OsCYP2, a chaperone involved in degradation of auxin‐responsive proteins,plays crucial roles in rice lateral root initiation

Auxin plays a pivotal role in many facets of plant development. It acts by inducing the interaction between auxin‐responsive [auxin (AUX)/indole‐3‐acetic acid (IAA)] proteins and the ubiquitin protein ligase SCF^TIR to promote the degradation of the AUX/IAA proteins. Other cofactors and chaperones that participate in auxin signaling remain to be identified. Here, we characterized rice (Oryza sativa) plants with mutations in a cyclophilin gene (OsCYP2). cyp2 mutants showed defects in auxin responses and exhibited a variety of auxin‐related growth defects in the root. In cyp2 mutants, lateral root initiation was blocked after nuclear migration but before the first anticlinal division of the pericycle cell. Yeast two‐hybrid and in vitro pull‐down results revealed an association between OsCYP2 and the co‐chaperone Suppressor of G2 allele of skp1 (OsSGT1). Luciferase complementation imaging assays further supported this interaction. Similar to previous findings in an Arabidopsis thaliana SGT1 mutant (atsgt1b), degradation of AUX/IAA proteins was retarded in cyp2 mutants treated with exogenous 1‐naphthylacetic acid. Our results suggest that OsCYP2 participates in auxin signal transduction by interacting with OsSGT1.     

In vivo metabolism of labelled indole-3-acetic acid during polar transport in etiolated hypocotyls of Lupinus albus: Relationship with growth

The in vivo metabolism of indole-3-acetic acid (IAA) in etiolated hypocotyls of lupin (Lupinus albus L., from Bari, Italy) was investigated by appliying IAA labelled with two radioisotopes ([1-¹⁴C]-IAA+[5-³H]-IAA) to the apical end of decapitated seedlings, followed by extraction of the radioactivity in the different regions along the hypocotyl. This method allowed detection of IAA decarboxylation in zones distant from the cut surface and, therefore, containing intact cells. When IAA was added directly in solution to the cut surface, decarboxylation was high especially in those hypocotyl regions where transient accumulations characteristic of the polar transport of IAA occurred. In 10-day-old seedlings such accumulations were observed both in the elongation zone (2^nd, 3^rd, and 4^th cm) and in the non elongating basal zone (8^th, 9^th and 10^th cm). When the IAA, instead, was applied with an agar block deposited on the cut surface, IAA metabolism (decarboxylation as well as conjugation) was increased but almost exclusively in tissues within 10 mm of the cut surface. In both kinds of experiment, the increase in IAA decarboxylation seemed to coincide with a decrease in the transport of IAA, since in the assay without agar the transient accumulations of radioactivity were probably due to a decrease in the transport velocity, while in the assay with agar the transport intensity was much lower than in the assay without agar. These results point to a competitive relationship between IAA metabolism and transport. Consequently, it is suggested that hypocotyl regions that probably use auxin for development processes (e.g., cell elongation and differentiation) may have a more intense IAA metabolism in parallel with their higher IAA concentrations.     

An unexpected extended conformation for the third TPR motif of the peroxin PEX5 from Trypanosoma brucei

A number of helix-rich protein motifs are involved in a variety of critical protein-protein interactions in living cells. One of these is the tetratrico peptide repeat (TPR) motif that is involved, amongst others, in cell cycle regulation, chaperone function and post-translation modifications. So far, these helix-rich TPR motifs have always been observed to be a compact unit of two helices interacting with each other in antiparallel fashion. Here, we describe the structure of the first three TPR-motifs of the peroxin PEX5 from Trypanosoma brucei, the causative agent of sleeping sickness. Peroxins are proteins involved in peroxisome, glycosome and glyoxysome biogenesis. PEX5 is the receptor of the proteins targeted to these organelles by the "peroxisomal targeting signal-1", a C-terminal tripeptide called PTS-1. The first two of the three TPR-motifs of T. brucei PEX5 appear to adopt the canonical antiparallel helix hairpin structure. In contrast, the third TPR motif of PEX5 has a dramatically different conformation in our crystals: the two helices that were supposed to form a hairpin are folded into one single 44 A long continuous helix. Such a conformation has never been observed before for a TPR motif. This raises interesting questions including the potential functional importance of a "jack-knife" conformational change in TPR motifs.     

Auxin and chloroplast movements

Auxin is involved in a wide spectrum of physiological processes in plants, including responses controlled by the blue light photoreceptors phototropins: phototropic bending and stomatal movement. However, the role of auxin in phototropin‐mediated chloroplast movements has never been studied. To address this question we searched for potential interactions between auxin and the chloroplast movement signaling pathway using different experimental approaches and two model plants, Arabidopsis thaliana and Nicotiana tabacum. We observed that the disturbance of auxin homeostasis by shoot decapitation caused a decrease in chloroplast movement parameters, which could be rescued by exogenous auxin application. In several cases, the impairment of polar auxin transport, by chemical inhibitors or in auxin carrier mutants, had a similar negative effect on chloroplast movements. This inhibition was not correlated with changes in auxin levels. Chloroplast relocations were also affected by the antiauxin p‐chlorophenoxyisobutyric acid and mutations in genes encoding some of the elements of the SCF^TIR1‐Aux/IAA auxin receptor complex. The observed changes in chloroplast movement parameters are not prominent, which points to a modulatory role of auxin in this process. Taken together, the obtained results suggest that auxin acts indirectly to regulate chloroplast movements, presumably by regulating gene expression via the SCF^TIR1‐Aux/IAA‐ARF pathway. Auxin does not seem to be involved in controlling the expression of phototropins.     

