Good neighbours : the role of non-lignified cells in Arabidopsis lignification

Lignin is a critical structural component of plants, providing vascular integrity and mechanical strength. Lignin precursors, monolignols, must be exported to the extracellular matrix where random oxidative coupling produces a complex lignin polymer. The objectives of this study were twofold: to determine the timing of lignification, with respect to programmed cell death during Arabidopsis thaliana primary xylem development, and to determine which cells are contributing to the lignification of tracheary elements and fibres. This thesis demonstrates that lignin deposition is not exclusively a post-mortem event, but also occurs prior to programmed cell death. Radiolabelled monolignols were not detected in the cytoplasm or vacuoles of tracheary elements or neighbours. To experimentally define which cells in lignifying tissues contribute to lignification in intact plants, a microRNA against CINNAMOYL CoA-REDUCTASE1, driven by the promoter from CELLULOSE SYNTHASE 7 (proCESA7:miRNA CCR1), was used to silence monolignol biosynthesis in cells developing secondary cell walls. When monolignol biosynthesis was knocked down specifically in the cells with thickened secondary cell walls, but not in the neighbouring cells, lignin was still deposited in the xylem secondary cell walls. This indicates that “good neighbour” cells are sufficient to produce lignin in the vascular bundles. Surprisingly, this was not the case in the interfascicular fibres, where a dramatic reduction in cell wall lignification demonstrates that these extra-xylary fibers undergo cell autonomous lignification. When a fibre-specific promoter (proAtPEROXIDASE64) was used to drive the miRNA, autonomous extraxylary fibre lignification was again observed, as was non-cell autonomous lignification between xylary fibres and neighbouring tracheary elements. These effects may have reflected compensatory mechanisms in response to lignin downregulation, so to demonstrate that discrete cell populations, such as xylem parenchyma, do contribute to lignification, genes encoding enzymes catalyzing the synthesis of novel monolignol conjugates were introduced into wild-type Arabidopsis using cell population-specific promoters. The detection of novel monolignol conjugates in the cell wall by chemical analysis and fluorescence microscopy supported the contribution of tracheary elements and fibres to lignification and also revealed that xylary parenchyma cells are producing monolignol substrates and acting as “good neighbours” to tracheary elements and xylary fibres during lignification.Science, Faculty ofBotany, Department ofGraduat