Auxin Transport Inhibitor Research Articles

Tobacco roots increasing diameter and secondary lateral density in response to drought stress

Exploring the responses of root morphology and its physiological mechanisms under drought stress is significant for further improving water and nutrient absorption in roots. Here, we simulated drought through hydroponics combined with PEG treatments in tobacco to characterize the changes in tobacco root architecture. Our results showed the total root length, first lateral root number, and first lateral root length were significantly reduced upon increasing drought severity, but the average root diameter and secondary lateral root density increased under certain drought conditions. The change of auxin content in roots under drought stress was correlated with the root diameter and second lateral root density responses. Exogenous addition of the auxin analog (NAA) and the auxin transport inhibitor (NPA), as well as DR5:GUS staining experiments further demonstrated that auxin participated in this physiological process. Meanwhile, brassinolide (BR) exhibited a similar trend. Exogenous addition of BR (EBR) and the BR synthesis inhibitor BRZ experiments demonstrated that BR may participate upstream of auxin under drought stress. PEG treatment significantly up-regulated NtBRI1 at 9–24 h, and promoted the up-regulation of NtBSK2 and NtBSK3 at 48 h and 24 h, respectively, these genes may contribute to the change in root morphology under drought stress. This study shows that auxin and BR are involved in the changes in root morphology in tobacco exposed to drought stress. The elucidation of the molecular mechanism at play thus represents a future target for breeding drought-tolerant tobacco varieties.

Flavonols affect the interrelated glucosinolate and camalexin biosynthetic pathways in Arabidopsis thaliana.

Flavonols are structurally and functionally diverse biomolecules involved in plant biotic and abiotic stress tolerance, pollen development, and inhibition of auxin transport. However, their effects on global gene expression and signaling pathways are unclear. To explore the roles of flavonol metabolites in signaling, we performed comparative transcriptome and targeted metabolite profiling of seedlings from the flavonol-deficient Arabidopsis loss-of-function mutant flavonol synthase1 (fls1) with and without exogenous supplementation of flavonol derivatives (kaempferol, quercetin, and rutin). RNA-seq results indicated that flavonols modulate various biological and metabolic pathways, with significant alterations in camalexin and aliphatic glucosinolate synthesis. Flavonols negatively regulated camalexin biosynthesis but appeared to promote the accumulation of aliphatic glucosinolates via transcription factor-mediated up-regulation of biosynthesis genes. Interestingly, upstream amino acid biosynthesis genes involved in methionine and tryptophan synthesis were altered under flavonol deficiency and exogenous supplementation. Quercetin treatment significantly up-regulated aliphatic glucosinolate biosynthesis genes compared with kaempferol and rutin. In addition, expression and metabolite analysis of the transparent testa7 mutant, which lacks hydroxylated flavonol derivatives, clarified the role of quercetin in the glucosinolate biosynthesis pathway. This study elucidates the molecular mechanisms by which flavonols interfere with signaling pathways, their molecular targets, and the multiple biological activities of flavonols in plants.

LIPID TRANSFER PROTEIN4 regulates cotton ceramide content and activates fiber cell elongation.

Cell elongation is a fundamental process for plant growth and development. Studies have shown lipid metabolism plays important role in cell elongation; however, the related functional mechanisms remain largely unknown. Here, we report that cotton (Gossypium hirsutum) LIPID TRANSFER PROTEIN4 (GhLTP4) promotes fiber cell elongation via elevating ceramides (Cers) content and activating auxin-responsive pathways. GhLTP4 was preferentially expressed in elongating fibers. Over-expression and down-regulation of GhLTP4 led to longer and shorter fiber cells, respectively. Cers were greatly enriched in GhLTP4-overexpressing lines and decreased dramatically in GhLTP4 down-regulating lines. Moreover, auxin content and transcript levels of indole-3-acetic acid (IAA)-responsive genes were significantly increased in GhLTP4-overexpressing cotton fibers. Exogenous application of Cers promoted fiber elongation, while NPA (N-1-naphthalic acid, a polar auxin transport inhibitor) counteracted the promoting effect, suggesting that IAA functions downstream of Cers in regulating fiber elongation. Furthermore, we identified a basic helix-loop-helix transcription factor, GhbHLH105, that binds to the E-box element in the GhLTP4 promoter region and promotes the expression of GhLTP4. Suppression of GhbHLH105 in cotton reduced the transcripts level of GhLTP4, resulting in smaller cotton bolls and decreased fiber length. These results provide insights into the complex interactions between lipids and auxin-signaling pathways to promote plant cell elongation.

