Auxin Transport Research Articles

The ldp1 Mutation Affects the Expression of Auxin-Related Genes and Enhances SAM Size in Rice.

Panicle type is one of the important factors affecting rice (Oryza sativa L.) yield, and the identification of regulatory genes in panicle development can provide significant insights into the molecular network involved. This study identified a large and dense panicle 1 (ldp1) mutant produced from the Wuyunjing 7 (WYJ7) genotype, which displayed significant relative increases in panicle length, number of primary and secondary branches, number of grains per panicle, grain width, and grain yield per plant. Scanning electron microscopy results showed that the shoot apical meristem (SAM) of ldp1 was relatively larger at the bract stage (BM), with a significantly increased number of primary (PBM) and secondary branch (SBM) meristematic centers, indicating that the ldp1 mutation affects early stages in SAM development Comparative RNA-Seq analysis of meristem tissues from WYJ7 and ldp1 at the BM, PBM, and SBM developmental stages indicated that the number of differentially expressed genes (DEGs) were highest (1407) during the BM stage. Weighted gene coexpression network analysis (WGCNA) revealed that genes in one module (turquoise) are associated with the ldp1 phenotype and highly expressed during the BM stage, suggesting their roles in the identity transition and branch differentiation stages of rice inflorescences. Hub genes involved in auxin synthesis and transport pathways, such as OsAUX1, OsAUX4, and OsSAUR25, were identified. Moreover, GO and KEGG analysis of the DEGs in the turquoise module and the 1407 DEGs in the BM stage revealed that a majority of genes involved in tryptophan metabolism and auxin signaling pathway were differentially expressed between WYJ and ldp1. The genetic analysis indicated that the ldp1 phenotype is controlled by a recessive monogene (LDP1), which was mapped to a region between 16.9 and 18.1 Mb on chromosome seven. This study suggests that the ldp1 mutation may affect the expression of key genes in auxin synthesis and signal transduction, enhance the size of SAM, and thus affect panicle development. This study provides insights into the molecular regulatory network underlying rice panicle morphogenesis and lays an important foundation for further understanding the function and molecular mechanism of LDP1 during panicle development.

Partially knocking out NtPDK1a/1b/1c/1d simultaneously in Nicotiana tabacum using CRISPR/CAS9 technology results in auxin-related developmental defects

The eukaryotic AGC protein kinase subfamily (protein kinase A/ protein kinase G/ protein kinase C-family) is involved in regulating numerous biological processes across kingdoms, including growth and development, and apoptosis. PDK1(3-phosphoinositide-dependent protein kinase 1) is a conserved serine/threonine kinase in eukaryotes, which is both a member of AGC kinase and a major regulator of many other downstream AGC protein kinase family members. Although extensively investigated in model plant Arabidopsis, detailed reports for tobacco PDK1s have been limited. To better understand the functions of PDK1s in tobacco, CRISPR/CAS9 transgenic lines were generated in tetraploid N. tabacum, cv. Samsun (NN) with 5–7 of the 8 copies of 4 homologous PDK1 genes in tobacco genome (NtPDK1a/1b/1c/1d homologs) simultaneously knocked out. Numerous developmental defects were observed in these NtPDK1a/1b/1c/1d CRISPR/CAS9 lines, including cotyledon fusion leaf shrinkage, uneven distribution of leaf veins, convex veins, root growth retardation, and reduced fertility, all of which reminiscence of impaired polar auxin transport. The severity of these defects was correlated with the number of knocked out alleles of NtPDK1a/1b/1c/1d. Consistent with the observation in Arabidopsis, it was found that the polar auxin transport, and not auxin biosynthesis, was significantly compromised in these knockout lines compared with the wild type tobacco plants. The fact that no homozygous plant with all 8 NtPDK1a/1b/1c/1d alleles being knocked out suggested that knocking out 8 alleles of NtPDK1a/1b/1c/1d could be lethal. In conclusion, our results indicated that NtPDK1s are versatile AGC kinases that participate in regulation of tobacco growth and development via modulating polar auxin transport. Our results also indicated that CRISPR/CAS9 technology is a powerful tool in resolving gene redundancy in polyploidy plants.

