Christian Chervin

Chromoplast Differentiation: Current Status and Perspectives

Chromoplasts are carotenoid-accumulating plastids conferring color to many flowers and fruits as well as to some tubers and roots. Chromoplast differentiation proceeds from preexisting plastids, most often chloroplasts. One of the most prominent changes is remodeling of the internal membrane system associated with the formation of carotenoid-accumulating structures. During the differentiation process the plastid genome is essentially stable and transcriptional activity is restricted. The buildup of the chromoplast for specific metabolic characteristics is essentially dependent upon the transcriptional activity of the nucleus. Important progress has been made in terms of mediation of the chloroplast-to-chromoplast transition with the discovery of the crucial role of the Or gene. In this article we review recent developments in the structural, biochemical and molecular aspects of chromoplast differentiation and also consider the reverse differentiation of chromoplasts into chloroplast-like structures during the regreening process occurring in some fruit. Future perspectives toward a full understanding of chromoplast differentiation include in-depth knowledge of the changes occurring in the plastidial proteome during chromoplastogenesis, elucidation of the role of hormones and the search for signals that govern the dialog between the nuclear and the chromoplastic genome.

SlARF4, an Auxin Response Factor Involved in the Control of Sugar Metabolism during Tomato Fruit Development

Tomato fruit development is subject to connections between auxin signaling, chloroplastic activity, and sugar metabolism. Successful completion of fruit developmental programs depends on the interplay between multiple phytohormones. However, besides ethylene, the impact of other hormones on fruit quality traits remains elusive. A previous study has shown that down-regulation of SlARF4, a member of the tomato (Solanum lycopersicum) auxin response factor (ARF) gene family, results in a dark-green fruit phenotype with increased chloroplasts (Jones et al., 2002). This study further examines the role of this auxin transcriptional regulator during tomato fruit development at the level of transcripts, enzyme activities, and metabolites. It is noteworthy that the dark-green phenotype of antisense SlARF4-suppressed lines is restricted to fruit, suggesting that SlARF4 controls chlorophyll accumulation specifically in this organ. The SlARF4 underexpressing lines accumulate more starch at early stages of fruit development and display enhanced chlorophyll content and photochemical efficiency, which is consistent with the idea that fruit photosynthetic activity accounts for the elevated starch levels. SlARF4 expression is high in pericarp tissues of immature fruit and then undergoes a dramatic decline at the onset of ripening concomitant with the increase in sugar content. The higher starch content in developing fruits of SlARF4 down-regulated lines correlates with the up-regulation of genes and enzyme activities involved in starch biosynthesis, suggesting their negative regulation by SlARF4. Altogether, the data uncover the involvement of ARFs in the control of sugar content, an essential feature of fruit quality, and provide insight into the link between auxin signaling, chloroplastic activity, and sugar metabolism in developing fruit.

Ethylene Control of Fruit Ripening: Revisiting the Complex Network of Transcriptional Regulation

The plant hormone ethylene controls fruit ripening through a complex network of transcriptional regulations and interplay between multiple signaling pathways. The plant hormone ethylene plays a key role in climacteric fruit ripening. Studies on components of ethylene signaling have revealed a linear transduction pathway leading to the activation of ethylene response factors. However, the means by which ethylene selects the ripening-related genes and interacts with other signaling pathways to regulate the ripening process are still to be elucidated. Using tomato (Solanum lycopersicum) as a reference species, the present review aims to revisit the mechanisms by which ethylene regulates fruit ripening by taking advantage of new tools available to perform in silico studies at the genome-wide scale, leading to a global view on the expression pattern of ethylene biosynthesis and response genes throughout ripening. Overall, it provides new insights on the transcriptional network by which this hormone coordinates the ripening process and emphasizes the interplay between ethylene and ripening-associated developmental factors and the link between epigenetic regulation and ethylene during fruit ripening.

