Non-invasive quantification of endogenous root auxin transport using an integrated flux microsensor technique

Abstract

SUMMARYIndole-3-acetic acid (IAA) is a primary phytohormone that regulates multiple aspects of plant development. Because polar transport of IAA is an essential determinant of organogenesis and dynamic tropic growth, methods to monitor IAA movement in vivo are in demand. A self-referencing electrochemical microsensor was optimized to non-invasively measure endogenous IAA flux near the surface of Zea mays roots without the addition of exogenous IAA. Enhanced sensor surface modification, decoupling of acquired signals, and integrated flux analyses were combined to provide direct, real time quantification of endogenous IAA movement in B73 maize inbred and brachytic2 (br2) auxin transport mutant roots. BR2 is localized in epidermal and hypodermal tissues at the root apex. br2 roots exhibit reduced shootward IAA transport at the root apex in radiotracer experiments and reduced gravitropic growth. IAA flux data indicates that maximal transport occurs in the distal elongation zone of maize roots, and net transport in/out of br2 roots was decreased compared to B73. Integration of short term real time flux data in this zone revealed oscillatory patterns, with B73 exhibiting shorter oscillatory periods and greater amplitude than br2. IAA efflux and influx were inhibited using 1-N-naphthylphthalamic acid (NPA), and 2-naphthoxyacetic acid (NOA), respectively. A simple harmonic oscillation model of these data produced a correlation between modeled and measured values of 0.70 for B73 and 0.69 for br2. These results indicate that this technique is useful for real-time IAA transport monitoring in surface tissues and that this approach can be performed simultaneously with current live imaging techniques.

The Signal Transducer NPH3 Integrates the Phototropin1 Photosensor with PIN2-Based Polar Auxin Transport in Arabidopsis Root Phototropism

“…This different behavior was not unexpected, as PIN2 is not expressed in Arabidopsis hypocotyls (Chen et al, 1998). Moreover, in vivo measurements of auxin flow in the root tips showed that the root apex transition zone (0.1 to 0.3 mm from the root tip in Arabidopsis; Verbelen et al, 2006;Baluš ka et al, 2010) is the most active region with respect to polar auxin flow Baluš ka et al, 2010;McLamore et al, 2010;McLamore and Porterfield, 2011) and gravity-stimulated peak cell elongation rates (Chavarria-Krauser et al, 2008). A BL illumination significantly increased the peak value of the auxin flux rate in the root apex transition zone of both dark-grown wildtype and pin2 seedlings, among which the response of the pin2 mutant was less significant than that of the wild type.…”

“…To determine the possible mechanism for the altered phototropic responses of the pin2, nph3, phot1, and phot1phot2 mutants, we further measured auxin flux profiles in vivo in the root apical region using a noninvasive microelectrode system (Mancuso et al, , 2007McLamore et al, 2010), which indicated that all lines have a net rhizosphere auxin influx in the root tip region (Figures 2A and 2B Two-way ANOVAs were performed to compare significant differences between measured peak values of auxin flux rates. Light conditions and different mutant lines were the two sources of variation; both caused extremely significant differences (P < 0.001).…”

“…Measurement of the rhizosphere auxin flux reflex activity of cell-to-cell auxin transport, which occurs in the apoplast, provides a quantitative analysis of polar auxin transport (McLamore et al, 2010;McLamore and Porterfield, 2011). A carbon nanotube auxin-selective microelectrode was used to monitor auxin fluxes in Arabidopsis roots as described previously Santelia et al, 2005;Bouchard et al, 2006;Bailly et al, 2008;Kim et al, 2010).…”

“…The measurement of rhizosphere auxin flux in this noninvasive microelectrode system was based on the application of Fick's law [J = -D(DC/ DX)] (J, flux rate of analytes; DC, concentration differences; DX, distance between two positions; D, diffusion coefficient, D auxin = 0.677 3 10 25 cm 2 s -1 at 258C) McLamore et al, 2010). Measurement of the rhizosphere auxin flux reflex activity of cell-to-cell auxin transport, which occurs in the apoplast, provides a quantitative analysis of polar auxin transport (McLamore et al, 2010;McLamore and Porterfield, 2011).…”

Biogenic Synthesis of Nanoparticles and Potential Applications: An Eco- Friendly Approach

“…Nanosensors have been developed that reacts with auxin. This is a step forward in auxin research, as it helps scientists know how plant roots acclimatize to their environment, particularly to marginal soils [95]. Nanotechnology for crop biotechnology: Nanocapsules can facilitate successful incursion of herbicides through cuticles and tissues, allowing slow and regular discharge of the active substances.…”