Synthesis and hydrolysis of auxins and their conjugates with different side-chain lengths: are all products active auxins?

Plants need hormone substances to regulate a plethora of responses during their life cycle. One major hormone class is called auxin, which is involved in many developmental processes. Besides the major auxin indole-3-acetic acid, there are other auxin-like molecules present in some, but not in all plants, an example would be chlorinated IAA in legumes. Among these are also the auxins with longer chains, indole-3-propionic acid and indole-3-butyric acid. The auxin-dependent growth response is dependent on the concentration of the compound. While lower concentrations are mainly growth promoting, high concentrations are actually inhibiting some developmental processes. Therefore, tight control of the auxin concentration is essential for proper growth and development. This can be achieved by altering the amount of active auxin via transport, biosynthesis, degradation or reversible conjugation to small molecules. In addition, plants use auxin during their interaction with the environment, for example during abiotic stresses such as salt, temperature or water stress to adapt the growth responses specifically. Furthermore, auxin is involved in the development of plant disease symptoms, such as tumor growth or aberrant tissue formation. However, together with other plant hormones such as salicylic acid auxin can also modulate disease progression or resistance in different plant – microbe combinations. </p

Auxin Amidohydrolases – From Structure to Function: Revisited

The control of plant growth and development is a well-coordinated process between exogenous and endogenous signals. Auxins are plant hormones belonging to the endogenous signals, which control a vast array of different processes. While auxins are growth promoting at low concentrations, higher levels are often inhibitory. Therefore, the tight control of auxin concentrations in a given plant tissue is essential. Among several processes that participate in auxin homeostasis, we focused herein on the process of reversible auxin conjugation that considers the synthesis of inactive auxin conjugates, which can be hydrolyzed back to the active form by so called auxin conjugate hydrolases. Although these proteins have been known for quite some time, their role in plants is still not clear, especially since novel hydrolases with different substrate specificities have been isolated. Thus, we have revisited the knowledge about auxin hydrolases, from their structure and biochemistry to the role in plant development and in dealing with unfavorable climate conditions. This work is licensed under a Creative Commons Attribution 4.0 International License

High-level expression of a viscotoxin in Arabidopsis thaliana gives enhanced resistance against Plasmodiophora brassicae

Viscotoxins are a group of toxic thionins found in several mistletoe species. The constitutive CaMV-Ω promoter was used to drive the expression of the viscotoxin A3 cDNA from Viscum album in transgenic Arabidopsis thaliana C24. Lines with high viscotoxin A3 levels in all parts of the plant were selected and tested for resistance against the clubroot pathogen Plasmodiophora brassicae. The transgenic lines were more resistant to infection by this pathogen than the parental lin

A Novel Target (Oxidation Resistant 2) in Arabidopsis thaliana to Reduce Clubroot Disease Symptoms via the Salicylic Acid Pathway without Growth Penalties

The clubroot disease (Plasmodiophora brassicae) is one of the most damaging diseases worldwide among brassica crops. Its control often relies on resistant cultivars, since the manipulation of the disease hormones, such as salicylic acid (SA) alters plant growth negatively. Alternatively, the SA pathway can be increased by the addition of beneficial microorganisms for biocontrol. However, this potential has not been exhaustively used. In this study, a recently characterized protein Oxidation Resistant 2 (OXR2) from Arabidopsis thaliana is shown to increase the constitutive pathway of SA defense without decreasing plant growth. Plants overexpressing AtOXR2 (OXR2-OE) show strongly reduced clubroot symptoms with improved plant growth performance, in comparison to wild type plants during the course of infection. Consequently, oxr2 mutants are more susceptible to clubroot disease. P. brassicae itself was reduced in these galls as determined by quantitative real-time PCR. Furthermore, we provide evidence for the transcriptional downregulation of the gene encoding a SA-methyltransferase from the pathogen in OXR2-OE plants that could contribute to the phenotype.Fil: Mencia, Regina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; ArgentinaFil: Welchen, Elina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Agrobiotecnología del Litoral. Universidad Nacional del Litoral. Instituto de Agrobiotecnología del Litoral; ArgentinaFil: Auer, Susann. Technische Universität Dresden; AlemaniaFil: Ludwig Müller, Jutta. Technische Universität Dresden; Alemani

Genetically transformed roots: From plant disease to biotechnological resource

Hairy root syndrome is a disease that is induced by Agrobacterium rhizogenes infection and characterized by a proliferation of excessively branching roots. However, in the past 30 years A. rhizogenes-mediated transformation has also provided a valuable platform for studying biosynthesis pathways in plants. Furthermore, the genetically transformed root cultures are becoming increasingly attractive, cost-effective options for mass-producing desired plant metabolites and expressing foreign proteins. Numerous proof-of-concept studies have demonstrated the feasibility of scaling up hairy-root-based processes while maintaining their biosynthetic potential. Recently, hairy roots have also shown immense potential for applications in phytoremediation, that is, plant-based decontamination of polluted environments. This review highlights recent progress and limitations in the field, and outlines future perspectives for the industrial exploitation of hairy roots.Fil: Georgiev, Milen I.. Bulgarian Academy of Sciences; Bulgaria. Leiden University; Países BajosFil: Agostini, Elizabeth. Universidad Nacional de Río Cuarto; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba; ArgentinaFil: Ludwig Müller, Jutta. Technische Universitat Dresden; AlemaniaFil: Xu, Jianfeng. University of Arkansas for Medical Sciences; Estados Unido