Ethylene Action Inhibition Improves Adventitious Root Induction in Adult Chestnut Tissues.

Phase change refers to the process of maturation and transition from the juvenile to the adult stage. In response to this shift, certain species like chestnut lose the ability to form adventitious roots, thereby hindering the successful micropropagation of adult plants. While auxin is the main hormone involved in adventitious root formation, other hormones, such as ethylene, are also thought to play a role in its induction and development. In this study, experiments were carried out to determine the effects of ethylene on the induction and growth of adventitious roots. The analysis was performed in two types of chestnut microshoots derived from the same tree, a juvenile-like line with a high rooting ability derived from basal shoots (P2BS) and a line derived from crown branches (P2CR) with low rooting responses. By means of the application of compounds to modify ethylene content or inhibit its signalling, the potential involvement of this hormone in the induction of adventitious roots was analysed. Our results show that ethylene can modify the rooting competence of mature shoots, while the response in juvenile material was barely affected. To further characterise the molecular reasons underlying this maturation-derived shift in behaviour, specific gene expression analyses were developed. The findings suggest that several mechanisms, including ethylene signalling, auxin transport and epigenetic modifications, relate to the modulation of the rooting ability of mature chestnut microshoots and their recalcitrant behaviour.

Rapid translocation of NGR proteins driving polarization of PIN-activating D6 protein kinase during root gravitropism.

Root gravitropic bending represents a fundamental aspect of terrestrial plant physiology. Gravity is perceived by sedimentation of starch-rich plastids (statoliths) to the bottom of the central root cap cells. Following gravity perception, intercellular auxin transport is redirected downwards leading to an asymmetric auxin accumulation at the lower root side causing inhibition of cell expansion, ultimately resulting in downwards bending. How gravity-induced statoliths repositioning is translated into asymmetric auxin distribution remains unclear despite PIN auxin efflux carriers and the Negative Gravitropic Response of roots (NGR) proteins polarize along statolith sedimentation, thus providing a plausible mechanism for auxin flow redirection. In this study, using a functional NGR1-GFP construct, we visualized the NGR1 localization on the statolith surface and plasma membrane (PM) domains in close proximity to the statoliths, correlating with their movements. We determined that NGR1 binding to these PM domains is indispensable for NGR1 functionality and relies on cysteine acylation and adjacent polybasic regions as well as on lipid and sterol PM composition. Detailed timing of the early events following graviperception suggested that both NGR1 repolarization and initial auxin asymmetry precede the visible PIN3 polarization. This discrepancy motivated us to unveil a rapid, NGR-dependent translocation of PIN-activating AGCVIII kinase D6PK towards lower PMs of gravity-perceiving cells, thus providing an attractive model for rapid redirection of auxin fluxes following gravistimulation.

Rapid translocation of NGR proteins driving polarization of PIN-activating D6 protein kinase during root gravitropism

Root gravitropic bending represents a fundamental aspect of terrestrial plant physiology. Gravity is perceived by sedimentation of starch-rich plastids (statoliths) to the bottom of the central root cap cells. Following gravity perception, intercellular auxin transport is redirected downwards leading to an asymmetric auxin accumulation at the lower root side causing inhibition of cell expansion, ultimately resulting in downwards bending. How gravity-induced statoliths repositioning is translated into asymmetric auxin distribution remains unclear despite PIN auxin efflux carriers and the Negative Gravitropic Response of roots (NGR) proteins polarize along statolith sedimentation, thus providing a plausible mechanism for auxin flow redirection. In this study, using a functional NGR1-GFP construct, we visualized the NGR1 localization on the statolith surface and plasma membrane (PM) domains in close proximity to the statoliths, correlating with their movements. We determined that NGR1 binding to these PM domains is indispensable for NGR1 functionality and relies on cysteine acylation and adjacent polybasic regions as well as on lipid and sterol PM composition. Detailed timing of the early events following graviperception suggested that both NGR1 repolarization and initial auxin asymmetry precede the visible PIN3 polarization. This discrepancy motivated us to unveil a rapid, NGR-dependent translocation of PIN-activating AGCVIII kinase D6PK towards lower PMs of gravity-perceiving cells, thus providing an attractive model for rapid redirection of auxin fluxes following gravistimulation.

