Ethylene and auxin control the Arabidopsis response to decreased light intensity

Morphological responses of plants to shading have long been studied as a function of light quality, in particular the ratio of red to far red light that affects phytochrome activity. However, changes in light quantity are also expected to be important for the shading response because plants have to adapt to the reduction in overall energy input. Here, we present data on the involvement of auxin and ethylene in the response to low light intensities. Decreased light intensities coincided with increased ethylene production in Arabidopsis rosettes. This response was rapid because the plants reacted within minutes. In addition, ethylene- and auxin-insensitive mutants are impaired in their reaction to shading, which is reflected by a defect in leaf elevation and an aberrant leaf biomass allocation. On the molecular level, several auxin-inducible genes are up-regulated in wild-type Arabidopsis in response to a reduction in light intensity, including the primary auxin response gene IAA3 and a protein with similarity to AUX22 and the 1-aminocyclopropane-1-carboxylic acid synthase genes ACS6, ACS8, and ACS9 that are involved in ethylene biosynthesis. Taken together, the data show that ethylene and auxin signaling are required for the response to low light intensities.     

Auxin and the response of pea roots to auxin transport inhibitors: morphactin

The auxin transport inhibitor methyl-2-chloro-9-hydroxyfluorene-9-carboxylate (CFM), a morphactin, inhibits negative geotropism, causes cellular swelling, and induces root hair formation in roots of intact Pisum sativum L. seedlings. In excised pea root tips, CFM inhibits elongation more than increase in fresh weight (swell ratio = 1.3 at 20 mum CFM). CFM growth inhibition was expressed in the presence of ethylene. Indoleacetic acid (IAA) prevented the expression of CFM growth inhibition possibly because IAA inhibited the accumulation of CFM into the tissue sections. CFM inhibited the accumulation of IAA and 2,4-dichlorophenoxyacetic acid into excised root tips. Applying Leopold's (1963. Brookhaven Symp. Biol. 16: 218-234) model for polar auxin transport, this result suggests a possible explanation for CFM inhibition of geotropism in pea roots, i.e. disruption of auxin transport by interfering with auxin binding.     

Interactions between auxin transport and the actin cytoskeleton in developmental polarity of Fucus distichus embryos in response to light and gravity

Land plants orient their growth relative to light and gravity through complex mechanisms that require auxin redistribution. Embryos of brown algae use similar environmental stimuli to orient their developmental polarity. These studies of the brown algae Fucus distichus examined whether auxin and auxin transport are also required during polarization in early embryos and to orient growth in already developed tissues. These embryos polarize with the gravity vector in the absence of a light cue. The auxin, indole-3-acetic acid (IAA), and auxin efflux inhibitors, such as naphthylphthalamic acid (NPA), reduced environmental polarization in response to gravity and light vectors. Young rhizoids are negatively phototropic, and NPA also inhibits rhizoid phototropism. The effect of IAA and NPA on gravity and photopolarization is maximal within 2.5 to 4.5 h after fertilization (AF). Over the first 6 h AF, auxin transport is relatively constant, suggesting that developmentally controlled sensitivity to auxin determines the narrow window during which NPA and IAA reduce environmental polarization. Actin patches were formed during the first hour AF and began to photolocalize within 3 h, coinciding with the time of NPA and IAA action. Treatment with NPA reduced the polar localization of actin patches but not patch formation. Latrunculin B prevented environmental polarization in a time frame that overlaps the formation of actin patches and IAA and NPA action. Latrunculin B also altered auxin transport. Together, these results indicate a role for auxin in the orientation of developmental polarity and suggest interactions between the actin cytoskeleton and auxin transport in F. distichus embryos.     

The auxin transport inhibitor response 3 (tir3) allele of BIG and auxin transport inhibitors affect the gibberellin status of Arabidopsis

The Arabidopsis gene BIG (formerly DOC1/TIR3/UMB1/ASA1) is known to encode a huge calossin-like protein that is required for polar auxin transport (PAT). Mutations at this locus, in addition to reducing PAT, can alter the sensitivity of plants to several hormones and light. The tir3-1 allele of BIG reduces the response of plants to application of the gibberellin (GA) precursors ent-kaurenoic acid and GA12 and its semidwarf phenotype is partially reversed by C19-GAs. The effects of auxin transport inhibitors (ATIs) on GA 20-oxidation was examined in wild-type and tir3-1 seedlings. 1-N-naphthylphthalamic acid (NPA) and triiodobenzoic acid lead to overexpression of the GA-biosynthetic gene AtGA20ox1 comparable in magnitude to the overexpression observed in seedlings treated with paclobutrazol, a GA biosynthesis inhibitor. In contrast to that of AtGA20ox1, overexpression of AtGA20ox2 is pronounced only in paclobutrazol-treated Col and Ler, and is less in tir3-1 and in all NPA-treated seedlings. Thus the effects of BIG and ATIs on the expression of genes encoding GA 20-oxidases are complex, and suggest that at least in some tissues ATIs, directly or indirectly, may reduce the level of bioactive GA and/or alter GA signal transduction.     

