Effects of phosphorus, indolebutyric acid and naphthylphthalamic acid on the lateral root growth of Poncirus trifoliata

To explore the influence of phosphorus (P), indolebutyric acid (IBA, Auxin) and Naphthylphthalamic acid (NPA, Auxin transport inhibitor) on plant lateral root (LR) formation, Poncirus trifoliata seedlings at two P levels, low P (LP) and control treatment (CK), which was applied with IBA and NPA, and the regulative effects of P level, IBA and NPA on LR formation of trifoliate orange were investigated. The results showed that LP level significantly reduced the plant biomass, LR number and length. NPA significantly decreased the plant biomass, LR number and length, while IBA did not significantly influence these parameters. These data suggested that auxin signaling pathway could be involved in the regulation of P level on LR formation, and the auxin transportation should be the key factor in LR formation of trifoliate orange.

Integrated Transcriptome and Metabolome Analysis Reveals an Essential Role for Auxin in Hypocotyl Elongation during End-of-Day Far-Red Treatment of Cucurbita moschata (Duch. Ex Lam.)

Long, robust hypocotyls are important for facilitating greenhouse transplant production. The use of far-red light at the end of the day (end-of-day far-red, EOD-FR) is known to prompt hypocotyl elongation, but the mechanism of EOD-FR-mediated hypocotyl elongation in pumpkin remains unclear. Here, we found that hypocotyl length, parenchymal cell size in hypocotyls, and plant IAA levels were significantly greater in pumpkin after EOD-FR treatment. This effect was counteracted by the application of the polar auxin transport inhibitor 1-N-naphthylphthalamic acid. Integrated transcriptomic and metabolomic analysis of pumpkin hypocotyls revealed that the expression of auxin-related genes changed significantly after EOD-FR treatment, and the contents of the auxin biosynthetic precursors tryptophan and indole were also significantly higher. Our results show that auxin plays an essential role in EOD-FR-mediated hypocotyl elongation, shed light on the mechanisms of EOD-FR mediated hypocotyl elongation, and provide a theoretical basis for the use of EOD-FR in facility cultivation.

Mode of Action of 1-Naphthylphthalamic Acid in Conspicuous Local Stem Swelling of Succulent Plant, Bryophyllum calycinum: Relevance to the Aspects of Its Histological Observation and Comprehensive Analyses of Plant Hormones

The mode of action of 1-naphthylphthalamic acid (NPA) to induce conspicuous local stem swelling in the area of its application to the growing internode in intact Bryophyllum calycinum was studied based on the aspects of histological observation and comprehensive analyses of plant hormones. Histological analyses revealed that NPA induced an increase in cell size and numerous cell divisions in the cortex and pith, respectively, compared to untreated stem. In the area of NPA application, vascular tissues had significantly wider cambial zones consisting of 5–6 cell layers, whereas phloem and xylem seemed not to be affected. This indicates that stem swelling in the area of NPA application is caused by stimulation of cell division and cell enlargement mainly in the cambial zone, cortex, and pith. Comprehensive analyses of plant hormones revealed that NPA substantially increased endogenous levels of indole-3-acetic acid (IAA) in the swelling area. NPA also increased endogenous levels of cytokinins, jasmonic acid, and its precursor, 12-oxo-phytodienoic acid, but did not increase abscisic acid and gibberellin levels. It was shown, using radiolabeled 14C-IAA, that NPA applied to the middle of internode segments had little effect on polar auxin transport, while 2,3,5-triiodobenzoic acid substantially inhibited it. These results strongly suggest that NPA induces changes in endogenous levels of plant hormones, such as IAA, cytokinins, and jasmonic acid, and their hormonal crosstalk results in a conspicuous local stem swelling. The possible different mode of action of NPA from other polar auxin transport inhibitors in succulent plants is extensively discussed.

Crosstalk between auxin and gibberellin during stalk elongation in flowering Chinese cabbage

Plant growth and development are tightly regulated by phytohormones. However, little is known about the interaction between auxin and gibberellin acid (GA) during flower stalk elongation and how it is directly related to organ formation. Therefore, the effects of indole acetic acid (IAA) and GA3 treatments and their interaction on flower stalk elongation in flowering Chinese cabbage were investigated. The growth of flowering Chinese cabbage is regulated by IAA and GA3, and the opposite results were observed after treatments with uniconazole (GA synthesis inhibitor) and N-1-naphthylphthalamic acid (NPA) (auxin transport inhibitor). Anatomical analysis of the pith region in stalks revealed that IAA promoted expansion via signal transduction and transport pathways. GA3 regulated the elongation of flower stalks by controlling GA synthesis and partially controlling the IAA signaling pathway. GA3 also had a stronger effect on stalk elongation than IAA. The results of qRT-PCR and histological analysis revealed that GA3 and IAA induced the expansion of cell walls by activating the expression of genes encoding cell wall structural proteins such as Expansin (EXP). These findings provide new insights into the mechanism of stalk formation regulated by the combination of IAA and GA3.