Expression of auxin transporter genes in flax (Linum usitatissimum) fibers during gravity response.

Gravitropism is an adaptive reaction of plants associated with the ability of various plant organs to be located and to grow in a certain direction relative to the gravity vector, while usually the asymmetric distribution of the phytohormone auxin is a necessary condition for the gravitropical bending of plant organs. Earlier, we described significant morphological changes in phloem fibers with a thickened cell wall located on different sides of the stem in the area of the gravitropic curvature. The present study is the first work devoted to the identification of genes encoding auxin transporters in cells at different stages of development and during gravity response. In this study, the flax genes encoding the AUX1/LAX, PIN-FORMED, PIN-LIKES, and ABCB auxin transporters were identified. A comparative analysis of the expression of these genes in flax phloem fibers at different stages of development revealed increased expression of some of these genes at the stage of intrusive growth (LusLAX2 (A, B), LuxPIN1-D, LusPILS7 (C, D)), at the early stage of tertiary cell wall formation (LusAUX1 (A, D), LusABCB1 (A, B), LusABCB15-A, LusPIN1 (A, B), LusPIN4-A, and LusPIN5-A), and at the late stage of tertiary cell wall development (LusLAX3 (A, B)). It was shown that in the course of gravitropism, the expression of many genes, including those responsible for the influx of auxin in cells (LusAUX1-D), in the studied families increased. Differential expression of auxin transporter genes was revealed during gravity response in fibers located on different sides of the stem (upper (PUL) and lower (OPP)). The difference was observed due to the expression of genes, the products of which are responsible for auxin intracellular transport (LusPILS3, LusPILS7-A) and its efflux (LusABCB15-B, LusABCB19-B). It was noted that the increased expression of PIN genes and ABCB genes was more typical of fibers on the opposite side. The results obtained allow us to make an assumption about the presence of differential auxin content in the fibers of different sides of gravistimulated flax plants, which may be determined by an uneven outflow of auxin. This study gives an idea of auxin carriers in flax and lays the foundation for further studies of their functions in the development of phloem fiber and in gravity response.

SlBEL11 regulates flavonoid biosynthesis, thus fine-tuning auxin efflux to prevent premature fruit drop in tomato.

Auxin regulates flower and fruit abscission, but how developmental signals mediate auxin transport in abscission remains unclear. Here, we reveal the role of the transcription factor BEL1-LIKE HOMEODOMAIN11 (SlBEL11) in regulating auxin transport during abscission in tomato (Solanum lycopersicum). SlBEL11 is highly expressed in the fruit abscission zone, and its expression increases during fruit development. Knockdown of SlBEL11 expression by RNA interference (RNAi) caused premature fruit drop at the breaker (Br) and 3 d post-breaker (Br+3) stages of fruit development. Transcriptome and metabolome analysis of SlBEL11-RNAi lines revealed impaired flavonoid biosynthesis and decreased levels of most flavonoids, especially quercetin, which functions as an auxin transport inhibitor. This suggested that SlBEL11 prevents premature fruit abscission by modulating auxin efflux from fruits, which is crucial for the formation of an auxin response gradient. Indeed, quercetin treatment suppressed premature fruit drop in SlBEL11-RNAi plants. DNA affinity purification sequencing (DAP-seq) analysis indicated that SlBEL11 induced expression of the transcription factor gene SlMYB111 by directly binding to its promoter. Chromatin immunoprecipitation-quantitative polymerase chain reaction and electrophoretic mobility shift assay showed that S. lycopersicum MYELOBLASTOSIS VIRAL ONCOGENE HOMOLOG111 (SlMYB111) induces the expression of the core flavonoid biosynthesis genes SlCHS1, SlCHI, SlF3H, and SlFLS by directly binding to their promoters. Our findings suggest that the SlBEL11-SlMYB111 module modulates flavonoid biosynthesis to fine-tune auxin efflux from fruits and thus maintain an auxin response gradient in the pedicel, thereby preventing premature fruit drop.

