Partial depolymerization of genetically modified potato tuber periderm reveals intermolecular linkages in suberin polyester.

Suberin is a biopolyester found in specialized plant tissues, both internal and external, with key frontier physiological functions. The information gathered so far from its monomer and oligomer composition, and in situ studies made by solid state techniques, haven't solved the enigma of how the suberin polyester is assembled as a macromolecule. To investigate how monomers are linked in suberin, we analyzed oligomer fragments solubilized by the partial depolymerization of suberin from potato (Solanum tuberosum) tuber periderms. The structure of the suberin oligomers, namely which monomers they included, and the type and frequency of the inter-monomer ester linkages, was assessed by ESI-MS/MS and high resolution NMR analysis. The analyzed potato periderms included the one from wild type (cv. Desirée) and from plants where suberin-biosynthesis genes were downregulated in chain elongation (StKCS6), ω-hydroxylation (CYP86A33) and feruloylation (FHT). Two building blocks were identified as possible key structures in the macromolecular development of the potato periderm suberin: glycerol - α,ω-diacid - glycerol, as the core of a continuous suberin aliphatic polyester; and glycerol - ω-hydroxyacid - ferulic acid, anchoring this polyaliphatic matrix at its periphery to the vicinal polyaromatics, through linking to ferulic acid. The silencing of the StKCS6 gene led to non-significant alterations in suberin structure, showing the relatively minor role of the very-long chain (>C28) fatty acids in potato suberin composition. The silencing of CYP86A33 gene impaired significantly suberin production and disrupted the biosynthesis of acylglycerol structures, proving the relevance of the latter and thus of the glycerol - α,ω-diacid - glycerol unit for the typical suberin lamellar organization. The silencing of the FHT gene led to a lower frequency of ferulate linkages in suberin polyester but to more polyphenolic guaiacyl units as seen by FTIR analyses in the intact polymer.

Cloning of two individual cDNAS encoding 9-cis-epoxycarotenoid dioxygenase from Gentiana lutea, their tissue-specific expression and physiological effect in transgenic tobacco.

Two 9-cis-epoxycarotenoid dioxygenase (NCED) cDNAs have been cloned from a petal library of Gentiana lutea. Both cDNAs carry a putative transit sequence for chloroplast import and differ mainly in their length and the 5'-flanking regions. GlNCED1 was evolutionary closely related to Arabidopsis thaliana NCED6 whereas GlNCED2 showed highest homology to tomato NCED1 and A. thaliana NCED3. The amounts of GlNCED2 transcript were below Northern detection in G. lutea. In contrast, GlNCED1 was specifically expressed at higher levels in developing flowers when petals start appearing. By genetic engineering of tobacco with coding regions of either gene under a constitutive promoter, their function was further analyzed. Although mRNA of both genes was detectable in the corresponding transgenic plants, a physiological effect was only found for GlNCED1 but not for GlNCED2. In germination experiments of GlNCED1 transgenic lines, delayed radicle formation and cotyledon appearance were observed. However, the transformants exhibited no improved tolerance against desiccation stress. In contrast to other plants with over-expressed NCEDs, prolonged delay of seed germination is the only abscisic-acid-related phenotypic effect in the GlNCED1 transgenic lines.