REGULATION OF OLEORESINOSIS IN GRAND FIR (ABIES GRANDIS) - DIFFERENTIAL TRANSCRIPTIONAL CONTROL OF MONOTERPENE, SESQUITERPENE, AND DITERPENE SYNTHASE GENES IN RESPONSE TO WOUNDING

Grand fit (Abies grandis Lindl.) has been developed as a model system for the study of wound-induced oleoresinosis in conifers as a response to insect attack. Oleoresin is a roughly equal mixture of turpentine (85% monoterpenes [C-10] and 15% sesquiterpenes [C-15]) and rosin (dit erpene [C-20] resin acids) that acts to seal wounds and is toxic to bo th invading insects and their pathogenic fungal symbionts. The dynamic regulation of wound-induced oleoresin formation was studied over 29 d at the enzyme level by in vitro assay of the three classes of synthas es directly responsible for the formation of monoterpenes, sesquiterpe nes, and diterpenes from the corresponding C-10, C-15, and C-20 prenyl diphosphate precursors, and at the gene level by RNA-blot hybridizati on using terpene synthase class-directed DNA probes. In overall appear ance, the shapes of the time-course curves for all classes of synthase activities are similar, suggesting coordinate formation of all of the terpenoid types. However, closer inspection indicates that the monote rpene synthases arise earlier, as shown by an abbreviated time course over 6 to 48 h. RNA-blot analyses indicated that the genes for all thr ee classes of enzymes are transcriptionally activated in response to w ounding, with the monoterpene synthases up-regulated first (transcript s detectable 2 h after wounding), in agreement with the results of cel l-free assays of monoterpene synthase activity, followed by the coordi nately regulated sesquiterpene synthases and diterpene synthases (tran scription beginning on d 3-4). The differential timing in the producti on of oleoresin components of this defense response is consistent with the immediate formation of monoterpenes to act as insect toxins and t heir later generation at solvent levels for the mobilization of resin acids responsible for wound sealing.