The exocyst complex contributes to PIN auxin efflux carrier recycling and polar auxin transport in Arabidopsis

In land plants polar auxin transport is one of the substantial processes guiding whole plant polarity and morphogenesis. Directional auxin fluxes are mediated by PIN auxin efflux carriers, polarly localized at the plasma membrane. The polarization of exocytosis in yeast and animals is assisted by the exocyst: an octameric vesicle‐tethering complex and an effector of Rab and Rho GTPases. Here we show that rootward polar auxin transport is compromised in roots of Arabidopsis thaliana loss‐of‐function mutants in the EXO70A1 exocyst subunit. The recycling of PIN1 and PIN2 proteins from brefeldin–A compartments is delayed after the brefeldin‐A washout in exo70A1 and sec8 exocyst mutants. Relocalization of PIN1 and PIN2 proteins after prolonged brefeldin‐A treatment is largely impaired in these mutants. At the same time, however, plasma membrane localization of GFP:EXO70A1, and the other exocyst subunits studied (GFP:SEC8 and YFP:SEC10), is resistant to brefeldin‐A treatment. In root cells of the exo70A1 mutant, a portion of PIN2 is internalized and retained in specific, abnormally enlarged, endomembrane compartments that are distinct from VHA‐a1‐labelled early endosomes or the trans‐Golgi network, but are RAB‐A5d positive. We conclude that the exocyst is involved in PIN1 and PIN2 recycling, and thus in polar auxin transport regulation.     

Impaired sterol ester synthesis alters the response of Arabidopsis thaliana to Phytophthora infestans

Non‐host resistance of Arabidopsis thaliana against Phytophthora infestans, the causal agent of late blight disease of potato, depends on efficient extracellular pre‐ and post‐invasive resistance responses. Pre‐invasive resistance against P. infestans requires the myrosinase PEN2. To identify additional genes involved in non‐host resistance to P. infestans, a genetic screen was performed by re‐mutagenesis of pen2 plants. Fourteen independent mutants were isolated that displayed an enhanced response to Phytophthora (erp) phenotype. Upon inoculation with P. infestans, two mutants, pen2‐1 erp1‐3 and pen2‐1 erp1‐4, showed an enhanced rate of mesophyll cell death and produced excessive callose deposits in the mesophyll cell layer. ERP1 encodes a phospholipid:sterol acyltransferase (PSAT1) that catalyzes the formation of sterol esters. Consistent with this, the tested T‐DNA insertion lines of PSAT1 are phenocopies of erp1 plants. Sterol ester levels are highly reduced in all erp1/psat1 mutants, whereas sterol glycoside levels are increased twofold. Excessive callose deposition occurred independently of PMR4/GSL5 activity, a known pathogen‐inducible callose synthase. A similar formation of aberrant callose deposits was triggered by the inoculation of erp1 psat1 plants with powdery mildew. These results suggest a role for sterol conjugates in cell non‐autonomous defense responses against invasive filamentous pathogens.     

Critical consideration on the relationship between auxin transport and calcium transients in gravity perception of Arabidopsis seedlings

Plants regulate their growth and morphogenesis in response to gravity field, known as gravitropism. In the early process of gravitropism, changes in the gravity vector (gravistimulation) are transduced into certain intracellular signals, termed gravity perception. The plant hormone auxin is not only a crucial factor to represent gravitropism but also a potential signaling molecule for gravity perception. Another strong candidate for the signaling molecule is calcium ion of which cytoplasmic concentration ([Ca²⁺]_c) is known to increase in response to gravistimulation. However, relationship between these two factors, say which is in the first place, has been controversial. This issue is addressed here mainly based on recent progress including our latest studies. Gravistimulation by turning plants 180° induced a two-peaked [Ca²⁺]_c-increase lasting for several minutes in Arabidopsis seedlings expressing apoaequorin; only the second peak was sensitive to the gravistimulation. Peak amplitudes of the [Ca²⁺]_c-increase were attenuated by the 10 µM auxin transport inhibitor (TIBA) and vesicle trafficking inhibitor (BFA), whereas the onset time and rate of rise of the second peak were not significantly altered. This result indicates that polar auxin transport is not involved in the initial phase of the second [Ca²⁺]_c-increase. It is likely that the gravi-induced [Ca²⁺]_c-increase constitutes an upstream event of the auxin transport, but may positively be modulated by auxin since its peak amplitude is attenuated by the inhibition of auxin transport.Key words: auxin, calcium, gravity perception, gravitropism, pin-formed (PIN) protein, Arabidopsis thaliana     

Evidence for altered polar and lateral auxin transport in the gravity persistent signal (gps) mutants of Arabidopsis