Comparative transcriptome analysis reveals the regulatory effects of exogenous auxin on lateral root development and tanshinone accumulation in Salvia miltiorrhiza.

The physiological and transcriptome analysis revealed that auxin was a positive regulator of lateral root development and tanshinone accumulation in Salvia miltiorrhiza. Roots of S. miltiorrhiza are widely used as medicinal materials in China, and the root morphology and content of bioactive compounds [such as phenolic acids and diterpenoid quinones (tanshinones)] are the main factors to determine the quality of this herb. Auxin regulates root development and secondary metabolism in many plant species, but little is known about its function in S. miltiorrhiza. In this study, S. miltiorrhiza seedlings were treated (exogenous application) with the auxin indole-3-acetic acid (IAA) and the polar auxin transport inhibitor N-1-naphthylphthalamic acid (NPA) to investigate the regulatory roles of auxin in S. miltiorrhiza. The results indicated that exogenous IAA promoted both lateral root development and tanshinones biosynthesis in S. miltiorrhiza. The NPA application suppressed the lateral root development but showed no obvious effects on tanshinones accumulation. Based on the RNA-seq analysis, expressions of genes related to auxin biosynthesis and signaling transduction were altered in both treated groups. Coincidental with the enhanced content of tanshinones, transcripts of several key enzyme genes in the tanshinones biosynthetic pathway were stimulated after the exogenous IAA application. The expression profiles of seven common transcription factor domain-containing gene families were analyzed, and the results implied that some AP2/ERF genes were probably responsible for the auxin-induced lateral root development in S. miltiorrhiza. These findings shed new light on the regulatory roles of auxin on root development and bioactive compounds biosynthesis in S. miltiorrhiza, and lay the groundwork for future research into the detailed molecular mechanism underlying these biological functions.

Tomato miR156-targeted SlSBP15 represses shoot branching by modulating hormone dynamics and interacting with GOBLET and BRANCHED1b.

The microRNA156 (miR156)/SQUAMOSA PROMOTER-BINDING PROTEIN-LIKE (SPL/SBP) regulatory hub is highly conserved among phylogenetically distinct species, but how it interconnects multiple pathways to converge to common integrators controlling shoot architecture is still unclear. Here, we demonstrated that the miR156/SlSBP15 node modulates tomato shoot branching (SB) by connecting multiple phytohormones with classical genetic pathways regulating both axillary bud (AB) development and outgrowth. MiR156-overexpressing plants (156-OE) displayed high SB, whereas plants overexpressing a miR156-resistant SlSBP15 allele (rSBP15) showed arrested SB. Importantly, the rSBP15 allele was able to partially restore the wild-type SB phenotype in 156-OE background. rSBP15 plants have tiny ABs, and their activation is dependent on shoot apex-derived auxin transport inhibition. Hormonal measurements revealed that Indole-3-acetic acid (IAA) and abscisic acid (ABA) concentrations were lower in 156-OE and higher in rSBP15 ABs, respectively. Genetic and molecular data indicated that SlSBP15 regulates AB development and outgrowth by inhibiting auxin transport and GOBLET (GOB) activity, and by interacting with tomato BRANCHED1b (SlBRC1b) to control ABA levels within ABs. Collectively, our data provide a new mechanism by which the miR156/SPL/SBP hub regulates SB, and suggest that modulating SlSBP15 activity might have potential applications in shaping tomato shoot architecture.

