The Biochemistry And Genetics Of Plant Surface Lipid Biosynthesis And Assembly

The waxy cuticle that covers the aerial epidermis of plants provides protection from desiccation and environmental stresses and was likely a critical adaptation that allowed plants to colonize land. The cuticle consists of various organic solvent soluble waxes that are embedded within, and layered upon, a non-soluble polyester matrix of cutin. Despite the fundamental importance of the cuticle, relatively little is known about its synthesis and metabolism. Tomato (Solanum lycopersicum) is an emerging model species for studying cuticle biology is, as it has extensive genetic resources and a fruit cuticle that is substantial and easily isolated. This work describes the results of three studies that leverage the attributes of tomato as a model for cuticle biology. First, a survey of the morphology and chemistry of the fruit cuticles in 6 wild relatives of tomato is described. While these species are separated by less than 7 million years of evolution, they are endemic to diverse environments, and substantial differences in cuticle morphology and composition were observed. The genetic basis of one of these traits was further dissected by chemical analysis of a S. habrochaites introgression line population. The second study used a proteomic approach to identify proteins associated with the cuticle of expanding tomato fruits. This study identified several candidates for the extracellular transport and assembly of cuticle precursors. One of iii these was independently identified by forward genetics of the cutin deficient 1 (cd1) mutant and shown to be required for accumulation of the cutin polymer. The CD1 gene encodes an extracellular GDSL-motif lipase/hydrolase family protein, and the final study describes the biochemical characterization of the CD1 protein and the cd1 mutant. In extracts from the cd1 mutant, but not the wild type, the 2-monoacylglycerol (2-MAG) derivative of the major cutin monomer of tomato was identified by GC-MS. In vitro biochemical characterization of CD1 demonstrated acyltransferase activity with a preference for a 2-MAG acyl donor. Collectively, these results suggest that CD1 catalyzes extracellular cutin polymerization via successive transesterification of 2-MAG derivatives of cutin monomers, thus indicating for the first time the monomeric precursor and enzymatic mechanism of cutin polymerization. i