Regulation of tissue growth in plants - A mathematical modeling study on shade avoidance response in Arabidopsis hypocotyls.

Plant growth is a plastic phenomenon controlled both by endogenous genetic programs and by environmental cues. The embryonic stem, the hypocotyl, is an ideal model system for the quantitative study of growth due to its relatively simple geometry and cellular organization, and to its essentially unidirectional growth pattern. The hypocotyl of Arabidopsis thaliana has been studied particularly well at the molecular-genetic level and at the cellular level, and it is the model of choice for analysis of the shade avoidance syndrome (SAS), a growth reaction that allows plants to compete with neighboring plants for light. During SAS, hypocotyl growth is controlled primarily by the growth hormone auxin, which stimulates cell expansion without the involvement of cell division. We assessed hypocotyl growth at cellular resolution in Arabidopsis mutants defective in auxin transport and biosynthesis and we designed a mathematical auxin transport model based on known polar and non-polar auxin transporters (ABCB1, ABCB19, and PINs) and on factors that control auxin homeostasis in the hypocotyl. In addition, we introduced into the model biophysical properties of the cell types based on precise cell wall measurements. Our model can generate the observed cellular growth patterns based on auxin distribution along the hypocotyl resulting from production in the cotyledons, transport along the hypocotyl, and general turnover of auxin. These principles, which resemble the features of mathematical models of animal morphogen gradients, allow to generate robust shallow auxin gradients as they are expected to exist in tissues that exhibit quantitative auxin-driven tissue growth, as opposed to the sharp auxin maxima generated by patterning mechanisms in plant development.

Arsenite and arsenate stress differently affect auxin distribution in rice roots and brassinosteroids restore it sustaining root system plasticity

Rice is a worldwide cultivated crop that serves as an important source of food for the human population, but it is also the simplest route for arsenic (As) contamination of the food chain. The As inorganic forms, arsenate [As(V)] and arsenite [As(III)], are the highly toxic As species found in the soil and the most easily absorbed by the roots. The absorption of As(V) prevails in aerobic soils while that of As(III) is favored in anaerobic soils. As(V) is converted to As(III) in the roots, although small amounts of As(V) also remain in the plant organs. The root system is the first target of the action of both As forms. The mechanisms of action of As(V) and As(III) are still widely unknown. Understanding them is essential for selecting rice genotypes with a lower capacity of As uptake and transport to the caryopses, thus improving food safety. Auxin is the phytohormone necessary for the development and plasticity of the root system, and its action is modulated by endogenous/exogenous brassinosteroids (BRs), mainly under stress conditions. The research aim was to deepen the knowledge of the mechanisms triggered by As(III) or As(V) in rice roots with particular attention to the role played by the interaction between auxin transport and BRs. We show that As(III) is the main As species present in rice roots regardless of the As(III) or As(V) forms supplied to the growing medium. Arsenic alters auxin distribution in both adventitious and lateral roots, but strongly in the latter ones. The application of an exogenous BR, the 24-epibrassinolide (eBL), combined with As(III) or As(V) strongly increases the expression of the OsPIN2 and OsAUX1 genes involved in auxin transport, thus contributing to restore the correct auxin distribution altered by As, and mainly by As(III), with higher effects on the LRs. Moreover, eBL increases the antioxidant activity in the roots in the presence of As, but only when combined with As(V).

Nitric oxide acts upstream of indole-3-acetic acid in ameliorating arsenate stress in tomato seedlings