Chemical inhibition of Arabidopsis PIN-FORMED auxin transporters by the anti-inflammatory drug naproxen

The phytohormone auxin plays central roles in many growth and developmental processes in plants. Development of chemical tools targeting the auxin pathway is useful for both plant biology and agriculture. Here we reveal that naproxen, a synthetic compound with anti-inflammatory activity in humans, acts as an auxin transport inhibitor targeting PIN-FORMED (PIN) transporters in plants. Physiological experiments indicate that exogenous naproxen treatment affects pleiotropic auxin-regulated developmental processes. Additional cellular and biochemical evidence indicates that naproxen suppresses auxin transport, specifically PIN-mediated auxin efflux. Moreover, biochemical and structural analyses confirm that naproxen binds directly to PIN1 protein via the same binding cavity as the indole-3-acetic acid substrate. Thus, by combining cellular, biochemical, and structural approaches, this study clearly establishes that naproxen is a PIN inhibitor and elucidates the underlying mechanisms. Further use of this compound may advance our understanding of the molecular mechanisms of PIN-mediated auxin transport and expand our toolkit in auxin biology and agriculture.

The cellulose–lignin balance mediated by auxin signal transduction affects the twisted growth of Yunnan pine trunk

Trunk twisting, a special phenomenon observed during plant growth, leads to remarkable changes in plant phenotypic plasticity and adaptability. However, it is a debate whether the trunk twisting belongs to environmental alteration or genetic variation. To understand the mechanisms underlying trunk twisting in Yunnan pine (Pinus yunnanensis Franch.), through a diallel cross experiment, we first determined that trunk twisting was controlled by recessive genes. Anatomical analysis identified that straight and twisty types differed significantly in xylem and phloem. RNA-seq of materials enriched by laser microdissection revealed several metabolic pathways with significant enrichment in twisty pines, including auxin signal transduction, photosynthetic carbon fixation and sucrose metabolism, etc. Application of exogenous auxin and auxin transport inhibitors (TIBA) can only change the growth rate, but cannot change the twisted or straight of the trunk. When auxin signaling inhibitors (auxinole) were added, the straight pines produce a tendency to twisty growth. Combined with enzyme activity assay and immunohistochemistry, we propose a working model. ARF can not only downregulate POR to block chlorophyll synthesis and photosynthesis but also upregulated Susy expression, and allows a large amount of sucrose to synthesize cellulose. Nevertheless, due to downregulated CBH expression and abnormal cellulolysis, cellulose accumulates and the lignin content decreases, eventually making the trunk highly prone to twisted growth. These results reveal the molecular mechanism of trunk twisting, and suggest that ARF expression method can be vital in trunk shape screening during the early growth stages of Yunnan pines.

Design and synthesis of strong root gravitropism inhibitors with no concomitant growth inhibition

Herein, we describe a highly potent gravitropic bending inhibitor with no concomitant growth inhibition. Previously, we reported that (2Z,4E)-5-phenylpenta-2,4-dienoic acid (ku-76) selectively inhibits root gravitropic bending of lettuce radicles at 5 μM. Based on the structure–activity relationship study of ku-76 as a lead compound, we designed and synthesized various C4-substituted analogs of ku-76. Among the analogs, 4-phenylethynyl analog exhibited the highest potency for gravitropic bending inhibition, which was effective at only 0.01 μM. Remarkably, 4-phenylethynyl analog is much more potent than the known inhibitor, NPA. Substitution in the para position on the aromatic ring of 4-phenylethynyl group was tolerated without diminished activity. In addition, evaluation using Arabidopsis indicated that 4-phenylethynyl analog inhibits gravitropism by affecting auxin distribution in the root tips. Based on the effects on Arabidopsis phenotypes, 4-phenylethynyl analog may be a novel inhibitor that differs in action from the previously reported auxin transport inhibitors.

Auxin participates in regulating the leaf curl development of Wucai (Brassica campestris L.).

