Cuticular waxes play a pivotal role in limiting transpirational water loss across the plant surface. The correlation between the chemical composition of the cuticular waxes and their function as a transpiration barrier is still unclear. In the present study, intact tomato fruits (Lycopersicon esculentum) are used, due to their astomatous surface, as a novel integrative approach to investigate this composition- function relationship: wax amounts and compositions of tomato were manipulated before measuring unbiased cuticular transpiration. First, successive mechanical and extractive wax-removal steps allowed the selective modification of epi- and intracuticular wax layers. The epicuticular film consisted exclusively of very-long-chain aliphatics, while the intracuticular compartment contained large quantities of pentacyclic triterpenoids as well. Second, applying reverse genetic techniques, a loss-of-function mutation with a transposon insertion in a very-long-chain fatty acid elongase beta-ketoacyl-CoA synthase was isolated and characterized. Mutant leaf and fruit waxes were deficient in n-alkanes and aldehydes with chain lengths beyond C30, while shorter chains and branched hydrocarbons were not affected. The mutant fruit wax also showed a significant increase in intracuticular triterpenoids. Removal of the epicuticular wax layer, accounting for one-third of the total wax coverage on wild-type fruits, had only moderate effects on transpiration. By contrast, reduction of the intracuticular aliphatics in the mutant to approximately 50% caused a 4-fold increase in permeability. Hence, the main portion of the transpiration barrier is located in the intracuticular wax layer, largely determined by the aliphatic constituents, but modified by the presence of triterpenoids, whereas epicuticular aliphatics play a minor role.

Cuticular waxes play a pivotal role in limiting transpirational water loss across the primary plant surface. The astomatous fruits of the tomato (Lycopersicon esculentum) 'MicroTom' and its lecer6 mutant, defective in a b-ketoacyl-coenzyme A synthase, which is involved in very-long-chain fatty acid elongation, were analyzed with respect to cuticular wax load and composition. The developmental course of fruit ripening was followed. Both the 'MicroTom' wild type and lecer6 mutant showed similar patterns of quantitative wax accumulation, although exhibiting considerably different water permeances. With the exception of immature green fruits, the lecer6 mutant exhibited about 3-to 8-fold increased water loss per unit time and fruit surface area when compared to the wild type. This was not the case with immature green fruits. The differences in final cuticular barrier properties of tomato fruits in both lines were fully developed already in the mature green to early breaker stage of fruit development. When the qualitative chemical composition of fruit cuticular waxes during fruit ripening was investigated, the deficiency in a b-ketoacyl-coenzyme A synthase in the lecer6 mutant became discernible in the stage of mature green fruits mainly by a distinct decrease in the proportion of n-alkanes of chain lengths . C 28 and a concomitant increase in cyclic triterpenoids. This shift in cuticular wax biosynthesis of the lecer6 mutant appears to be responsible for the simultaneously occurring increase of water permeance. Changes in cutin composition were also investigated as a function of developmental stage. This integrative functional approach demonstrates a direct relationship between cuticular transpiration barrier properties and distinct chemical modifications in cuticular wax composition during the course of tomato fruit development.

Maintaining the integrity of the cuticular transpiration barrier even at elevated temperatures is of vital importance especially for hot-desert plants. Currently, the temperature dependence of the leaf cuticular water permeability and its relationship with the chemistry of the cuticles are not known for a single desert plant. This study investigates whether (i) the cuticular permeability of a desert plant is lower than that of species from non-desert habitats, (ii) the temperature-dependent increase of permeability is less pronounced than in those species and (iii) whether the susceptibility of the cuticular permeability barrier to high temperatures is related to the amounts or properties of the cutin or the cuticular waxes. We test these questions with Rhazya stricta using the minimum leaf water vapour conductance (gmin) as a proxy for cuticular water permeability. gmin of R. stricta (5.41 × 10−5 m s−1 at 25 °C) is in the upper range of all existing data for woody species from various non-desert habitats. At the same time, in R. stricta, the effect of temperature (15–50 °C) on gmin (2.4-fold) is lower than in all other species (up to 12-fold). Rhazya stricta is also special since the temperature dependence of gmin does not become steeper above a certain transition temperature. For identifying the chemical and physical foundation of this phenomenon, the amounts and the compositions of cuticular waxes and cutin were determined. The leaf cuticular wax (251.4 μg cm−2) is mainly composed of pentacyclic triterpenoids (85.2% of total wax) while long-chain aliphatics contribute only 3.4%. In comparison with many other species, the triterpenoid-to-cutin ratio of R. stricta (0.63) is high. We propose that the triterpenoids deposited within the cutin matrix restrict the thermal expansion of the polymer and, thus, prevent thermal damage to the highly ordered aliphatic wax barrier even at high temperatures.

The efficacy of the cuticular transpiration barrier and its resistance to elevated temperatures are significantly higher in a typical water-saver than in a water-spender plant growing in hot desert.

Nonspecific lipid transfer proteins reversibly bind different types of lipid molecules in a hydrophobic cavity. They facilitate phospholipid transfer between membranes in vitro, play a role in cuticle and possibly in suberin formation, and might be involved in plant pathogen defense signaling. This study focuses on the role of the lipid transfer protein AtLTPI-4 in crown gall development. Arabidopsis (Arabidopsis thaliana) crown gall tumors, which develop upon infection with the virulent Agrobacterium tumefaciens strain C58, highly expressed AtLTPI-4. Crown galls of the atltpI-4 loss-of-function mutant were much smaller compared with those of wild-type plants. The gene expression pattern and localization of the protein to the plasma membrane pointed to a function of AtLTPI-4 in cell wall suberization. Since Arabidopsis crown galls are covered by a suberin-containing periderm instead of a cuticle, we analyzed the suberin composition of crown galls and found a reduction in the amounts of long-chain fatty acids (C 18:0 ) in the atltpI-4 mutant. To demonstrate the impact of AtLtpI-4 on extracellular lipid composition, we expressed the protein in Arabidopsis epidermis cells. This led to a significant increase in the very-long-chain fatty acids C 24 and C 26 in the cuticular wax fraction. Homology modeling and lipid-protein-overlay assays showed that AtLtpI-4 protein can bind these very-long-chain fatty acids. Thus, AtLtpI-4 protein may facilitate the transfer of long-chain as well as very-long-chain fatty acids into the apoplast, depending on the cell type in which it is expressed. In crown galls, which endogenously express AtLtpI-4, it is involved in suberin formation.