Chloroplast to chromoplast transition in tomato fruit: spectral confocal microscopy analyses of carotenoids and chlorophylls in isolated plastids and time-lapse recording on intact live tissue

BACKGROUND AND AIMS There are several studies suggesting that tomato (Solanum lycopersicum) chromoplasts arise from chloroplasts, but there is still no report showing the fluorescence of both chlorophylls and carotenoids in an intermediate plastid, and no video showing this transition phase. METHODS Pigment fluorescence within individual plastids, isolated from tomato fruit using sucrose gradients, was observed at different ripening stages, and an in situ real-time recording of pigment fluorescence was performed on live tomato fruit slices. KEY RESULTS At the mature green and red stages, homogenous fractions of chloroplasts and chromoplasts were obtained, respectively. At the breaker stage, spectral confocal microscopy showed that intermediate plastids contained both chlorophylls and carotenoids. Furthermore, an in situ real-time recording (a) showed that the chloroplast to chromoplast transition was synchronous for all plastids of a single cell; and (b) confirmed that all chromoplasts derived from pre-existing chloroplasts. CONCLUSIONS These results give details of the early steps of tomato chromoplast biogenesis from chloroplasts, with the formation of intermediate plastids containing both carotenoids and chlorophylls. They provide information at the sub-cellular level on the synchronism of plastid transition and pigment changes.

Carotenoid accumulation during tomato fruit ripening is modulated by the auxin-ethylene balance

BackgroundTomato fruit ripening is controlled by ethylene and is characterized by a shift in color from green to red, a strong accumulation of lycopene, and a decrease in β-xanthophylls and chlorophylls. The role of other hormones, such as auxin, has been less studied. Auxin is retarding the fruit ripening. In tomato, there is no study of the carotenoid content and related transcript after treatment with auxin.ResultsWe followed the effects of application of various hormone-like substances to “Mature-Green” fruits. Application of an ethylene precursor (ACC) or of an auxin antagonist (PCIB) to tomato fruits accelerated the color shift, the accumulation of lycopene, α-, β-, and δ-carotenes and the disappearance of β-xanthophylls and chlorophyll b. By contrast, application of auxin (IAA) delayed the color shift, the lycopene accumulation and the decrease of chlorophyll a. Combined application of IAA + ACC led to an intermediate phenotype. The levels of transcripts coding for carotenoid biosynthesis enzymes, for the ripening regulator Rin, for chlorophyllase, and the levels of ethylene and abscisic acid (ABA) were monitored in the treated fruits. Correlation network analyses suggest that ABA, may also be a key regulator of several responses to auxin and ethylene treatments.ConclusionsThe results suggest that IAA retards tomato ripening by affecting a set of (i) key regulators, such as Rin, ethylene and ABA, and (ii) key effectors, such as genes for lycopene and β-xanthophyll biosynthesis and for chlorophyll degradation.

Influence of low oxygen storage on aroma compounds of whole pears and crushed pear flesh.

Controlled atmosphere storage is known to decrease pome fruit aroma. Here results are presented showing that low O2 storage (3 kPa) for 2 months reduced character impact compounds of ‘Packham’s Triumph’ pears, namely methyl and ethyl decadienoates, during subsequent ripening. This reduction was detected in both whole pears and crushed pear flesh, used here to approximately reproduce mastication. The relative abundance of more than 20 volatile compounds is presented. Analyses after grouping the results by alcohol or acyl moiety suggest that the synthesis of the decadienoate moiety was more depressed than the methyl or ethyl moieties. Other esters were less abundant after low O2 storage, particularly various acetates (butyl, hexyl, heptyl). Their reduction was particularly revealed in crushed flesh which had been incubated at 40°C. Phenyl ethanol and phenylethyl acetate levels were also reduced after low O2 storage.