Wucai (Brassica campestris L. ssp. chinensis var. rosularis Tsen) belongs to the Brassica genus of the Cruciferae family, and its leaf curl is a typical feature that distinguishes Wucai from other nonheading cabbage subspecies. Our previous research found that plant hormones were involved in the development of the leaf curl in Wucai. However, the molecular mechanisms and the hormones regulating the formation of leaf curl in Wucai have not yet been reported. This study aimed to understand the molecular functions related to hormone metabolism during the formation of leaf curl in Wucai. A total of 386 differentially expressed genes (DEGs) were identified by transcriptome sequencing of two different morphological parts of the same leaf of Wucai germplasm W7-2, and 50 DEGs were found to be related to plant hormones, which were mainly involved in the auxin signal transduction pathway. Then, we measured the content of endogenous hormones in two different forms of the same leaf of Wucai germplasm W7-2. A total of 17 hormones with differential content were identified, including auxin, cytokinins, jasmonic acids, salicylic acids, and abscisic acid. And we found that treatment with auxin transport inhibitor N-1-naphthylphthalamic acid can affect the leaf curl phenotype of Wucai and pak choi (Brassica rapa L. subsp. Chinensis). These results indicated that plant hormones, especially auxin, are involved in developing the leaf curl of Wucai. Our findings provide a potentially valuable reference for future research on the development of leaf curls.

Identification of new potential downstream transcriptional targets of the strigolactone pathway including glucosinolate biosynthesis.

Strigolactones regulate shoot branching and many aspects of plant growth, development, and allelopathy. Strigolactones are often discussed alongside auxin because they work together to inhibit shoot branching. However, the roles and mechanisms of strigolactones and how they act independently of auxin are still elusive. Additionally, there is still much in general to be discovered about the network of molecular regulators and their interactions in response to strigolactones. Here, we conducted an experiment in Arabidopsis with physiological treatments and strigolactone mutants to determine transcriptional pathways associated with strigolactones. The three physiological treatments included shoot tip removal with and without auxin treatment and treatment of intact plants with the auxin transport inhibitor, N-1-naphthylphthalamic acid (NPA). We identified the glucosinolate biosynthesis pathway as being upregulated across strigolactone mutants indicating strigolactone-glucosinolate crosstalk. Additionally, strigolactone application cannot restore the highly branched phenotype observed in glucosinolate biosynthesis mutants, placing glucosinolate biosynthesis downstream of strigolactone biosynthesis. Oxidative stress genes were enriched across the experiment suggesting that this process is mediated through multiple hormones. Here, we also provide evidence supporting non-auxin-mediated, negative feedback on strigolactone biosynthesis. Increases in strigolactone biosynthesis gene expression seen in strigolactone mutants could not be fully restored by auxin. By contrast, auxin could fully restore auxin-responsive gene expression increases, but not sugar signaling-related gene expression. Our data also point to alternative roles of the strigolactone biosynthesis genes and potential new signaling functions of strigolactone precursors. In this study, we identify a strigolactone-specific regulation of glucosinolate biosynthesis genes indicating that the two are linked and may work together in regulating stress and shoot ranching responses in Arabidopsis. Additionally, we provide evidence for non-auxinmediated feedback on strigolactone biosynthesis and discuss this in the context of sugar signaling.

Auxin efflux carrier ZmPIN1a modulates auxin reallocation involved in nitrate-mediated root formation

BackgroundAuxin plays a crucial role in nitrate (NO3–)-mediated root architecture, and it is still unclear that if NO3– supply modulates auxin reallocation for regulating root formation in maize (Zea mays L.). This study was conducted to investigate the role of auxin efflux carrier ZmPIN1a in the root formation in response to NO3– supply.ResultsLow NO3– (LN) promoted primary root (PR) elongation, while repressed the development of lateral root primordia (LRP) and total root length. LN modulated auxin levels and polar transport and regulated the expression of auxin-responsive and -signaling genes in roots. Moreover, LN up-regulated the expression level of ZmPIN1a, and overexpression of ZmPIN1a enhanced IAA efflux and accumulation in PR tip, while repressed IAA accumulation in LRP initiation zone, which consequently induced LN-mediated PR elongation and LR inhibition. The inhibition rate of PR length, LRP density and number of ZmPIN1a-OE plants was higher than that of wild-type plants after auxin transport inhibitor NPA treatment under NN and LN conditions, and the degree of inhibition of root growth in ZmPIN1a-OE plants was more obvious under LN condition.ConclusionThese findings suggest that ZmPIN1a was involved in modulating auxin levels and transport to alter NO3–-mediated root formation in maize.

