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DEVELOPMENT AND HORMONE ACTION

Nitric Oxide and Cyclic GMP Are Messengers in the Indole Acetic Acid-Induced Adventitious Rooting Process¹

Instituto de Investigaciones Biológicas, Facultad de Ciencias Exactas y Naturales, Universidad Nacional de Mar del Plata, CC 1245, 7600 Mar del Plata, Argentina

This report describes part of the signaling pathway and some of the molecules involved in the auxin-induced adventitious root formation in cucumber (Cucumis sativus). Previous results showed that nitric oxide (NO) mediates the auxin response during adventitious root formation (Pagnussat et al., 2002). To determine the order of action of indole acetic acid (IAA) and NO within the signal transduction pathway and to elucidate the target molecules that are downstream of NO action, cucumber hypocotyl cuttings were submitted to a pretreatment leading to endogenous auxin depletion. The auxin depletion treatment provoked a 3-fold reduction of the root number in comparison to the nondepleted explants. The NO-donor sodium nitroprusside was able to promote adventitious rooting in auxin-depleted explants, whereas the specific NO scavenger cPTIO prevented the effect of sodium nitroprusside. The endogenous NO level was monitored in both control and auxin-depleted explants using a NO-specific fluorescent probe. The NO level was 3.5-fold higher in control (nondepleted) explants than in auxin-depleted ones. The exogenous application of IAA restored the NO concentration to the level found in nondepleted explants. Because NO activates the enzyme guanylate cyclase (GC), we analyzed the involvement of the messenger cGMP in the adventitious root development mediated by IAA and NO. The GC inhibitor LY83583 reduced root development induced by IAA and NO, whereas the cell-permeable cGMP derivative 8-Br-cGMP reversed this effect. The endogenous level of cGMP is regulated by both the synthesis via GC and its degradation by the phosphodiesterase activity. When assayed, the phosphodiesterase inhibitor sildenafil citrate was able to induce adventitious rooting in both nondepleted and auxin-depleted explants. Results indicate that NO operates downstream of IAA promoting adventitious root development through the GC-catalyzed synthesis of cGMP.

¹ This work was supported by grants to L.L. from Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), from Agencia Nacional de Promoción Científica y Tecnológica, from Fundación Antorchas and from Universidad Nacional de Mar del Plata (UNMdP), Argentina. L.L. is a career member and G.C.P. is a research postdoctoral fellow from CONICET Argentina. M.L.L. is a student fellow from UNMdP Argentina.

^* Corresponding author; e-mail lolama{at}mdp.edu.ar; fax 54–223–475–3150.

Received February 14, 2003; returned for revision February 26, 2003; accepted March 7, 2003.

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