Ethanol triggers grape gene expression leading to anthocyanin accumulation during berry ripening

Recent studies have shown that low doses of ethanol stimulate the maturation of some fruits. The present work showed that spraying Cabernet Sauvignon grapes, with 5% ethanol at veraison enhances the anthocyanin accumulation. Veraison is the time when the berries turn from green to purple. HPLC analysis showed a marked increase in the total concentrations of the derivatives of delphinidin, cyanidin, petunidin, peonidin and malvidin from the fourth day after the ethanol treatment until harvest. This was not linked to a difference in berry weight in comparison to controls. Two distinct expression patterns were found for anthocyanin biosynthesis genes in the treated and untreated berries. For one group, consisting of chalcone synthase, flavanone-3-hydroxylase, dihydroxyflavonol-4-reductase and leucoanthocyanidin dioxygenase, the expression was inhibited or unchanged by the ethanol treatment, whereas for UDP glucose-flavonoid 3-O-glucosyltransferase (UFGT) there was a marked increase in expression from 1 to 20 days after ethanol treatment. These results suggest that the UFGT gene is a key factor in the observed anthocyanin accumulation following ethanol treatment.

Could studies on cell responses to low oxygen levels provide improved options for fruit storage and disinfestation

Abstract Cellular responses of harvested horticultural products to reduced partial pressure of oxygen in controlled atmospheres (CA) or modified atmospheres (MA) storage include reduced energy charge, reduced energy availability for repair and maintenance, cytoplasmic acidification, and activation of glycolysis and toxic glycolytic end products. These responses are compared with those of root cells to flooding and of vertebrate and invertebrate animal cells to oxygen deficit. The capacity to sense incipient oxygen stress and, when time, temperature and stress level permit, adapt and so avoid cell damage, is emphasised. The applications of CA and MA technologies could be extended if adaptive changes in the plant products were fostered during the early stages of storage. The use of a combination of heat and oxygen deficit for disinfestation is another possibility. A need for more fundamental studies of energy metabolism in horticultural products in CA is recognised.

Metabolic and Molecular Events Occurring during Chromoplast Biogenesis

Chromoplasts are nonphotosynthetic plastids that accumulate carotenoids. They derive from other plastid forms, mostly chloroplasts. The biochemical events responsible for the interconversion of one plastid form into another are poorly documented. However, thanks to transcriptomics and proteomics approaches, novel information is now available. Data of proteomic and biochemical analysis revealed the importance of lipid metabolism and carotenoids biosynthetic activities. The loss of photosynthetic activity was associated with the absence of the chlorophyll biosynthesis branch and the presence of proteins involved in chlorophyll degradation. Surprisingly, the entire set of Calvin cycle and of the oxidative pentose phosphate pathway persisted after the transition from chloroplast to chromoplast. The role of plastoglobules in the formation and organisation of carotenoid-containing structures and that of the Or gene in the control of chromoplastogenesis are reviewed. Finally, using transcriptomic data, an overview is given the expression pattern of a number of genes encoding plastid-located proteins during tomato fruit ripening.

Predicting the substrate specificity of a glycosyltransferase implicated in the production of phenolic volatiles in tomato fruit

The volatile compounds that constitute the fruit aroma of ripe tomato (Solanum lycopersicum) are often sequestered in glycosylated form. A homology‐based screen was used to identify the gene SlUGT5, which is a member of UDP‐glycosyltransferase 72 family and shows specificity towards a range of substrates, including flavonoid, flavanols, hydroquinone, xenobiotics and chlorinated pollutants. SlUGT5 was shown to be expressed primarily in ripening fruit and flowers, and mapped to chromosome I in a region containing a QTL that affected the content of guaiacol and eugenol in tomato crosses. Recombinant SlUGT5 protein demonstrated significant activity towards guaiacol and eugenol, as well as benzyl alcohol and methyl salicylate; however, the highest in vitro activity and affinity was shown for hydroquinone and salicyl alcohol. NMR analysis identified isosalicin as the only product of salicyl alcohol glycosylation. Protein modelling and substrate docking analysis were used to assess the basis for the substrate specificity of SlUGT5. The analysis correctly predicted the interactions with SlUGT5 substrates, and also indicated that increased hydrogen bonding, due to the presence of a second hydrophilic group in methyl salicylate, guaiacol and hydroquinone, appeared to more favourably anchor these acceptors within the glycosylation site, leading to increased stability, higher activities and higher substrate affinities.