Naphthylphthalamic acid associates with and inhibits PIN auxin transporters

N-1-naphthylphthalamic acid (NPA) is a key inhibitor of directional (polar) transport of the hormone auxin in plants. For decades, it has been a pivotal tool in elucidating the unique polar auxin transport-based processes underlying plant growth and development. Its exact mode of action has long been sought after and is still being debated, with prevailing mechanistic schemes describing only indirect connections between NPA and the main transporters responsible for directional transport, namely PIN auxin exporters. Here we present data supporting a model in which NPA associates with PINs in a more direct manner than hitherto postulated. We show that NPA inhibits PIN activity in a heterologous oocyte system and that expression of NPA-sensitive PINs in plant, yeast, and oocyte membranes leads to specific saturable NPA binding. We thus propose that PINs are a bona fide NPA target. This offers a straightforward molecular basis for NPA inhibition of PIN-dependent auxin transport and a logical parsimonious explanation for the known physiological effects of NPA on plant growth, as well as an alternative hypothesis to interpret past and future results. We also introduce PIN dimerization and describe an effect of NPA on this, suggesting that NPA binding could be exploited to gain insights into structural aspects of PINs related to their transport mechanism.

Cadmium Inhibits Lateral Root Emergence in Rice by Disrupting OsPIN-Mediated Auxin Distribution and the Protective Effect of OsHMA3

Cadmium (Cd) strongly inhibits root growth, especially the formation of lateral roots (LRs). The mechanism of Cd inhibition on LR formation in rice (Oryza sativa) remains unclear. In this study, we found that LR emergence in rice was inhibited significantly by 1 µM Cd and almost completely arrested by 5 µM Cd. Cd suppressed both the formation and subsequent development of the lateral root primordium (LRP). By using transgenic rice expressing the auxin response reporters DR5::GUS and DR5rev::VENUS, we found that Cd markedly reduced the auxin levels in the stele and LRP. Cd rapidly downregulated the expression of the auxin efflux transporter genes OsPIN1b, OsPIN1c and OsPIN9 in the stele and LRP. The emergence of LRs in a rice cultivar with a null allele of OsHMA3 (Heavy Metal ATPase 3) was more sensitive to Cd than cultivars with functional alleles. Overexpression of functional OsHMA3 in rice greatly alleviated the inhibitory effect of Cd, but the protective effect of OsHMA3 was abolished by the auxin polar transport inhibitor 1-N-naphthylphthalamic acid. The results suggest that Cd inhibits LR development in rice by disrupting OsPIN-mediated auxin distribution to LRP and OsHMA3 protects against Cd toxicity by sequestering Cd into the vacuoles.

TDIF regulates auxin accumulation and modulates auxin sensitivity to enhance both adventitious root and lateral root formation in poplar trees

Of six TRACHEARY ELEMENT DIFFERENTIATION INHIBITORY FACTOR (TDIF)-encoding genes in poplar, PtTDIF1 is predominantly expressed in adventitious roots (ARs), and the other five PtTDIFs are preferentially expressed in lateral roots (LRs). Upon auxin application, expression of all PtTDIFs declined in ARs but transiently increased in LRs. Both exogenous TDIF peptides and overexpression of PtTDIFs in poplar positively regulated the initiation and elongation of LRs, and overexpression of PtTDIFs also increased the number of ARs. As visualized by the auxin-responsive marker DR5:GUS, TDIF had differential impacts on the auxin signaling activity in ARs and LRs, which was corroborated by the free indole-3-acetic acid (IAA) measurements in them. Shoot tips of PtTDIF2- and PtTDIFL2-overexpressing (together as PtTDIFsOE) trees revealed an enhanced IAA biosynthetic capacity, and removal of the aerial tissues dramatically diminished the root phenotypes of micro-propagated PtTDIFsOE trees. Furthermore, PtTDIFsOE poplars displayed an increased sensitivity for exogenous IAA, and N-1-naphthylphthalamic acid (NPA) completely blocked the TDIF-induced AR and LR formation. In PtTDIFsOE roots, several auxin-related LR initiation markers such as GATA23, LBD16 and LBD29 were transcriptionally upregulated, further supporting that TDIF regulates LR organogenesis by strengthening the spatiotemporal auxin cues and that dynamic interplays between hormones govern root branching and developmental plasticity in tree species.