Improvement of root architecture under abiotic stress through control of auxin homeostasis in Arabidopsis and Brassica crops

Auxin plays an important role in many aspects of plant development including stress responses. Here we briefly summarize how auxin is involved in salt stress, drought (i.e. mostly osmotic stress), waterlogging and nutrient deficiency in Brassica plants. In addition, some mechanisms to control auxin levels and signaling in relation to root formation (under stress) will be reviewed. Molecular studies are mainly described for the model plant Arabidopsis thaliana, but we also like to demonstrate how this knowledge can be transferred to agriculturally important Brassica species, such as Brassica rapa, Brassica napus and Brassica campestris. Moreover, beneficial fungi could play a role in the adaptation response of Brassica roots to abiotic stresses. Therefore, the possible influence of Piriformospora indica will also be covered since the growth promoting response of plants colonized by P. indica is also linked to plant hormones, among them auxin

The paramutated SULFUREA locus of tomato is involved in auxin biosynthesis

The tomato (Solanum lycopersicum) sulfurea mutation displays trans-inactivation of wild-type alleles in heterozygous plants, a phenomenon referred to as paramutation. Homozygous mutant plants and paramutated leaf tissue of heterozygous plants show a pigment-deficient phenotype. The molecular basis of this phenotype and the function of the SULFUREA gene (SULF) are unknown. Here, a comprehensive physiological analysis of the sulfurea mutant is reported which suggests a molecular function for the SULFUREA locus. It is found that the sulf mutant is auxin-deficient and that the pigment-deficient phenotype is likely to represent only a secondary consequence of the auxin deficiency. This is most strongly supported by the isolation of a suppressor mutant which shows an auxin overaccumulation phenotype and contains elevated levels of indole-3-acetic acid (IAA). Several lines of evidence point to a role of the SULF gene in tryptophan-independent auxin biosynthesis, a pathway whose biochemistry and enzymology is still completely unknown. Thus, the sulfurea mutant may provide a promising entry point into elucidating the tryptophan-independent pathway of IAA synthesis

Utjecaj strukturno srodnih flavonoida na ekspresiju hsp gena u ljudskim promieloidnim leukemijskim stanicama

Quercetin is a known specific inhibitor of hsp70 synthesis and thus might be a potent agent for enhancing the selective cytotoxicity of heat on tumour cells. A comparative analysis of the effects of quercetin and five structurally related flavonoids on hsp90α, hsp70A, hsp60 and hsp27 gene expression was carried out using human myeloid leukaemia cells (HL-60). The cells were preincubated with 50 μM quercetin, kaempferol, myricetin, taxifolin, isorhamnetin, methylquercetagetin or 0.1 % DMSO (controls) for 24 h at 37 °C before heat shock treatment (43 °C for 30 min). Total RNA was isolated from heat-stressed and unstressed cells and analysed by RT PCR. Hsp27 gene expression was inhibited by flavonoids more strongly than other hsp genes investigated in heat stressed as well as in unstressed cells. Among the hsp genes tested, only hsp60 was expressed above control level under the influence of taxifolin. Members of the hsp70 and hsp27 families are highly expressed in breast and lung cancer and leukaemias and they play a role in the acquired resistance to chemotherapy or radiation therapy combined with hyperthermia. Therefore, hsps present potential targets for cancer diagnosis and treatment. The present structure/activity study indicates that position, number and substitution of hydroxyl groups of the B ring and saturation of the C2-C3 bond are important factors affecting flavonoid activity on hsp gene expression. This study could help provide a basis for further design of specific inhibitors of hsp gene expression.Kvercetin je poznati specifični inhibitor sinteze hsp70 i kao takav mogući čimbenik povećanja selektivnog citotoksičnog učinka topline na tumorske stanice. Provedena je komparativna analiza učinka kvercetina i njemu strukturno srodnih flavonoida na ekspresiju hsp90α, hsp70A, hsp60 i hsp27 gena u stanicama humane mijeloidne leukemije (HL-60). Stanice su inkubirane s 50 μM kvercetina, kempferola, miricetina, taksifolina, izoramnetina, metilkvercetagetina ili 0,1% DMSO (kontrola) tijekom 24 h na 37 °C prije obrade toplinskim šokom (43 °C tijekom 30 minuta). Izolirana je ukupna RNA iz stanica koje su bile, kao i one koje nisu bile, podvrgnute toplinskom stresu te je provedena RT-PCR analiza. Ekspresija gena hsp27 bila je jače inhibirana flavonoidima nego ostali istraživani hsp geni, i to podjednako u stanicama podvrgnutim kao i u onima koje nisu bile podvrgnute toplinskom stresu. Među istraživanim hsp genima jedino je ekspresija hsp60 bila iznad kontrolne razine pod utjecajem taksifolina. Članovi porodica hsp70 i hsp27 snažno su ekspresirani kod raka dojke, pluća i leukemija te imaju bitnu ulogu u stečenoj rezistenciji stanica pri kemoterapiji ili terapiji zračenjem u kombinaciji s hipertermijom. Stoga bi hsp proteine trebalo istraživati pri dijagnosticiranju i liječenju raka. Prikazana studija strukture i aktivnosti upućuje na to da su položaj, broj i supstitucija hidroksilnih skupina na prstenu B kao i zasićeni vez C2–C3 kod flavonoida bitni čimbenici koji utječu na ekspresiju hsp gena. Ova bi studija mogla poslužiti kao osnova za daljnje strukturiranje specifičnih inhibitora ekspresije hsp gena