After their discovery, nitric oxide (NO) and indole-3-acetic acid (IAA) have been reported as game-changing cellular messengers for reducing abiotic stresses in plants. But, information regarding their shared signaling in regulating metal stress is still unclear. Herein, we have investigated about the joint role of NO and IAA in mitigation of arsenate [As(V)] toxicity in tomato seedlings. Arsenate being a toxic metalloid increases the NPQ level and cell death while decreasing the biomass accumulation, photosynthetic pigments, chlorophyll a fluorescence, endogenous NO content in tomato seedlings. However, application of IAA or SNP to the As(V) stressed seedlings improved growth together with less accumulation of arsenic and thus, preventing cell death. Interestingly, addition of c-PTIO, {2-(4-carboxyphenyl)-4, 4, 5, 5-tetramethylimidazoline-1-oxyl-3-oxide, a scavenger of NO} and 2, 3, 5-triidobenzoic acid (TIBA, an inhibitor of polar auxin transport) further increased cell death and inhibited activity of GST, leading to As(V) toxicity. However, addition of IAA to SNP and TIBA treated seedlings reversed the effect of TIBA resulting into decreased As(V) toxicity. These findings demonstrate that IAA plays a crucial and advantageous function in NO-mediated reduction of As(V) toxicity in seedlings of tomato. Overall, this study concluded that IAA might be acting as a downstream signal for NO-mediated reduction of As(V) toxicity in tomato seedlings.

Leaf Vein Patterning.

Leaves form veins whose patterns vary from a single vein running the length of the leaf to networks of staggering complexity where huge numbers of veins connect to other veins at both ends. For the longest time, vein formation was thought to be controlled only by the polar, cell-to-cell transport of the plant hormone auxin; recent evidence suggests that is not so. Instead, it turns out that vein patterning features are best accounted for by a combination of polar auxin transport, facilitated auxin diffusion through plasmodesma intercellular channels, and auxin signal transduction-though the latter's precise contribution remains unclear. Equally unclear remain the sites of auxin production during leaf development, on which that vein patterning mechanism ought to depend. Finally, whether that vein patterning mechanism can account for the variety of vein arrangements found in nature remains unknown. Addressing those questions will be the exciting challenge of future research.

Maternal nitric oxide homeostasis impacts female gametophyte development under optimal and stress conditions.

In adverse environments, the number of fertilizable female gametophytes (FGs) in plants is reduced, leading to increased survival of the remaining offspring. How the maternal plant perceives internal growth cues and external stress conditions to alter FG development remains largely unknown. We report that homeostasis of the stress signaling molecule nitric oxide (NO) plays a key role in controlling FG development under both optimal and stress conditions. NO homeostasis is precisely regulated by S-nitrosoglutathione reductase (GSNOR). Prior to fertilization, GSNOR protein is exclusively accumulated in sporophytic tissues and indirectly controls FG development in Arabidopsis (Arabidopsis thaliana). In GSNOR null mutants, NO species accumulated in the degenerating sporophytic nucellus, and auxin efflux into the developing FG was restricted, which inhibited FG development, resulting in reduced fertility. Importantly, restoring GSNOR expression in maternal, but not gametophytic tissues, or increasing auxin efflux substrate significantly increased the proportion of normal FGs and fertility. Furthermore, GSNOR overexpression or added auxin efflux substrate increased fertility under drought and salt stress. These data indicate that NO homeostasis is critical to normal auxin transport and maternal control of FG development, which in turn determine seed yield. Understanding this aspect of fertility control could contribute to mediating yield loss under adverse conditions.

Defying gravity: WEEP promotes negative gravitropism in peach trees by establishing asymmetric auxin gradients.

Trees with weeping shoot architectures are valued for their beauty and are a resource for understanding how plants regulate posture control. The peach (Prunus persica) weeping phenotype, which has elliptical downward arching branches, is caused by a homozygous mutation in the WEEP gene. Little is known about the function of WEEP despite its high conservation throughout Plantae. Here, we present the results of anatomical, biochemical, biomechanical, physiological, and molecular experiments that provide insight into WEEP function. Our data suggest that weeping peach trees do not have defects in branch structure. Rather, transcriptomes from the adaxial (upper) and abaxial (lower) sides of standard and weeping branch shoot tips revealed flipped expression patterns for genes associated with early auxin response, tissue patterning, cell elongation, and tension wood development. This suggests that WEEP promotes polar auxin transport toward the lower side during shoot gravitropic response, leading to cell elongation and tension wood development. In addition, weeping peach trees exhibited steeper root systems and faster lateral root gravitropic response. This suggests that WEEP moderates root gravitropism and is essential to establishing the set-point angle of lateral roots from the gravity vector. Additionally, size exclusion chromatography indicated that WEEP proteins self-oligomerize, like other proteins with sterile alpha motif domains. Collectively, our results from weeping peach provide insight into polar auxin transport mechanisms associated with gravitropism and lateral shoot and root orientation.