Plant shoots do not respond when they are reoriented relative to gravity at 4 degrees C. However, when returned to vertical at room temperature, these organs bend in response to the previous cold gravistimulation. The inflorescence stem of the Arabidopsis thaliana gravity persistent signal (gps) mutants respond abnormally after the cold gravistimulation: gps1 does not bend when returned to room temperature, gps2 bends the wrong way and gps3 over-responds, curving past the predicted angle. In wild type and the mutants, basipetal auxin transport in the inflorescence stem was abolished at 4 degrees C but restored when plants were returned to room temperature. In gps1, auxin transport was increased; in both gps2 and gps3, no significant difference was found when compared to wild type. Expression of the auxin-inducible P(IAA2)::GUS reporter gene, indicated that auxin-induced gene expression was redistributed to the lower side of the inflorescence stem in wild type after gravistimulation at 4 degrees C. In gps1, no asymmetries in P(IAA2)::GUS expression were seen. In gps2, P(IAA2)::GUS expression was localized to the upper side of the stem and in gps3, asymmetric P(IAA2):GUS expression was extended throughout the elongation zone of the inflorescence stem. These results are consistent with altered lateral Indole-3-acetic-acid (IAA) gradients being responsible for the phenotype of each mutant.     

The binding of auxin to the Arabidopsis auxin influx transporter AUX1

The cellular import of the hormone auxin is a fundamental requirement for the generation of auxin gradients that control a multitude of plant developmental processes. The AUX/LAX family of auxin importers, exemplified by AUX1 from Arabidopsis (Arabidopsis thaliana), has been shown to mediate auxin import when expressed heterologously. The quantitative nature of the interaction between AUX1 and its transport substrate indole-3-acetic acid (IAA) is incompletely understood, and we sought to address this in the present investigation. We expressed AUX1 to high levels in a baculovirus expression system and prepared membrane fragments from baculovirus-infected insect cells. These membranes proved suitable for determination of the binding of IAA to AUX1 and enabled us to determine a K(d) of 2.6 mum, comparable with estimates for the K(m) for IAA transport. The efficacy of a number of auxin analogues and auxin transport inhibitors to displace IAA binding from AUX1 has also been determined and can be rationalized in terms of their physiological effects. Determination of the parameters describing the initial interaction between a plant transporter and its hormone ligand provides novel quantitative data for modeling auxin fluxes.     

Entrainment of Arabidopsis roots to the light:dark cycle by light piping

Correct operation of the plant circadian clock is crucial for optimal growth and development. Recent evidence has shown that the plant clock is tissue specific and potentially hierarchical, implying that there are signalling mechanisms that can synchronise the clock in different tissues. Here, I have addressed the mechanism that allows the shoot and root clocks to be synchronised in light:dark cycles but not in continuous light. Luciferase imaging data from 2 different Arabidopsis accessions with 2 different markers show that the period of the root clock is much less sensitive to blue light than to red light. Decapitated roots were imaged either in darkness or with the top section of root tissue exposed to light. Exposure to red light reduced the period of the root tissue maintained in darkness, whereas exposure to blue light did not. The data indicate that light can be piped through root tissue to affect the circadian period of tissue in darkness. I propose that the synchronisation of shoots and roots in light:dark cycles is achieved by light piping from shoots to roots.     

The unconventional prefoldin RPB5 interactor mediates the gravitropic response by modulating cytoskeleton organization and auxin transport in Arabidopsis

Gravity-induced root curvature involves the asymmetric distribution of the phytohormone auxin. This response depends on the concerted activities of the auxin transporters such as PIN-FORMED(PIN)proteins for auxin efflux and AUXIN RESISTANT 1(AUX1) for auxin influx. However, how the auxin gradient is established remains elusive. Here we identified a new mutant with a short root, strong auxin distribution in the lateral root cap and an impaired gravitropic response. The causal gene encoded an Arab...     

Low-fluence red light increases the transport and biosynthesis of auxin

In plants, light is an important environmental signal that induces photomorphogenesis and interacts with endogenous signals, including hormones. We found that light increased polar auxin transport in dark-grown Arabidopsis (Arabidopsis thaliana) and tomato (Solanum lycopersicum) hypocotyls. In tomato, this increase was induced by low-fluence red or blue light followed by 1 d of darkness. It was reduced in phyA, phyB1, and phyB2 tomato mutants and was reversed by far-red light applied immediately after the red or blue light exposure, suggesting that phytochrome is involved in this response. We further found that the free indole-3-acetic acid (IAA) level in hypocotyl regions below the hook was increased by red light, while the level of conjugated IAA was unchanged. Analysis of IAA synthesized from [13C]indole or [13C]tryptophan (Trp) revealed that both Trp-dependent and Trp-independent IAA biosynthesis were increased by low-fluence red light in the top section (meristem, cotyledons, and hook), and the Trp-independent pathway appears to become the primary route for IAA biosynthesis after red light exposure. IAA biosynthesis in tissues below the top section was not affected by red light, suggesting that the increase of free IAA in this region was due to increased transport of IAA from above. Our study provides a comprehensive view of light effects on the transport and biosynthesis of IAA, showing that red light increases both IAA biosynthesis in the top section and polar auxin transport in hypocotyls, leading to unchanged free IAA levels in the top section and increased free IAA levels in the lower hypocotyl regions.