Genotype-dependent regulation of vitamin E biosynthesis in olive fruits as revealed through metabolic and transcriptional profiles.

Vitamin E is a general term used to describe a group of eight lipophilic compounds known as tocochromanols. These vitamin E variants are chemically categorised into two classes formed by α-, ß-, γ- and δ- tocopherols and tocotrienols isoforms, respectively. The present study describes the concurrent regulation of genes and metabolites orchestrating vitamin E biosynthesis in olive drupes of five distinctive Greek olive cultivars. A combination of analytical, biochemical and molecular approaches was employed in order to carry out comparative analyses, including real-time RT-qPCR for gene expression levels and HPLC analysis of metabolite content. Findings indicated that tocochromanol levels and composition, oil content, gene expression levels as well as total antioxidant activity were highly dependent on cultivar and, to a lesser extent, on fruit developmental stage. Specifically, cultivars 'Kalokairida' and 'Lianolia Kerkyras' demonstrated the highest vitamin E content. The latter possessed high tocochromanol content combined with highest overall antioxidant activity in all developmental stages, concomitant with the up-regulation expression profile of HPPD. The genotypic imprint versus the temporal contribution to vitamin E levels, as well as the potential link to lipid peroxidation amelioration, are discussed.

Influence of long-range Coulomb interaction in velocity map imaging.

The standard velocity-map imaging (VMI) analysis relies on the simple approximation that the residual Coulomb field experienced by the photoelectron ejected from a neutral or ion system may be neglected. Under this almost universal approximation, the photoelectrons follow ballistic (parabolic) trajectories in the externally applied electric field, and the recorded image may be considered as a 2D projection of the initial photoelectron velocity distribution. There are, however, several circumstances where this approximation is not justified and the influence of long-range forces must absolutely be taken into account for the interpretation and analysis of the recorded images. The aim of this paper is to illustrate this influence by discussing two different situations involving isolated atoms or molecules where the analysis of experimental images cannot be performed without considering long-range Coulomb interactions. The first situation occurs when slow (meV) photoelectrons are photoionized from a neutral system and strongly interact with the attractive Coulomb potential of the residual ion. The result of this interaction is the formation of a more complex structure in the image, as well as the appearance of an intense glory at the center of the image. The second situation, observed also at low energy, occurs in the photodetachment from a multiply charged anion and it is characterized by the presence of a long-range repulsive potential. Then, while the standard VMI approximation is still valid, the very specific features exhibited by the recorded images can be explained only by taking into consideration tunnel detachment through the repulsive Coulomb barrier.

Molecular characterization of a tomato endo-beta-1,4-glucanase gene expressed in mature pistils, abscission zones and fruit.

A cDNA (TAC1) and genomic clone (cel5) encoding an endo-beta-1,4-glucanase (EGase) were identified from tomato (Lycopersicon esculentum Mill., cv. Rutgers). The cel5 gene is expressed in pistils, flower pedicel and leaf abscission zones, and ripening fruit. The genomic sequence includes a 22 bp 5' upstream sequence that is conserved in a closely related peach EGase gene, ppEG1.

Three different polygalacturonases are expressed in tomato leaf and flower abscission, each with a different temporal expression pattern.

Abscission, or organ separation, is accompanied by a marked increase in hydrolases, which are responsible for the degradation of the middle lamella and the loosening of the primary cell wall surrounding cells in the separation layer. We recently reported on the cloning of a tomato (Lycopersicon esculentum) polygalacturonase (PG) cDNA, TAPG1, expressed during leaf and flower abscission. In addition to TAPG1, we have cloned two more PG cDNAs (TAPG2 and TAPG4) that are also expressed during leaf and flower abscission. The peptide sequences for the three abscission PGs are relatively similar (76-93% identity) yet different from the those of tomato fruit PG (38-41% identity). None of the three abscission PG mRNAs are expressed in fruit, stems, petioles, or anthers of fully open flowers. An RNase protection assay revealed that all three PGs are expressed in leaf and flower abscission zones and in pistils of fully open flowers. TAPG4 mRNA is detected much earlier than TAPG1 and TAPG2 mRNA during both leaf and flower abscission.

The mRNA for an ETR1 homologue in tomato is constitutively expressed in vegetative and reproductive tissues.

Dominant mutations in the Arabidopsis ETR1 gene block the ethylene signal transduction pathway. The ETR1 gene has been cloned and sequenced. Using the ETR1 cDNA as a probe, we identified a cDNA homologue (eTAE1) from tomato. eTAE1 contains an open reading frame encoding a polypeptide of 754 amino acid residues. The nucleic acid sequence for the coding sequence in eTAE1 is 74% identical to that for ETR1, and the deduced amino acid sequence is 81% identical and 90% similar. Genomic Southern blot analysis indicates that three or more ETR1 homologues exist in tomato. RNA blots show that eTAE1 mRNA is constitutively expressed in all the tissues examined, and its accumulation in leaf abscission zones was unaffected by ethylene, silver ions (an inhibitor of ethylene action) or auxin.

Cloning of a tomato polygalacturonase expressed in abscission.

Abscission, organ separation, is accompanied by cell wall breakdown in separation layer cells. In tomato (Lycopersicon esculentum), ethylene-induced abscission is correlated with an increase in polygalacturonase (PG) and endo-beta-1,4-D-glucanase (cellulase) activity. We have identified a putative, abscission-specific cDNA clone for PG, pTAPG1. The TAPG1 cDNA has 43% identity at the amino acid level with the tomato fruit PG. Genomic blot analysis suggests that the gene for TAPG1 is a member of a small subfamily of PG genes that is distinct from the tomato fruit PG. The TAPG1 cDNA hybridizes to mRNA expressed during the course of ethylene-induced leaf and flower abscission. A high level of PG transcript accumulation coincides with the occurrence of abscission. Auxin, an abscission inhibitor, and silver thiosulfate, an ethylene action inhibitor, suppressed accumulation of mRNA in leaf abscission zones complementary to the TAPG1 cDNA. Expression of TAPG1 transcripts is several-fold higher in flower abscission zones than in leaf abscission zones. The identification of cDNAs that encode abscission-specific PG provide and additional tool to study the regulation of abscission and cell wall dissolution in separation layer cells.

Cellulase occurs in multiple active forms in ripe avocado fruit mesocarp.

The existence of multiple forms of avocado (Persea americana Mill. cv Hass) cellulase in crude protein extracts of ripe avocado fruit is reported. Cellulase was separated into at least 11 multiple forms by native isoelectric focusing in the pH range between 4 and 7 and visualized by both activity staining using Congo red and immunostaining. The enzyme components were acidic proteins with isoelectric points in the range of pH 5.10 to 6.80, the predominant forms having isoelectric points of 5.60, 5.80, 5.95, and 6.20. All 11 forms were immunologically related with molecular masses of 54 kilodaltons.