Tomato (Solanum lycopersicum) WRKY23 enhances salt and osmotic stress tolerance by modulating the ethylene and auxin pathways in transgenic Arabidopsis

Osmotic stress is one of the biggest problems in agriculture, which adversely affects crop productivity. Plants adopt several strategies to overcome osmotic stresses that include transcriptional reprogramming and activation of stress responses mediated by different transcription factors and phytohormones. We have identified a WRKY transcription factor from tomato, SlWRKY23, which is induced by mannitol and NaCl treatment. Over-expression of SlWRKY23 in transgenic Arabidopsis enhances osmotic stress tolerance to mannitol and NaCl and affects root growth and lateral root number. Transgenic Arabidopsis over-expressing SlWRKY23 showed reduced electrolyte leakage and higher relative water content than Col-0 plants upon mannitol and NaCl treatment. These lines also showed better membrane integrity with lower MDA content and higher proline content than Col-0. Responses to mannitol were governed by auxin as treatment with TIBA (auxin transport inhibitor) negatively affected the osmotic tolerance in transgenic lines by inhibiting lateral root growth. Similarly, responses to NaCl were controlled by ethylene as treatment with AgNO3 (ethylene perception inhibitor) inhibited the stress response to NaCl by suppressing primary and lateral root growth. The study shows that SlWRKY23, a osmotic stress inducible gene in tomato, imparts tolerance to mannitol and NaCl stress through interaction of the auxin and ethylene pathways.

RAV1 mediates cytokinin signaling for regulating primary root growth in Arabidopsis.

Root growth dynamics is an outcome of complex hormonal crosstalk. The primary root meristem size, for example, is determined by antagonizing actions of cytokinin and auxin. Here we show that RAV1, a member of the AP2/ERF family of transcription factors, mediates cytokinin signaling in roots to regulate meristem size. The rav1 mutants have prominently longer primary roots, with a meristem that is significantly enlarged and contains higher cell numbers, compared with wild-type. The mutant phenotype could be restored on exogenous cytokinin application or by inhibiting auxin transport. At the transcript level, primary cytokinin-responsive genes like ARR1, ARR12 were significantly downregulated in the mutant root, indicating impaired cytokinin signaling. In concurrence, cytokinin induced regulation of SHY2, an Aux/IAA gene, and auxin efflux carrier PIN1 was hindered in rav1, leading to altered auxin transport and distribution. This effectively altered root meristem size in the mutant. Notably, CRF1, another member of the AP2/ERF family implicated in cytokinin signaling, is transcriptionally repressed by RAV1 to promote cytokinin response in roots. Further associating RAV1 with cytokinin signaling, our results demonstrate that cytokinin upregulates RAV1 expression through ARR1, during post-embryonic root development. Regulation of RAV1 expression is a part of secondary cytokinin response that eventually represses CRF1 to augment cytokinin signaling. To conclude, RAV1 functions in a branch pathway downstream to ARR1 that regulates CRF1 expression to enhance cytokinin action during primary root development in Arabidopsis.

МОРФОГЕНЕЗ IN VITRO В КАЛЛУСАХ ЯЧМЕНЯ: ВЛИЯНИЕ НАФТИЛФТАЛАМОВОЙ КИСЛОТЫ

Morphogenesis in vitro in barley calli of the cv. Steptoe and its ABA-deficient mutant AZ34 under the influence of an inhibitor of polar auxin transport, N-1-naphthylphthalamic acid (NPA), was studied by light microscopy The formation of «polymeric» embryoids (polyembryoids), characterized by the presence of multiple, often fused (fasciated) organs in varying degrees, in calli of cv. Steptoe and the formation of fasciated roots in calli of AZ34 were revealed. The distribution of IAA in calli was studied by the immunolocalization method. It was found that the most intense staining for IAA in both genotypes was found in fasciated roots. The important role of the polar transport of auxins in the regulation of plant morphogenesis in vitro has been confirmed.

Position of meristems and the angles of the cell division plane regulate the uniqueness of lateral organ shape.

