UBC Theses and Dissertations
Functional characterization and spatial and temporal patterns of expression of genes involved in gibberellin and diterpene resin acid biosynthesis in white spruce Storey, Kathryn Madeleine
Conifers produce large quantities of diterpene resin acids (DRAs) as major components of the constitutive and induced oleoresin defense system. Like all vascular plants, conifers also produce gibberellin (GA) diterpene phytohormones, which influence growth and development. Conifers thus provide an interesting biological system for comparing the GA and DRA diterpene biosynthetic pathways. Despite serving different functions in growth and defense, respectively, the GA and DRA biosynthetic pathways are biochemically similar, utilizing the same isoprenoid precursors, evolutionarily related diterpene synthases (diTPSs), and functionally similar cytochrome P450 monooxygenases (CYP450s) to produce structurally similar diterpene intermediates and diterpene acid products. Functional characterization of central diTPS genes (ent-copalyl diphosphate synthase [CPS], ent-kaurene synthase [KS], levopimaradiene/abietadiene synthase [LAS]) and CYP450 genes (ent-kaurene oxidase [CYP701] and CYP720B4) in white spruce (Picea glauca), described in this thesis, allowed for comparative analysis of GA and DRA pathway genes. This thesis characterized the DXS (1-deoxy-D-xylulose 5-phosphate synthase) gene family in white spruce as additional analysis of the isoprenoid biosynthetic pathway producing the common precursor to both GAs and DRAs. Transcript expression of genes was analyzed to understand their seasonal (year-long time course of apical bud and shoot development), sample-specific (e.g. needle, stem, bud, bark/phloem, wood/xylem), and stress-specific (methyl jasmonate [MeJA] exposure) spatial and temporal patterns. Functionality of the DRA pathway was also assessed via quantification of DRA products. Expression of DRA and GA pathway genes was generally spatially separated. Expression of DRA genes was low in photosynthetic tissues but up-regulated during the time of year when trees are most likely to encounter seasonal attack from insect pests; expression declined sharply well before dormancy showing a strong seasonality to DRA production. GA related genes had broader expression across sample types and throughout the year, but spatially were mainly allocated to photosynthetic tissues. GA and DRA pathway genes all showed differential responses to MeJA treatment, and within corresponding sample types, age also played a role in expression. These studies improve our understanding of the organization of conifer chemical defenses, showing distinct differences compared with GA gene expression, and providing information on the spatial, seasonal and stress-responsive expression of DRA pathway genes.
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