling to design root architecture and improve soil nutrient utilization in crop plants

Rapid and uniform seedling establishment is a critical phase of crop growth in natural habitats,and affects the growth and yield of crop plants under abiotic conditions such as drought stress[92].Grass crops such as rice and maize possess fibrous root systems.After seed germination,the SR is the main organ of rice seedling for absorbing nutrients and water,and SR elongation induced by SLs is beneficial to seedling establishment and growth under N-and Pi-deficiency conditions[56].Exogenous application of SLs or enhancement of SL signaling pathway by manipulation of SL-related genes can induce well-developed SR in seedlings,and guarantee rapid and uniform seedling establishment and yield of crops under field conditions[93].At later plant growth and developmental stages,AR and CR are crucial organs for the uptake of nutrients and the development of the fibrous root system of rice plants.The elongation of AR and CR by elevated endogenous SLs under N-or Pi-deficiency conditions increased nutrient uptake in rice plants[52,56].These results implied that exogenous application of SLs or enhancement of SL signaling pathway could be used to increase the nutrient absorption capacity of crop plants by increasing AR/CR length,and accordingly could be considered candidates for selection in future breeding programs.

Similar results were also observed for eudicot plants such as Arabidopsis,where SLs and SL signaling promoted PR elongation[59,61,62].Exogenous application of SLs or enhancement of SL signaling pathway could potentially increase water and nutrient absorption from surface soil during early seedling establishment and deep soil during later development stages of eudicot crop plants.Long RH and high RH density would also increase water and nutrient(N and Pi)absorption from the soil,suggesting that SL signaling can promote water,N,and Pi uptake in plants by promoting RH development[59,61].In summary,manipulation of SL signaling could be used to increase crop resistance to drought and nutrient deficiency via modulation of root morphology.

Several ways can be used to enhance SL signaling pathway,such as exogenous application of endogenous SLs and their analogs(such as GR24)has been widely used in laboratory experiments.However,the high cost and low stability of these chemicals has limited their use on crop plants in the field.Investigation of stable SL mimics or analogs,such as 4-Br-debranone and 2-NO2-debranone,will facilitate SL research in agriculture[94,95].Overexpression of MAX3 and MAX4 homologs,which are key SL biosynthesis genes involved in feedback regulation[33,96],might be used to increase endogenous levels of SLs or to knock out or downregulate SMXL6,7,and 8 genes to reduce the suppression of SL signaling in crop plants[36,37].

7.Concluding and perspective remarks

SLs are synthesized mainly in roots,and their levels become elevated when either N or Pi is lacking in the soil.Under normal conditions,SLs induce the elongation of PR and RH in Arabidopsis and SR in rice,and suppress LR development in Arabidopsis,pea,and rice.The effects of SLs on formation and elongation of AR are either positive or negative,depending on the plant species and growth period.The observation that some downstream transcription factors and target genes of the SL-signaling pathway are also involved in other hormone signaling pathways supports a hypothesis of cross-regulation between SL and other hormone signaling in root growth and development.Some early experiments aimed at characterizing SL function by exogenous application of the mixture rac-GR24,which mimics both SLs and KARs and initiates their signal transduction,remind us to consider both possible pathways in those studies for more accurate interpretation.Investigations of SL-or KAR-specific mutant plants in future studies would provide more accurate results than exogenous applications of rac-GR24 alone.

CRediT authorship contribution statement

Huwei Sun:Formal analysis,Visualization,Writing-original draft.Weiqiang Li:Formal analysis,Visualization,Writing-original draft.David J.Burritt:Conceptualization,Writing-review &editing.Hongtao Tian:Conceptualization,Writing-review&editing.Heng Zhang:Conceptualization,Writing-review & editing.Xiaohan Liang:Conceptualization,Writing-review & editing.Yuchen Miao:Conceptualization,Writing-review & editing.Mohammad Golam Mostofa:Conceptualization,Writing-review& editing.Lam-Son Phan Tran:Conceptualization,Writingreview & editing.

Declaration of competing interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgments

This work was funded by the National Natural Science Foundation of China(31601821 and 31770300),the Strategic Priority Research Program of the Chinese Academy of Sciences(XDA28110100),the National Key Research and Development Program of China (2018YFE0194000,2018YFD0100304,2016YFD0101006),and the Special Fund for Henan Agriculture Research System(HARS-22-03-G3).