As a genetic platform, tomato (Solanum lycopersicum) benefits from rich germplasm collections and ease of cultivation and transformation that enable the analysis of biological processes impossible to investigate in other model species. To facilitate the assembly of an open genetic toolbox designed to study Solanaceae, we initiated a joint collection of publicly available gene manipulation tools. We focused on the characterization of promoters expressed at defined time windows during fruit development, for the regulated expression or silencing of genes of interest. Five promoter sequences were captured as entry clones compatible with the versatile MultiSite Gateway format: PPC2, PG, TPRP, and IMA from tomato and CRC from Arabidopsis (Arabidopsis thaliana). Corresponding transcriptional fusions were made with the GUS gene, a nuclear-localized GUS-GFP reporter, and the chimeric LhG4 transcription factor. The activity of the promoters during fruit development and in fruit tissues was confirmed in transgenic tomato lines. Novel Gateway destination vectors were generated for the transcription of artificial microRNA (amiRNA) precursors and hairpin RNAs under the control of these promoters, with schemes only involving Gateway BP and LR Clonase reactions. Efficient silencing of the endogenous phytoene desaturase gene was demonstrated in transgenic tomato lines producing a matching amiRNA under the cauliflower mosaic virus 35S or PPC2 promoter. Lastly, taking advantage of the pOP/LhG4 two-component system, we found that well-characterized flower-specific Arabidopsis promoters drive the expression of reporters in patterns generally compatible with heterologous expression. Tomato lines and plasmids will be distributed through a new Nottingham Arabidopsis Stock Centre service unit dedicated to Solanaceae resources.

The physiological disorder hyperhydricity occurs frequently in tissue culture and causes several morphological abnormalities such as thick, brittle, curled, and translucent leaves. It is well known that hyperhydric shoots are characterized by a high water content, but how this is related to the abnormalities is not clear. It was observed that water accumulated extensively in the apoplast of leaves of hyperhydric Arabidopsis seedlings and flooded apoplastic air spaces almost completely. In hyperhydric Arabidopsis seedlings, the volume of apoplastic air was reduced from 85% of the apoplast to only 15%. Similar results were obtained with hyperhydric shoots of statice. The elevated expression of hypoxia-responsive genes in hyperhydric seedlings showed that the water saturation of the apoplast decreased oxygen supply. This demonstrates a reduced gas exchange between the symplast and its surroundings, which will consequently lead to the accumulation of gases in the symplast, for example ethylene and methyl jasmonate. The impairment of gas exchange probably brings about the symptoms of hyperhydricity. Interestingly, stomatal aperture was reduced in hyperhydric plants, a previously reported response to injection of water into the apoplast. Closure of the stomata and the accumulation of water in the apoplast may be the reasons why seedlings with a low level of hyperhydricity showed improved acclimatization after planting into soil.

Growth of tomato fruits is determined by cell division and cell expansion, which are tightly controlled by factors that drive the core cell cycle. The cyclin-dependent kinases (CDKs) and their interacting partners, the cyclins, play a key role in the progression of the cell cycle. In this study the role of CDKA1, CDKB1, and CDKB2 in fruit development was characterized by fruit-specific overexpression and down-regulation. CDKA1 is expressed in the pericarp throughout development, but is strongly up-regulated in the outer pericarp cell layers at the end of the growth period, when CDKB gene expression has ceased. Overexpression of the CDKB genes at later stages of development and the down-regulation of CDKA1 result in a very similar fruit phenotype, showing a reduction in the number of cell layers in the pericarp and alterations in the desiccation of the fruits. Expression studies revealed that CDKA1 is down-regulated by the expression of CDKB1/2 in CDKB1 and CDKB2 overexpression mutants, suggesting opposite roles for these types of CDK proteins in tomato pericarp development.