Cyanide Restores N Gene–Mediated Resistance to Tobacco Mosaic Virus in Transgenic Tobacco Expressing Salicylic Acid Hydroxylase

Abstract

Salicylhydroxamic acid (SHAM), an inhibitor of alternative oxidase (AOX), blocks salicylic acid-induced resistance to tobacco mosaic virus (TMV) but does not inhibit pathogenesis-related PR-1 protein synthesis or resistance to fungal and bacterial pathogens. We found that the synthetic resistance-inducing chemical 2,6-dichloroisonicotinic acid also induced Aox transcript accumulation and SHAM-sensitive resistance to TMV. The respiratory inhibitors antimycin A and KCN also induced Aox transcript accumulation and resistance to TMV but did not induce PR-1 accumulation. Tobacco plants of the TMV-resistant cultivar Samsun NN transformed with the salicylic acid hydroxylase ( nahG ) gene could no longer restrict virus to the inoculation site, resulting in spreading necrosis instead of discrete necrotic lesions. Treatment with KCN restored TMV localization and normal lesion morphology. SHAM antagonized this effect, allowing virus escape and spreading necrosis to resume. The results demonstrate the importance of the SHAM-sensitive (potentially AOX-dependent) signal transduction pathway in mediating virus localization early in the hypersensitive response. INTRODUCTIONSalicylic acid (SA) plays a pivotal role in plant disease resistance (reviewed in Raskin, 1992;Ryals et al., 1996;Durner et al., 1997). When plants are challenged with an avirulent pathogen, the interaction of the products of the pathogen's avirulence and the host's resistance genes triggers resistance via a signal transduction system that includes SA as a key component (Delaney et al., 1994;Staskawicz et al., 1995;Baker et al., 1997). In the case of tobacco cultivars possessing the N resistance gene, inoculation with tobacco mosaic virus (TMV) induces the hypersensitive response (HR)-rapid programmed host cell death at the infection site. The spread of TMV is restricted to a small zone adjacent to the necrotic lesions, and the triggering of the HR is accompanied by an increase in SA levels (Malamy et al., 1990). In addition, parts of the plant distal to the primary infection site nonetheless acquire an enhanced degree of resistance to secondary infection, and this systemic acquired resistance (SAR) is preceded by a rise in the concentration of SA (Malamy et al., 1990).SAR is effective against TMV and a broad spectrum of other pathogens (Delaney et al., 1994). In tobacco plants containing the N gene, SAR against TMV is characterized by the production of fewer and smaller necrotic lesions in response to secondary infection with the virus (Ross, 1961a(Ross, , 1961b. The rise in SA levels and the induction of SAR are accompanied by the activation of a range of genes, including those that encode pathogenesis-related (PR) proteins (Bowles, 1990). The occurrence of PR proteins has been used in the past as a molecular marker for SAR induction (Ward et al., 1991;Kessmann et al., 1994). However, although many PR proteins are antibacterial or antifungal, none has yet been shown to act against viruses. Furthermore, the induction of PR protein accumulation and the induct...

Alternative Respiratory Pathway under Drought is Partially Mediated by Hydrogen Peroxide and Contributes to Antioxidant Protection in Wheat Leaves

“…Salicylhydroxamic acid (SHAM), a well-known inhibitor of AOX activity, has been widely used on intact tissues to inhibit AOX activity (Chivasa et al, 1997;Chivasa and Carr, 1998;Naylor et al, 1998). In a recent work (Bartoli et al, 2005), treatment with 1 mM of SHAM inhibited cyanide-insensitive oxygen uptake by near 70% in both well-irrigated and water -stressed wheat leaves.…”

Programmed cell death, mitochondria and the plant hypersensitive response

“…AOX catalyses electron flow directly from ubiquinol to oxygen, thereby creating an electron shunt that bypasses complexes III and IV of the IMM and results in a cyanide-insensitive electron-transfer pathway. AOX activation by treatment with cyanide during the HR may help to suppress cell death during the propagative phase of lesion formation and thus restrict the size of the necrotic zone 26 . Antisense suppression of AOX resulted in hypersensitivity to antimycin A, an inhibitor of complex III 27 .…”

Influence of irradiation on cyanide-resistant respiration and AOX1 multi-gene family expression during greening of etiolated rice seedlings

“…Salicylhydroxamic acid (SHAM), an inhibitor of AOX activity, has been used in AOX studies in intact tissues (Chivasa et al 1997, Chivasa and Carr 1998, Naylor et al 1998. To determine the appropriate concentration of SHAM, we mimicked a titration experiment similarly to a most recent work (Bartoli et al 2005).…”