Leaf meristem is a cell proliferative zone present in the lateral organ primordia. In this study, we examined how cell proliferative zones in primordia of planar floral organs and polar auxin transport inhibitor (PATI)-treated leaf organs differ from those of non-treated foliage leaves of Arabidopsis thaliana, with a focus on the accumulation pattern of ANGUSTIFOLIA3 (AN3) protein, a key element for leaf meristem positioning. We found that PATI-induced leaf shape changes were correlated with cell division angle but not with meristem positioning/size or AN3 localisation. In contrast, different shapes between sepals and petals compared with foliage leaves were associated with both altered meristem position, due to altered AN3 expression patterns, and different distributions of cell division angles. A numerical simulation showed that meristem position majorly affected the final shape but biased cell division angles had a minor effect. Taken together, these results suggest that the unique shapes of different lateral organs depend on the position of the meristem in the case of floral organs and cell division angles in the case of leaf organs with different auxin flow.

Mapping of the Classical Mutation rosette Highlights a Role for Calcium in Wound-Induced Rooting.

Removal of the root system induces the formation of new roots from the remaining shoot. This process is primarily controlled by the phytohormone auxin, which interacts with other signals in a yet unresolved manner. Here, we study the classical tomato mutation rosette (ro), which lacks shoot-borne roots. ro mutants were severely inhibited in formation of wound-induced roots (WiRs) and had reduced auxin transport rates. We mapped ro to the tomato ortholog of the Arabidopsis thaliana BIG and the mammalians UBR4/p600. RO/BIG is a large protein of unknown biochemical function. In A. thaliana, BIG was implicated in regulating auxin transport and calcium homeostasis. We show that exogenous calcium inhibits WiR formation in tomato and A. thaliana ro/big mutants. Exogenous calcium antagonized the root-promoting effects of the auxin indole-3-acetic-acid but not of 2,4-dichlorophenoxyacetic acid, an auxin analog that is not recognized by the polar transport machinery, and accumulation of the auxin transporter PIN-FORMED1 (PIN1) was sensitive to calcium levels in the ro/big mutants. Consistent with a role for calcium in mediating auxin transport, both ro/big mutants and calcium-treated wild-type plants were hypersensitive to treatment with polar auxin transport inhibitors. Subcellular localization of BIG suggests that, like its mammalian ortholog, it is associated with the endoplasmic reticulum. Analysis of subcellular morphology revealed that ro/big mutants exhibited disruption in cytoplasmic streaming. We suggest that RO/BIG maintains auxin flow by stabilizing PIN membrane localization, possibly by attenuating the inhibitory effect of Ca2+ on cytoplasmic streaming.

The Germin-like protein OsGER4 is involved in promoting crown root development under exogenous jasmonic acid treatment in rice.

In rice (Oryza sativa L.), crown roots (CRs) have many important roles in processes such as root system expansion, water and mineral uptake, and adaptation to environmental stresses. Phytohormones such as auxin, cytokinin, and ethylene are known to control CR initiation and development in rice. However, the role of jasmonic acid (JA) in CR development remained elusive. Here, we report that JA promotes CR development by regulating OsGER4, a rice Germin-like protein. Root phenotyping analysis revealed that exogenous JA treatment induced an increase in CR number in a concentration-dependent manner. A subsequent genome-wide association study and gene expression analyses pinpointed a strong association between the Germin-like protein OsGER4 and the increase in CR number under exogenous JA treatment. The ProGER4::GUS reporter line showed that OsGER4 is a hormone-responsive gene involved in various stress responses, mainly confined to epidermal and vascular tissues during CR primordia development and to vascular bundles of mature crown and lateral roots. Notable changes in OsGER4 expression patterns caused by the polar auxin transport inhibitor NPA support its connection to auxin signaling. Phenotyping experiments with OsGER4 knockout mutants confirmed that this gene is required for CR development under exogenous JA treatment. Overall, our results provide important insights into JA-mediated regulation of CR development in rice.

Leaf vein patterning is regulated by the aperture of plasmodesmata intercellular channels.