Growth Inhibition and Recovery in Roots Following Temporary Treatment with Auxin

Continuous recording (streak photography) of elongation of roots treated with IAA (10^–6–10^–7M) showed that removal of IAA from the nutrient solution resulted in a rapid resumption of elongation, unless the IAA treatment was shorter than 60 min. If it was shorter, the recovery was delayed, so that it occurred about 1 hour after the beginning of the treatment, independently of the duration of the treatment, down to 4 min. This behavior of roots was observed in all the species investigated (corn, pea, sunflower, onion), also in response to NAA and 2,4-D. This time lag in recovery of elongation after brief auxin treatment is discussed in connection with the radial concentration gradient of auxin in the root imposed by external auxin. The possible role of a radial gradient of auxin (concentration decreasing with distance from the center) in the control of root elongation is suggested.     

The Role of Auxin Metabolism in Root Meristem Regeneration by Pinus lambertiana Embryo Cuttings

Since the existence of root promoting substances that consist of a complex between auxin and another molecule has been suggested, we have examined the role of auxin conversion products in root regeneration by Pinus lambertiana embryo cuttings. Auxin conversion products were detected using radioactive forms of the auxins IAA (indoIe-3-acetic acid), NAA (a-napthaleneacetic acid) and 2,4-D (2,4-dichlorophenoxyacetic acid). 10^?7M NAA was more effective than 10^?6M IAA at promoting rooting, yet it formed conversion products much less rapidly. Also continuous exposure to IAA was necessary for optimum root formation. Based on these and other findings, we conclude that free auxin, and not the conversion products we detected, is essential to root meristem formation.     

A saturable site responsible for polar transport of indole-3-acetic acid in sections of maize coleoptiles

The velocity of transport and shape of a pulse of radioactive indole-3-acetic acid (IAA) applied to a section of maize (Zea mays L.) coleoptile depends strongly on the concentration of nonradioactive auxin in which the section has been incubated before, during, and after the radioactive pulse. A pulse of [³H]IAA disperses slowly in sections incubated in buffer (pH 6) alone; but when 0.5–5 M IAA is included, the pulse achieves its maximum velocity of about 2 cm h^-1. At still higher IAA concentrations in the medium, a transition occurs from a discrete, downwardly migrating pulse to a slowly advancing profile. Specificity of IAA in the latter effect is indicated by the observation that benzoic acid, which is taken up to an even greater extent than IAA, does not inhibit movement of [³H]IAA. These results fully substantiate the hypothesis that auxin transport consists of a saturable flux of auxin anions (A^-) in parallel with a nonsaturable flux of undissociated IAA (HA), with both fluxes operating down their respective concentration gradients. When the anion site saturates, the movement of [³H]IAA is nonpolar and dominated by the diffusion of HA. Saturating polar transport also results in greater cellular accumulation of auxin, indicating that the same site mediates the cellular efflux of A^-. The transport inhibitors napthylphthalamic acid and 2,3,5-triiodobenzoic acid specifically block the polar A^- component of auxin transport without affecting the nonsaturable component. The transport can be saturated at any point during its passage through the section, indicating that the carriers are distributed throughout the tissue, most likely in the plasmalemma of each cell.Abbreviations A^- auxin anion - HA undissociated auxin - IAA indole-3-acetic acid - NPA N-1-napthylphthalamic acid - TIBA 2,3,5-triiodobenzoic acid     

Different Behaviour of Indole-3-Acetic Acid and Indole-3-Butyric Acid in Stimulating Lateral Root Development in Rice (Oryza sativa L.)

The plant hormone auxin has been shown to be involved in lateral root development and application of auxins, indole-3-acetic acid (IAA) and indole-3-butyric acid (IBA), increases the number of lateral roots in several plants. We found that the effects of two auxins on lateral root development in the indica rice (Oryza sativa L. cv. IR8) were totally different from each other depending on the application method. When the roots were incubated with an auxin solution, IAA inhibited lateral root development, while IBA was stimulatory. In contrast, when auxin was applied to the shoot, IAA promoted lateral root formation, while IBA did not. The transport of [³H]IAA from shoot to root occurred efficiently (% transported compared to supplied) but that of [³H]IBA did not, which is consistent with the stimulatory effect of IAA on lateral root production when applied to the shoot. The auxin action of IBA has been suggested to be due to its conversion to IAA. However, in rice IAA competitively inhibited the stimulatory effect of IBA on lateral root formation when they were applied to the incubation solution, suggesting that the stimulatory effect of IBA on lateral root development is not through its conversion to IAA.