Transient efflux inhibition improves plant regeneration by natural auxins.

Plant genome editing and propagation are important tools in crop breeding and production. Both rely heavily on the development of efficient in vitro plant regeneration systems. Two prominent regeneration systems that are widely employed in crop production are somatic embryogenesis (SE) and de novo shoot regeneration. In many of the protocols for SE or shoot regeneration, explants are treated with the synthetic auxin analog 2,4-dichlorophenoxyacetic acid (2,4-D), since natural auxins, such as indole-3-acetic acid (IAA) or 4-chloroindole-3-acetic acid (4-Cl-IAA), are less effective or even fail to induce regeneration. Based on previous reports that 2,4-D, compared to endogenous auxins, is not effectively exported from plant cells, we investigated whether efflux inhibition of endogenous auxins could convert these auxins into efficient inducers of SE in Arabidopsis immature zygotic embryos (IZEs). We show that natural auxins and synthetic analogs thereof become efficient inducers of SE when their efflux is transiently inhibited by co-application of the auxin transport inhibitor naphthylphthalamic acid (NPA). Moreover, IZEs of auxin efflux mutants pin2 or abcb1 abcb19 show enhanced SE efficiency when treated with IAA or efflux-inhibited IAA, confirming that auxin efflux reduces the efficiency of Arabidopsis SE. Importantly, in contrast to the 2,4-D system, where only 50-60% of the embryos converted to seedlings, all SEs induced by transport-inhibited natural auxins converted to seedlings. Efflux-inhibited IAA, like 2,4-D, also efficiently induced SE from carrot suspension cells, whereas IAA alone could not, and efflux-inhibited 4-Cl-IAA significantly improved de novo shoot regeneration in Brassica napus. Our data provides new insights into the action of 2,4-D as an efficient inducer of plant regeneration but also shows that replacing this synthetic auxin for efflux-inhibited natural auxin significantly improves different types of plant regeneration, leading to a more synchronized and homogenous development of the regenerated plants.

Far-red light-enhanced apical dominance stimulates flower and fruit abortion in sweet pepper.

Far-red radiation affects many plant processes, including reproductive organ abortion. Our research aimed to determine the role of apical dominance in far-red light-induced flower and fruit abortion in sweet pepper (Capsicum annuum L.). We conducted several climate room experiments where plants were grown under white- or red-rich LED light, with or without additional far-red light. Additional far-red light enhanced apical dominance: it increased auxin levels in the apices of dominant shoots, and caused a greater difference in internode length and apical auxin levels between dominant and subordinate shoots. Additional far-red light stimulated fruit abortion in intact plants but not in decapitated plants, suggesting a crucial role of shoot apices in this effect. However, reducing basipetal auxin transport in the stems with N-1-naphthylphthalamic acid did not influence far-red light-stimulated fruit abortion, although auxin levels in the stem were largely reduced. Applying the synthetic auxin 1-naphthaleneacetic acid on decapitated apices did not influence fruit abortion. However, applying the auxin biosynthesis inhibitor yucasin to shoot apices reduced fruit abortion regardless of the light conditions, accompanied by slight shoot growth retardation. These findings suggest that the basipetal auxin stream does not mediate far-red light-stimulated fruit abortion. Far-red light-stimulated fruit abortion was associated with reduced sucrose accumulation and lower invertase activities in flowers. We suggest that under additional far-red light conditions, increased auxin levels in shoot apices promote fruit abortion probably through enhanced competition for assimilates between apices and flowers, which limits assimilate import into flowers.