To form tissue networks, animal cells migrate and interact through proteins protruding from their plasma membranes. Plant cells can do neither, yet plants form vein networks. How plants do so is unclear, but veins are thought to form by the coordinated action of the polar transport and signal transduction of the plant hormone auxin. However, plants inhibited in both pathways still form veins. Patterning of vascular cells into veins is instead prevented in mutants lacking the function of the GNOM (GN) regulator of auxin transport and signaling, suggesting the existence of at least one more GN-dependent vein-patterning pathway. Here we show that in Arabidopsis such a pathway depends on the movement of auxin or an auxin-dependent signal through plasmodesmata (PDs) intercellular channels. PD permeability is high where veins are forming, lowers between veins and nonvascular tissues, but remains high between vein cells. Impaired ability to regulate PD aperture leads to defects in auxin transport and signaling, ultimately leading to vein patterning defects that are enhanced by inhibition of auxin transport or signaling. GN controls PD aperture regulation, and simultaneous inhibition of auxin signaling, auxin transport, and regulated PD aperture phenocopies null gn mutants. Therefore, veins are patterned by the coordinated action of three GN-dependent pathways: auxin signaling, polar auxin transport, and movement of auxin or an auxin-dependent signal through PDs. Such a mechanism of tissue network formation is unprecedented in multicellular organisms.

Division of cortical cells is regulated by auxin in Arabidopsis roots.

The root cortex transports water and nutrients absorbed by the root epidermis into the vasculature and stores substances such as starch, resins, and essential oils. The cortical cells are also deeply involved in determining epidermal cell fate. In Arabidopsis thaliana roots, the cortex is composed of a single cell layer generated by a single round of periclinal division of the cortex/endodermis initials. To further explore cortex development, we traced the development of the cortex by counting cortical cells. Unlike vascular cells, whose number increased during the development of root apical meristem (RAM), the number of cortical cells did not change, indicating that cortical cells do not divide during RAM development. However, auxin-induced cortical cell division, and this finding was confirmed by treatment with the auxin transport inhibitor N-1-naphthylphthalamic acid (NPA) and examining transgenic plants harboring CO2::ΔARF5, in which cortical expression of truncated AUXIN RESPONSE FACTOR5 (ΔARF5) induces auxin responses. NPA-induced cortical auxin accumulation and CO2::ΔARF5-mediated cortical auxin response induced anticlinal and periclinal cell divisions, thus increasing the number of cortical cells. These findings reveal a tight link between auxin and cortical cell division, suggesting that auxin is a key player in determining root cortical cell division.

The Plastidial DIG5 Protein Affects Lateral Root Development by Regulating Flavonoid Biosynthesis and Auxin Transport in Arabidopsis.

To reveal the mechanisms underlying root adaptation to drought stress, we isolated and characterized an Arabidopsis mutant, dig5 (drought inhibition of lateral root growth 5), which exhibited increased sensitivity to the phytohormone abscisic acid (ABA) for the inhibition of lateral root growth. The dig5 mutant also had fewer lateral roots under normal conditions and the aerial parts were yellowish with a lower level of chlorophylls. The mutant seedlings also displayed phenotypes indicative of impaired auxin transport, such as abnormal root curling, leaf venation defects, absence of apical hook formation, and reduced hypocotyl elongation in darkness. Auxin transport assays with [3H]-labeled indole acetic acid (IAA) confirmed that dig5 roots were impaired in polar auxin transport. Map-based cloning and complementation assays indicated that the DIG5 locus encodes a chloroplast-localized tRNA adenosine deaminase arginine (TADA) that is involved in chloroplast protein translation. The levels of flavonoids, which are naturally occurring auxin transport inhibitors in plants, were significantly higher in dig5 roots than in the wild type roots. Further investigation showed that flavonoid biosynthetic genes were upregulated in dig5. Introduction of the flavonoid biosynthetic mutation transparent testa 4 (tt4) into dig5 restored the lateral root growth of dig5. Our study uncovers an important role of DIG5/TADA in retrogradely controlling flavonoid biosynthesis and lateral root development. We suggest that the DIG5-related signaling pathways, triggered likely by drought-induced chlorophyll breakdown and leaf senescence, may potentially help the plants to adapt to drought stress through optimizing the root system architecture.