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Leaf attachment, wounding, and control of leaf expansion in arabidopsis by the plant hormone indole-3-acetic acid.

Indole-3-acetic acid (IAA) is a hormone responsible for controlling various aspects of plant growth including leaf expansion. Previous work has shown that increasing the IAA content of intact expanding leaves of Arabidopsis and Phaseolus, either through exogenous application or through trapping the endogenous hormone in leaves, results in inhibition of leaf growth (1). Paradoxically, other work has clearly shown that treatment of excised leaf strips from tobacco (Nicotiana) (2) and from beans (Phaseolus) (3) with IAA stimulates rather than inhibits growth.

Two hypothetical explanations might explain the apparent reversal of growth sensitivity to IAA by leaf tissues upon excision. It is possible that excision isolates the tissue from interaction with a second unstable growth controller that, in the intact plant, interacts with IAA to inhibit growth. Alternatively, wounding may induce a collapse in endogenous IAA levels in excised leaf tissues so that IAA levels, which are optimal for growth in the intact plant, become sub-optimal. To assess these possibilities, we have monitored growth responses to IAA of attached leaves, wounded attached leaves, excised leaves, and of excised leaf strips of the model plant system Arabidopsis.

For our experiments, stratified Arabidopsis seeds were sown in 288 well plug trays containing moist potting soil and grown in a growth chamber at 19[degrees]C, with continuous illumination (150 [micro]M [s.sup-1] [m.sup.-2]). After 10-14 days, plants were selected with both the first two true leaves 2.8-3.2 mm in diameter. For one set of experiments, scaled digital images of the intact attached leaves were prepared for subsequent determination of initial leaf area. For another set of experiments detached leaves were imaged. Initial images of excised leaf strips (0.7 mm wide cut transversely across the midpoint of the leaves) were prepared for a third set of experiments and for a fourth set of experiments wounded attached leaves (sliced transversely from leaf edge to near the midvein in three places) were imaged. For all experiments one of the first two leaves (selected randomly) from each plant served as the experimental (IAA treated) leaf and the other leaf served as a paired control. Treatment solutions included: full strength Murashige and Skoog media (Caisson Laboratories, Rexburg, ID), 10 mM KC1, 0.1 mM Mes/Btp (pH 6.0), +/- IAA at various concentrations (10 [micro]M, 50 [micro]M, 100 [micro]M, 300 [micro]M, and 1 mM). Following initial imaging attached leaves and wounded attached leaves received either a 5[micro]l drop of a control treatment solution (minus IAA) or a 5 [micro]l drop of the treatment solution containing IAA. Detached leaves and leaf strips were each incubated on 3 mL of the same solutions. One day (24 hours) later, the attached leaves and wounded leaves were detached from the plant and again imaged as were the detached leaves and excised leaf strips.

The results do not clearly differentiate between our two alternative hypotheses. As expected, growth of intact Arabidopsis leaves, though somewhat insensitive, is inhibited by IAA at higher concentrations. For example, at 300 [micro]M the area of IAA treated leaves increased 51.7 +/- 10.1% compared to 77.2 +/- 6.3 % for the controls (n=12). Also, as expected, the growth excised leaf strips was increased by IAA treatment across a range of concentrations 10 [micro]M and higher. The growth of wounded attached leaves was significantly inhibited by IAA applied at 50 [micro]M and higher. This result argues against the possibility of wounding inducing lower leaf IAA levels and supports the possibility that detachment is a requirement for IAA-induced growth increase. Growth of detached leaves floated on solution, however, were inhibited by IAA with growth responding to as little as 10 [micro]M IAA, indicating that detachment does not, by itself, reverse IAA sensitivity. It remains possible, however, that only substantial wounding, as experienced by leaf strip tissues, is sufficient to induce a decline in IAA level. We are currently collaborating with Jerry Cohen's lab at the University of Minnesota to test this possibility through analysis of the IAA content of variously treated leaf tissues.

This project is supported by NIH grant P20 RR016741 from the NCRR

Derek S. Lentz *, Morgan L. Grundstad, Michael Evanoff and Christopher P. Keller

Department of Biology, Minot State University, Minot, ND 58707

(1) Keller CP, Stahlberg R, Barkawi L, Cohen JD (2004) Plant Physiology 134: 1217-1226.

(2) Keller CP, Van Volkenburgh E (1997) Plant Physiology 113:603-610

(3) Keller CP (2007) Physiologia Plantarum 130:580-589
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Author:Lentz, Derek S.; Grundstad, Morgan L.; Evanoff, Michael; Keller, Christopher P.
Publication:Proceedings of the North Dakota Academy of Science
Article Type:Technical report
Geographic Code:1U4ND
Date:Apr 1, 2008
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