Auxin and hydrogen peroxide (H2O2) interaction differentially regulate seedling growth, Na+/K+ ratio and H2S homeostasis accompanying NaCl stress in etiolated sunflower (Helianthus annuus L. cv. Microgreen) seedling roots and cotyledons

Although H2S has been reported to modulate auxin and H2O2 signaling during abiotic stress, no reports decipher the interactive roles of exogenous auxin and H2O2 on the regulation of endogenous H2S homeostasis. In the present work, auxin (IAA) and hydrogen peroxide (H2O2) interaction appear to be crucial in regulating Na+/K+ homeostasis, recovery in seedling growth and H2S accumulation in NaCl-stressed sunflower seedlings. Interestingly, impairment in endogenous IAA transport (TIBA application) also impacts H2S accumulation in the roots and cotyledons. IAA supplementation is effective in reducing Na+/K+ ratio and electrolytic leakage in roots, which reverses in the presence of TIBA (polar auxin transport inhibitor) application. IAA and H2O2-mediated modulation of H2S content corroborates with l-DES activity in seedling roots and cotyledons. In contrast to cotyledons, roots exhibited a reduction in H2S content in the presence of treatments (IAA and H2O2). H2O2 supplementation seems to partially circumvent the changes caused by the disruption of auxin transport. Thus, we hypothesize that auxin might imply H2O2 as a crucial signaling component in the regulation of H2S levels in sunflower tissues. To sum up, IAA and H2O2 interaction regulates root growth, LR proliferation, Na+/K+ ratio, and H2S metabolism in sunflower seedling roots.

Characterization of OsPIN2 Mutants Reveal Novel Roles for Reactive Oxygen Species in Modulating Not Only Root Gravitropism but Also Hypoxia Tolerance in Rice Seedlings.

Tolerance to submergence-induced hypoxia is an important agronomic trait especially for crops in lowland and flooding-affected areas. Although rice (Oryza sativa) is considered a flood-tolerant crop, only limited cultivars display strong tolerance to prolonged submergence and/or hypoxic stress. Therefore, characterization of hypoxic resistant genes and/or germplasms have important theoretical and practical significance for rice breeding and sustained improvements. Previous investigations have demonstrated that loss-of-function of OsPIN2, a gene encoding an auxin efflux transporter, results in the loss of root gravitropism due to disrupted auxin transport in the root tip. In this study, we revealed a novel connection between OsPIN2 and reactive oxygen species (ROS) in modulating root gravitropism and hypoxia tolerance in rice. It is shown that the OsPIN2 mutant had decreased accumulation of ROS in root tip, due to the downregulation of glycolate oxidase encoding gene OsGOX6, one of the main H2O2 sources. The morphological defects of root including waved rooting and agravitropism in OsPIN2 mutant may be rescued partly by exogenous application of H2O2. The OsPIN2 mutant exhibited increased resistance to ROS toxicity in roots due to treatment with H2O2. Furthermore, it is shown that the OsPIN2 mutant had increased tolerance to hypoxic stress accompanied by lower ROS accumulation in roots, because the hypoxia stress led to over production of ROS in the roots of the wild type but not in that of OsPIN2 mutant. Accordingly, the anoxic resistance-related gene SUB1B showed differential expression in the root of the WT and OsPIN2 mutant in response to hypoxic conditions. Notably, compared with the wild type, the OsPIN2 mutant displayed a different pattern of auxin distribution in the root under hypoxia stress. It was shown that hypoxia stress caused a significant increase in auxin distribution in the root tip of the WT but not in that of the war1 mutant. In summary, these results suggested that OsPIN2 may play a role in regulating ROS accumulation probably via mediating auxin transport and distribution in the root tip, affecting root gravitropism and hypoxic tolerance in rice seedlings. These findings may contribute to the genetic improvement and identification of potential hypoxic tolerant lines in rice.

Evaluating the effects of azelaic acid in the metabolism of Arabidopsis thaliana seedlings through untargeted metabolomics and ionomics approaches.

