Total Wax Load Research Articles

Cuticular waxes on eceriferum mutants of Arabidopsis thaliana

We present cuticular wax chemical profiles for the leaves and stems of Arabidopsis wildtype Landsberg erecta and eleven isogenic e cer iferum mutants: cer5, cer10 to cer15, and cer17 to cer20. These cer mutants have wax profiles that are different from those of wildtype in chemical chain length distribution, amount per chemical class, and/or total wax load. Analyses of detailed leaf and stem wax profiles for these cer mutants have allowed us to place some of these mutants at specific steps in wax production. The cer13 gene is predicted to affect release of the 30 carbon fatty acid from the elongation complex or the reduction of C30 fatty acid to C30 aldehyde. The CER19 gene product is predicted to be involved in C28 to C30 fatty acyl-CoA elongation. The CER20 gene is predicted to affect the oxidation of C29 alkane to C29 secondary alcohol. Several predicted gene products affect only stem specific steps in the wax pathway.

Leaf sheath cuticular waxes on bloomless and sparse-bloom mutants of Sorghum bicolor

Leaf sheath cuticular waxes on wild-type Sorghum bicolor were approximately 96% free fatty acids, with the C 28 and C 30 acids being 77 and 20% of these acids, respectively. Twelve mutants with markedly reduced wax load were characterized for chemical composition. In all of the 12 mutants, reduction in the amount of C 28 and C 30 acids accounted for essentially all of the reduction in total wax load relative to wildtype. The bm2 mutation caused a 99% reduction in total waxes. The bm4, bm5, bm6, bm7 and h10 mutations caused more than 91% reduction in total waxes, whereas the remaining six mutants, bm9, bm11, h7, h11, h12 and h13, caused between 35 and 78% reduction in total wax load. Relative to wild-type, bm4 caused a large increase in the absolute amount of C 22, C 24 and C 26 acids, and reduction in the C 28 and longer acids, suggesting that bm4 may suppress elongation of C 26 acyl-CoA primarily. The h10 mutation increased the absolute amounts of the longest chain length acids, but reduced shorter acids, suggesting that h10 may suppress termination of acyl-CoA elongation. The bm6, bm9, bm11, h7, h11, h12 and h13 mutations increased the relative amounts, but not absolute amounts, of longer chain acids. Based on chemical composition alone, it is still uncertain which genes and their products were altered by these mutations. Nevertheless, these Sorghum cuticular wax mutants should provide a valuable resource for future studies to elucidate gene involvement in the biosynthesis of cuticular waxes, in particular, the very-long-chain fatty acids.

134 Seasonal Variation in Composition and Total Load of Leaf Blade Epicuticular Waxes on Hosta

Hosta variants for epicuticular waxes were selected based on variation in surface glaucousness, from highly glaucous to highly glossy. In an effort to determine seasonal variation in hosta waxes, gas-chromatography mass-spectrometry was used to perform detailed chemical analysis of the adaxial and abaxial leaf blade waxes four times points during the growing season, early spring, mid-spring, mid-summer, and autumn. These studies revealed that in all variants, the total wax loads increased dramatically during the period of leaf expansion in the spring, dropped roughly five fold by midsummer, and then accumulated slightly above summer levels into the fall season. The dominant wax constituent class on all hosta cultivars was primary alcohols. Changes in these alcohols were primarily responsible for seasonal changes in total wax load. In some variants, the shorter chain length alcohols were unusually high compared with alcohol distributions normally found on other plants. Besides primary alcohols, significant amounts of acids, aldehydes, and alkanes, were also found and shown to vary during the growing season. A possible association between these seasonal changes in wax profiles and hosta resistance to slugs is discussed.

KCS1 encodes a fatty acid elongase 3-ketoacyl-CoA synthase affecting wax biosynthesis in Arabidopsis thaliana.

An Arabidopsis fatty acid elongase gene, KCS1, with a high degree of sequence identity to FAE1, encodes a 3-ketoacyl-CoA synthase which is involved in very long chain fatty acid synthesis in vegetative tissues, and which also plays a role in wax biosynthesis. Sequence analysis of KCS1 predicted that this synthase was anchored to a membrane by two adjacent N-terminal, membrane-spanning domains. Analysis of a T-DNA tagged kcs1-1 mutant demonstrated the involvement of the KCS1 in wax biosynthesis. Phenotypic changes in the kcs1-1 mutant included thinner stems and less resistance to low humidity stress at a young age. Complete loss of KCS1 expression resulted in decreases of up to 80% in the levels of C26 to C30 wax alcohols and aldehydes, but much smaller effects were observed on the major wax components, i.e. the C29 alkanes and C29 ketones on leaves, stems and siliques. In no case did the loss of KCS1 expression result in complete loss of any individual wax component or significantly decrease the total wax load. This indicated that there was redundancy in the elongase KCS activities involved in wax synthesis. Furthermore, since alcohol, aldehyde, alkane and ketone levels were affected to varying degrees, involvement of the KCS1 synthase in both the decarbonylation and acyl-reduction wax synthesis pathways was demonstrated.

Epicuticular wax variation in ecotypes of Arabidopsis thaliana

Quantification of the epicuticular wax from the stems of 40 ecotypes of Arabidopsis thaliana showed a two-fold range in total wax load that was not correlated to known abiotic characteristics of the ecotype's origin of collection. Chemical analysis of these ecotypes revealed similar epicuticular wax profiles for all ecotypes except CT-1. In CT-1 the amount of 22 and 24 carbon length primary alcohols was increased by 16- and 8-fold, respectively, over that observed in the epicuticular wax averaged over all ecotypes.

Changes in epicuticular waxes on wildtype and eceriferum mutants in Arabidopsis during development

We analysed the leaf epicuticular wax chemical constituents on wildtype and eceriferum ( cer; cerl, cer3 and cer4) mutants of Arabidopsis thaliana at 7, 15, 25 and 40 days after germination. Changes during development are described for the partitioning of epicuticular waxes into the five major chemical classes; free carboxylic fatty acids, aldehydes, alkanes, primary alcohols and wax esters. Changes are described for cumulative total epicuticular wax loads per leaf area, percentages of total constituents in each of the five major chemical classes and chain length distributions for the two major chemical classes: alkanes and primary alcohols. Stem epicuticular waxes on wildtype and cer mutants were analysed at 25 and 40 days and found to be similar.