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The molecular basis of cellular auxin transport is still not fully understood. Although a number of carriers have been identified and proved to be involved in auxin transport, their regulation and possible activity of as yet unknown transporters remain unclear. Nevertheless, using single-cell-based systems it is possible to track the course of auxin accumulation inside cells and to specify and quantify some auxin transport parameters. The synthetic auxins 2,4-dichlorophenoxyacetic acid (2,4-D) and naphthalene-1-acetic acid (NAA) are generally considered to be suitable tools for auxin transport studies because they are transported specifically via either auxin influx or efflux carriers, respectively. Our results indicate that NAA can be metabolized rapidly in tobacco BY-2 cells. The predominant metabolite has been identified as NAA glucosyl ester and it is shown that all NAA metabolites were retained inside the cells. This implies that the transport efficiency of auxin efflux transporters is higher than previously assumed. By contrast, the metabolism of 2,4-D remained fairly weak. Moreover, using data on the accumulation of 2,4-D measured in the presence of auxin transport inhibitors, it is shown that 2,4-D is also transported by efflux carriers. These results suggest that 2,4-D is a promising tool for determining both auxin influx and efflux activities. Based on the accumulation data, a mathematical model of 2,4-D transport at a single-cell level is proposed. Optimization of the model provides estimates of crucial transport parameters and, together with its validation by successfully predicting the course of 2,4-D accumulation, it confirms the consistency of the present concept of cellular auxin transport.
- MeSH
- biologický transport MeSH
- kultivované buňky MeSH
- kyselina 2,4-dichlorfenoxyoctová chemie metabolismus MeSH
- kyseliny indoloctové chemie metabolismus MeSH
- naftaleny chemie metabolismus MeSH
- regulátory růstu rostlin chemie metabolismus MeSH
- tabák chemie cytologie metabolismus MeSH
- teoretické modely MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- MeSH
- biologický transport účinky léků MeSH
- buněčné dělení fyziologie účinky léků MeSH
- finanční podpora výzkumu jako téma MeSH
- kultivované buňky MeSH
- kyselina 2,4-dichlorfenoxyoctová farmakologie metabolismus MeSH
- kyseliny indoloctové metabolismus MeSH
- tabák metabolismus MeSH
- transportní proteiny MeSH
Polar auxin transport is a crucial process for control and coordination of plant development. Studies of auxin transport through plant tissues and organs showed that auxin is transported by a combination of phloem flow and the active, carrier-mediated cell-to-cell transport. Since plant organs and even tissues are too complex for determination of the kinetics of carrier-mediated auxin uptake and efflux on the cellular level, simplified models of cell suspension cultures are often used, and several tobacco cell lines have been established for auxin transport assays. However, there are very few data available on the specificity and kinetics of auxin transport across the plasma membrane for Arabidopsis thaliana suspension-cultured cells. In this report, the characteristics of carrier-mediated uptake (influx) and efflux for the native auxin indole-3-acetic acid and synthetic auxins, naphthalene-1-acetic and 2,4-dichlorophenoxyacetic acids (NAA and 2,4-D, respectively) in A. thaliana ecotype Landsberg erecta suspension-cultured cells (LE line) are provided. By auxin competition assays and inhibitor treatments, we show that, similarly to tobacco cells, uptake carriers have high affinity towards 2,4-D and that NAA is a good tool for studies of auxin efflux in LE cells. In contrast to tobacco cells, metabolic profiling showed that only a small proportion of NAA is metabolized in LE cells. These results show that the LE cell line is a useful experimental system for measurements of kinetics of auxin carriers on the cellular level that is complementary to tobacco cells.
- MeSH
- Arabidopsis cytologie růst a vývoj metabolismus MeSH
- biologický transport MeSH
- buněčné kultury MeSH
- fenotyp MeSH
- hypokotyl cytologie růst a vývoj metabolismus MeSH
- kotyledon cytologie růst a vývoj metabolismus MeSH
- kyselina 2,4-dichlorfenoxyoctová metabolismus MeSH
- kyseliny indoloctové metabolismus MeSH
- kyseliny naftalenoctové metabolismus MeSH
- listy rostlin cytologie růst a vývoj metabolismus MeSH
- metabolom MeSH
- regulátory růstu rostlin metabolismus MeSH
- semenáček cytologie růst a vývoj metabolismus MeSH
- tabák cytologie růst a vývoj metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
A surface plasmon resonance (SPR) biosensor for simultaneous detection of multiple organic pollutants exhibiting endocrine-disrupting activity, namely atrazine, benzo[a]pyrene, 2,4-dichlorophenoxyacetic acid (2,4-D) and 4-nonylphenol, is reported. The biosensor utilizes a multichannel SPR sensor based on wavelength modulation of SPR and wavelength division multiplexing (WDM) of sensing channels, antibodies as biorecognition element and a competitive immunoassay detection format. An analysis time of 45 min (including 30-min incubation of the sample with antibodies) and limits of detection as low as 0.05, 0.07, 0.16 and 0.26 ng mL(-1) are demonstrated for benzo[a]pyrene, atrazine, 2,4-D and 4-nonylphenol, respectively. The biosensor is also shown to be regenerable and suitable for repeated use.
- MeSH
- atrazin analýza MeSH
- benzopyren analýza MeSH
- biosenzitivní techniky metody přístrojové vybavení MeSH
- časové faktory MeSH
- endokrinní disruptory analýza MeSH
- fenoly analýza MeSH
- financování organizované MeSH
- imunoanalýza metody MeSH
- kyselina 2,4-dichlorfenoxyoctová analýza MeSH
- povrchová plasmonová rezonance metody přístrojové vybavení MeSH
- senzitivita a specificita MeSH
Developmental responses to auxin are regulated by facilitated uptake and efflux, but detailed molecular understanding of the carrier proteins is incomplete. We have used pharmacological tools to explore the chemical space that defines substrate preferences for the auxin uptake carrier AUX1. Total and partial loss-of-function aux1 mutants were assessed against wild-type for dose-dependent resistance to a range of auxins and analogues. We then developed an auxin accumulation assay with associated mathematical modelling to enumerate accurate IC50 values for a small library of auxin analogues. The structure activity relationship data were analysed using molecular field analyses to create a pharmacophoric atlas of AUX1 substrates. The uptake carrier exhibits a very high level of selectivity towards small substrates including the natural indole-3-acetic acid, and the synthetic auxin 2,4-dichlorophenoxyacetic acid. No AUX1 activity was observed for herbicides based on benzoic acid (dicamba), pyridinyloxyacetic acid (triclopyr) or the 6-arylpicolinates (halauxifen), and very low affinity was found for picolinic acid-based auxins (picloram) and quinolinecarboxylic acids (quinclorac). The atlas demonstrates why some widely used auxin herbicides are not, or are very poor substrates. We list molecular descriptors for AUX1 substrates and discuss our findings in terms of herbicide resistance management.
- MeSH
- Arabidopsis metabolismus MeSH
- biologické modely MeSH
- biotest MeSH
- herbicidy metabolismus MeSH
- indoly metabolismus MeSH
- inhibiční koncentrace 50 MeSH
- kořeny rostlin růst a vývoj MeSH
- kyselina 2,4-dichlorfenoxyoctová metabolismus MeSH
- kyseliny indoloctové metabolismus MeSH
- mutace genetika MeSH
- proteiny huseníčku metabolismus MeSH
- semenáček růst a vývoj MeSH
- substrátová specifita MeSH
- tabák cytologie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