Evidence for the Early Evolutionary Loss of the M20D Auxin Amidohydrolase Family from Mosses and Horizontal Gene Transfer from Soil Bacteria of Cryptic Hydrolase Orthologues to Physcomitrella patens

Inactive auxin conjugates are accumulated in plants and hydrolyzed to recover phytohormone action. A family of metallopeptidase orthologues has been conserved in Plantae to help regulate auxin homeostatic levels during growth and development. This hydrolase family was recently traced back to liverwort, the most ancient extant land plant lineage. Liverwort’s auxin hydrolase has little activity against auxin conjugate substrates and does not appear to actively regulate auxin. This finding, along with data that shows moss can synthesize auxin conjugates, led to examining another bryophyte lineage, Physcomitrella patens. We have identified and isolated three M20D hydrolase paralogues from moss. The isolated enzymes strongly recognize and cleave a variety of auxin conjugates, including those of indole butyric and indole propionic acids. These P. patens hydrolases not only appear to be “cryptic”, but they are likely to have derived from soil bacteria through Horizontal Gene Transfer. Additionally, support is presented that the plant-type M20D peptidase family may have been universally lost from mosses after divergence from the common ancestor with liverwort

Praćenje metabolizma flavonoida u humanim stanicama na temelju fluorescencije izazvane interakcijom kvercetina s proteinima

Despite the wealth of information concerning biological effects of flavonoids, a systematic approach to analyzing the molecular targets is still lacking and, for this reason, a rational evaluation of the risks or benefits of flavonoid-containing foods or of possible pharmaceutical applications is difficult. We have exploited the property of quercetin to elicit fluorescence when bound to specific target proteins and assayed several flavonoids with different modifications (methylation, hydroxylation, glycosylation). Quercetin target proteins can be visualized in living cells, but in vital human leukaemia cells (HL-60) the fluorescence decreases rapidly after labelling, while metabolically inactive apoptotic cells retain the fluorescence. These cytological differences were apparent under the fluorescent microscope and were quantified using flow cytometry. Metabolic conversion of quercetin in vital cells was confirmed and quantified by HPLC analysis. While apoptotic cells still contained considerable amounts of quercetin, vital cells rapidly metabolized the flavonoid (e.g., by methylation or glycosylation). Biochemical results are consistent with the cytological observations and support the conclusion that quercetin becomes rapidly converted to non-fluorogenic metabolites in vital cells. Loss of fluorescence in vital cells allows convenient monitoring and quantifying of the dynamics of quercetin metabolism in human cells.Unatoč mnoštvu informacija koje se odnose na biološke učinke flavonoida, sustavni pristup analizi njihovih ciljnih molekula još uvijek nedostaje. Iz toga razloga vrlo je teško racionalno vrednovati opasnosti ili koristi koje donosi hrana koja sadrži flavonoide kao i njihovu moguću farmakološku primjenu. Iskoristili smo svojstvo kvercetina da izazove fluorescenciju kada se veže za specifične ciljne proteine i analizirali nekoliko različito modificiranih flavonoida (metilacija, hidroksilacija, glikozilacija). Ciljni proteini za koje se kvercetin veže u živim stanicama mogu se vizualizirati na temelju fluorescencije. U živim stanicama humane leukemije (HL-60) fluorescencija naglo pada nakon označavanja flavonoidima, dok metabolički inaktivne apoptotične stanice zadržavaju fluorescenciju. Te su citološke razlike jasno zapažene pod fluorescencijskim mikroskopom, a kvantificirane su pomoću protočne citometrije. Metabolička pretvorba kvercetina u živim stanicama potvr|ena je i kvantificirana pomoću HPLC analiza. Dok apoptotične stanice zadržavaju značajnu količinu kvercetina, žive ga stanice brzo metaboliziraju (npr. metilacijom ili glikozilacijom). Ti su biokemijski rezultati u skladu s citološkim promatranjima i podupiru zaključak da se kvercetin u živim stanicama brzo pretvara u nefluorogene metabolite. Gubitak fluorescencije u živim stanicama omogućava praćenje i kvantifikaciju dinamike metabolizma kvercetina u humanim stanicama