The present study demonstrates that low concentrations of azelaic acid (AZA) significantly impact the metabolism of Arabidopsis thaliana seedlings, leading to imbalances in numerous minerals and metabolites due to AZA-induced stress. Untargeted metabolomic analyses were conducted on untreated and AZA-treated seedlings at two time points: 7 and 14 days after treatment initiation. The results revealed a general accumulation of sugars (e.g., glucose, mannose, xylose), amino acids (e.g., lysine, GABA, threonine, glutamine), and organic acids (e.g., glutaric acid, shikimic acid, succinic acid) in AZA treated-seedlings, suggesting that AZA triggers stress responses in Arabidopsis. Ionomic analysis revealed that AZA induces phosphorus deficiency, which plants compensate by increasing malate content in the roots. Additionally, AZA treatment induced putrescine accumulation within the root, a metabolic biomarker of potassium deficiency and plant stress. The metabolomic profile showed elevated levels of different specialized metabolites, such as nitrogen- and sulphur-containing compounds, and altered levels of various phytohormones, including jasmonates and brassinosteroids, implicated in plant protection under biotic and/or abiotic stresses. These findings support the hypothesis that AZA's mode of action is associated with an auxin imbalance, suggesting its function as an auxinic herbicide. The observed increases in starch and jasmonates, coupled with the disruptions in potassium homeostasis, are linked to the previously reported alterations in the auxin transport, root architecture and gravitropic root response. Statistical analyses were applied, including Kruskal-Wallis tests for ionomic data, as well as multifactor analysis, Principal Component Analysis, Orthogonal Partial Least Squares-Discriminant Analysis, and enrichment pathway analysis for metabolomic data, ensuring the robustness and validity of these findings.

Splicing defect of StDRO2 intron 1 promotes potato root growth by disturbing auxin transport to adapt to drought stress

The formation of root system architecture (RSA) plays a crucial role in plant growth. OsDRO1 is known to have a function in controlling RSA in rice, however, the role of potato StDRO2, a homolog of rice OsDRO1, in root growth remains unclear. In this study, we obtained potato dro2 mutant lines by Clustered Regularly Interspaced Short Palindromic Repeats-CRISPR-Associated 9 (CRISPR/Cas9)-mediated genome editing system. The mutant lines were generated from a splicing defect of the StDRO2 intron 1, which causes a nonsense mutation in StDRO2. Furthermore, the secondary structure of StDRO2 mRNA analyzed with RNAfold WebServer was altered in the dro2 mutant. Mutation of StDRO2 conveys potato adaptation through changing the RSA via alteration of auxin transport under drought stress. The potato dro2 lines showed higher plant height, longer root length, smaller root growth angle and increased tuber weight than the wild-type. The alteration of RSA was associated with a disturbance of IAA distribution in the dro2 mutant, and the levels of StPIN7 and StPIN10 detected by using real-time PCR were up-regulated in the roots of potato dro2 lines grown under drought stress. Moreover, the microRNAs (miRNAs) PmiREN024536 and PmiREN024486 targeted the StDRO2 gene, and auxin positively and negatively regulated the expression of StDRO2 and the miRNAs PmiREN024536 and PmiREN024486, respectively, in the potato roots. Our data shows that a regulatory network involving auxin, StDRO2, PmiREN024536 and PmiREN024486 can control RSA to convey potato fitness under drought stress.

D6PK plasma membrane polarity requires a repeated CXX(X)P motif and PDK1-dependent phosphorylation

D6 PROTEIN KINASE (D6PK) is a polarly localized plasma-membrane-associated kinase from Arabidopsis thaliana that activates polarly distributed PIN-FORMED auxin transporters. D6PK moves rapidly to and from the plasma membrane, independent of its PIN-FORMED targets. The middle D6PK domain, an insertion between kinase subdomains VII and VIII, is required and sufficient for association and polarity of the D6PK plasma membrane. How D6PK polarity is established and maintained remains to be shown. Here we show that cysteines from repeated middle domain CXX(X)P motifs are S-acylated and required for D6PK membrane association. While D6PK S-acylation is not detectably regulated during intracellular transport, phosphorylation of adjacent serine residues, in part in dependence on the upstream 3-PHOSPHOINOSITIDE-DEPENDENT PROTEIN KINASE, promotes D6PK transport, controls D6PK residence time at the plasma membrane and prevents its lateral diffusion. We thus identify new mechanisms for the regulation of D6PK plasma membrane interaction and polarity.