Sesquiterpene cyclase is not a determining factor for elicitor- and pathogen-induced capsidiol accumulation in tobacco

The induction of sesquiterpene cyclase, a key phytoalexin biosynthetic enzyme, and the accumulation of phytoalexins in relation to the induction of a hypersensitive response (HR) and cell necrosis in tobacco (Nicotiana tabacum L.) were investigated. When tobacco leaves were inoculated with virulent or avirulent isolates of Ralstonia solanacearum, steady-state levels of mRNA complementary to cDNA of the sensitivity-related (sts) gene str319 were dramatically induced. This cDNA clone is greater than 90% homologous with a gene coding for 5-epi-aristolochene synthase (EAS), previously described as a branch-point enzyme regulating the synthesis of capsidiol, the major sesquiterpenoid phytoalexin found in tobacco. Accumulation of EAS transcripts in leaves after inoculation with virulent and avirulent strains of R. solanacearum, or after treatment with necrotizing or non-necrotizing elicitins was rapid but transient, and restricted to the site of infiltration. Two highly similar sesquiterpene cyclase activities, 5-epi-aristolochene synthase and a vetispiradiene synthase-like activity, were found in extracts of elicitin-challenged and R. solanacearum-inoculated tobacco. Under all conditions tested, the induction of cyclase activity was closely correlated with induction of the cyclase mRNA level. In contrast, high levels of capsidiol were found only after treatment with the necrosis-inducing elicitin cryptogein, or after infiltration with HR-inducing bacterial strains. Low levels of capsidiol did accumulate after application of capsicein, an elicitin that induces little or no necrosis on tobacco, or after infection with a virulent bacterium. Hence, capsidiol accumulation, not 5-epi-aristolochene synthase gene expression or total sesquiterpene cyclase enzyme activity, appears to be a good marker for the HR of tobacco.