Salicylic acid (SA), a natural defensive signal chemical, and antimycin A, a cytochrome pathway inhibitor, induce resistance to Tobacco mosaic virus (TMV). Pharmacological evidence suggested signaling during resistance induction by both chemicals involved alternative oxidase (AOX), sole component of the alternative respiratory pathway (AP). Roles of the AP include regulation of intramitochondrial reactive oxygen species and maintenance of metabolic homeostasis. Transgenic tobacco (Nicotiana tabacum) with modified AP capacities (2-to 3-fold increased or decreased) showed no alteration in phenotype with respect to basal susceptibility to TMV or the ability to display SA-induced resistance to systemic viral disease. However, in directly inoculated tissue, antimycin A-induced TMV resistance was inhibited in plants with increased AP capacities, whereas SA and antimycin A-induced resistance was transiently enhanced in plant lines with decreased AP capacities. We conclude that SA-induced TMV resistance results from activation of multiple mechanisms, a subset of which are inducible by antimycin A and influenced by AOX. Other antiviral factors, potentially including the SA-inducible RNA-dependent RNA polymerase, are regulated by AOX-independent mechanisms.

SUMMARY The plant signal molecule salicylic acid (SA) can induce resistance to a wide range of pathogen types. In the case of viruses, SA can stimulate the inhibition of all three main stages in virus infection: replication, cell-to-cell movement and long-distance movement. Induction of resistance by SA appears to depend, in part, on downstream signalling via the mitochondrion. However, evidence has recently emerged that SA may stimulate a separate downstream pathway, leading to the induction of an additional mechanism of resistance based on RNA interference. In this review our aims are to document these recent advances and to suggest possible future avenues of research on SA-induced resistance to viruses.

Five economically important crop pests, Manduca sexta, Pieris brassicae, Mamestra brassicae, Spodoptera exigua, and Agrotis ipsilon, were tested at two stages of larval development for susceptibility to Bacillus thuringiensis toxins Cry1Ac, Cry1Ca, Cry1J, and Cry1Ba. Bioassay results for M. sexta showed that resistance to all four Cry toxins increased from the neonate stage to the third-instar stage; the increase in resistance was most dramatic for Cry1Ac, the potency of which decreased 37-fold. More subtle increases in resistance during larval development were seen in M. brassicae for Cry1Ca and in P. brassicae for Cry1Ac and Cry1J. By contrast, the sensitivity of S. exigua did not change during development. At both larval stages, A. ipsilon was resistant to all four toxins. Because aminopeptidase N (APN) is a putative Cry1 toxin binding protein, APN activity was measured in neonate and third-instar brush border membrane vesicles (BBMV). With the exception of S. exigua, APN activity was found to be significantly lower in neonates than in third-instar larvae and thus inversely correlated with increased resistance during larval development. The binding characteristics of iodinated Cry1 toxins were determined for neonate and third-instar BBMV. In M. sexta, the increased resistance to Cry1Ac and Cry1Ba during larval development was positively correlated with fewer binding sites in third-instar BBMV than in neonate BBMV. The other species-instar-toxin combinations did not reveal positive correlations between potency and binding characteristics. The correlation between binding and potency was inconsistent for the species-instar-toxin combinations used in this study, reaffirming the complex mode of action of Cry1 toxins.The ␦ endotoxins are a family of insecticidal proteins produced by Bacillus thuringiensis during sporulation. These toxins are typically found in parasporal crystals that are released into the environment with the bacterial spores. Numerous ␦ endotoxins produced by B. thuringiensis have been identified and are grouped on the basis of sequence homology and insect specificity (40). The Cry1 toxins are a group of ␦ endotoxins that principally target lepidopteran species, including several important crop pests.The mechanism of action of the Cry1 ␦ endotoxins begins with solubilization of the protoxin in the alkaline larval midgut, followed by proteolytic processing by midgut proteases (40). The stable 60-to 65-kDa toxins then bind to midgut receptors and insert into the apical membrane of brush border epithelial cells to form pores. These pores disrupt functional membrane processes and are ultimately responsible for larval death (40).Each type of Cry1 toxin has a unique spectrum of activity and targets only a small range of lepidopteran species. Within the small target ranges there are dramatic differences in potency between species that are often closely related (12,15,31). Indeed, the potency of a Cry1 toxin can significantly decrease as the larvae age (1, 38). Variations in the potencies of Cry1 toxins ...

The alternative oxidase (AOX) is the terminal oxidase of the cyanide-resistant alternative respiratory pathway in plants and has been implicated in resistance to viruses. When tobacco mosaic virus (TMV) vectors were used to drive very high levels of expression of either AOX or AOX mutated in its active site (AOX-E), virus spread was enhanced. This was visualized as the induction of larger hypersensitive-response lesions after inoculation onto NN-genotype tobacco than those produced by vectors bearing sequences of comparable length [the green fluorescent protein (gfp) gene sequence or antisense aox] or the 'empty' viral vector. Also, in the highly susceptible host Nicotiana benthamiana, systemic movement of TMV vectors expressing AOX or AOX-E was faster than that of TMV constructs bearing gfp or antisense aox sequences. Notably, in N. benthamiana, TMV.AOX and TMV.AOX-E induced symptoms that were severe and ultimately included cell death, whereas the empty vector, TMV.GFP and the TMV vector expressing antisense aox sequences never induced necrosis. The results show that, if expressed at sufficiently high levels, active and inactive AOX proteins can affect virus spread and symptomology in plants.

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