Maryse Laloi

NTR1 ENCODES A HIGH AFFINITY OLIGOPEPTIDE TRANSPORTER IN ARABIDOPSIS

Heterologous complementation of yeast mutants has enabled the isolation of genes encoding several families of amino acid transporters. Among them, NTR1 codes for a membrane protein with weak histidine transport activity. However at the sequence level, NTR1 is related to rather non‐specific oligopeptide transporters from a variety of species including Arabidopsis and to the Arabidopsis nitrate transporter CHL1. A yeast mutant deficient in oligopeptide transport was constructed allowing to show that NTR1 functions as a high affinity, low specificity peptide transporter. In siliques NTR1‐expression is restricted to the embryo, implicating a role in the nourishment of the developing seed.

Source-to-sink transport of sugar and regulation by environmental factors

Source-to-sink transport of sugar is one of the major determinants of plant growth and relies on the efficient and controlled distribution of sucrose (and some other sugars such as raffinose and polyols) across plant organs through the phloem. However, sugar transport through the phloem can be affected by many environmental factors that alter source/sink relationships. In this paper, we summarize current knowledge about the phloem transport mechanisms and review the effects of several abiotic (water and salt stress, mineral deficiency, CO2, light, temperature, air, and soil pollutants) and biotic (mutualistic and pathogenic microbes, viruses, aphids, and parasitic plants) factors. Concerning abiotic constraints, alteration of the distribution of sugar among sinks is often reported, with some sinks as roots favored in case of mineral deficiency. Many of these constraints impair the transport function of the phloem but the exact mechanisms are far from being completely known. Phloem integrity can be disrupted (e.g., by callose deposition) and under certain conditions, phloem transport is affected, earlier than photosynthesis. Photosynthesis inhibition could result from the increase in sugar concentration due to phloem transport decrease. Biotic interactions (aphids, fungi, viruses…) also affect crop plant productivity. Recent breakthroughs have identified some of the sugar transporters involved in these interactions on the host and pathogen sides. The different data are discussed in relation to the phloem transport pathways. When possible, the link with current knowledge on the pathways at the molecular level will be highlighted.

Insights into the Role of Specific Lipids in the Formation and Delivery of Lipid Microdomains to the Plasma Membrane of Plant Cells

The existence of sphingolipid- and sterol-enriched microdomains, known as lipid rafts, in the plasma membrane (PM) of eukaryotic cells is well documented. To obtain more insight into the lipid molecular species required for the formation of microdomains in plants, we have isolated detergent (Triton X-100)-resistant membranes (DRMs) from the PM of Arabidopsis (Arabidopsis thaliana) and leek (Allium porrum) seedlings as well as from Arabidopsis cell cultures. Here, we show that all DRM preparations are enriched in sterols, sterylglucosides, and glucosylceramides (GluCer) and depleted in glycerophospholipids. The GluCer of DRMs from leek seedlings contain hydroxypalmitic acid. We investigated the role of sterols in DRM formation along the secretory pathway in leek seedlings. We present evidence for the presence of DRMs in both the PM and the Golgi apparatus but not in the endoplasmic reticulum. In leek seedlings treated with fenpropimorph, a sterol biosynthesis inhibitor, the usual Δ5-sterols are replaced by 9β,19-cyclopropylsterols. In these plants, sterols and hydroxypalmitic acid-containing GluCer do not reach the PM, and most DRMs are recovered from the Golgi apparatus, indicating that Δ5-sterols and GluCer play a crucial role in lipid microdomain formation and delivery to the PM. In addition, DRM formation in Arabidopsis cells is shown to depend on the unsaturation degree of fatty acyl chains as evidenced by the dramatic decrease in the amount of DRMs prepared from the Arabidopsis mutants, fad2 and Fad3+, affected in their fatty acid desaturases.

The Vitis vinifera sugar transporter gene family: phylogenetic overview and macroarray expression profiling

BackgroundIn higher plants, sugars are not only nutrients but also important signal molecules. They are distributed through the plant via sugar transporters, which are involved not only in sugar long-distance transport via the loading and the unloading of the conducting complex, but also in sugar allocation into source and sink cells. The availability of the recently released grapevine genome sequence offers the opportunity to identify sucrose and monosaccharide transporter gene families in a woody species and to compare them with those of the herbaceous Arabidopsis thaliana using a phylogenetic analysis.ResultsIn grapevine, one of the most economically important fruit crop in the world, it appeared that sucrose and monosaccharide transporter genes are present in 4 and 59 loci, respectively and that the monosaccharide transporter family can be divided into 7 subfamilies. Phylogenetic analysis of protein sequences has indicated that orthologs exist between Vitis and Arabidospis. A search for cis-regulatory elements in the promoter sequences of the most characterized transporter gene families (sucrose, hexoses and polyols transporters), has revealed that some of them might probably be regulated by sugars. To profile several genes simultaneously, we created a macroarray bearing cDNA fragments specific to 20 sugar transporter genes. This macroarray analysis has revealed that two hexose (VvHT1, VvHT3), one polyol (VvPMT5) and one sucrose (VvSUC27) transporter genes, are highly expressed in most vegetative organs. The expression of one hexose transporter (VvHT2) and two tonoplastic monosaccharide transporter (VvTMT1, VvTMT2) genes are regulated during berry development. Finally, three putative hexose transporter genes show a preferential organ specificity being highly expressed in seeds (VvHT3, VvHT5), in roots (VvHT2) or in mature leaves (VvHT5).ConclusionsThis study provides an exhaustive survey of sugar transporter genes in Vitis vinifera and revealed that sugar transporter gene families in this woody plant are strongly comparable to those of herbaceous species. Dedicated macroarrays have provided a Vitis sugar transporter genes expression profiling, which will likely contribute to understand their physiological functions in plant and berry development. The present results might also have a significant impact on our knowledge on plant sugar transporters.

Interaction of grape ASR proteins with a DREB transcription factor in the nucleus.

MINT‐6743067: VvMSA (uniprotkb:Q94G23) and VvDREB (uniprotkb:A6XA90) physically interact (MI:0218) by bimolecular fluorescence complementation (MI:0809) MINT‐6743043: VvMSA (uniprotkb:Q94G23) physically interacts (MI:0218) with VvDREB (uniprotkb:A6XA90) by two hybrid (MI:0018)

Regulation of RhSUC2, a sucrose transporter, is correlated with the light control of bud burst in Rosa sp.

In roses, light is a central environmental factor controlling bud break and involves a stimulation of sugar metabolism. Very little is known about the role of sucrose transporters in the bud break process and its regulation by light. In this study, we show that sugar promotes rose bud break and that bud break is accompanied by an import of sucrose. Radio-labelled sucrose accumulation is higher in buds exposed to light than to darkness and involves an active component. Several sucrose transporter (RhSUC1, 2, 3 and 4) transcripts are expressed in rose tissues, but RhSUC2 transcript level is the only one induced in buds exposed to light after removing the apical dominance. RhSUC2 is preferentially expressed in bursting buds and stems. Functional analyses in bakers yeast demonstrate that RhSUC2 encodes a sucrose/proton co-transporter with a K(m) value of 2.99 mm at pH 4.5 and shows typical features of sucrose symporters. We therefore propose that bud break photocontrol partly depends upon the modulation of sucrose import into buds by RhSUC2.

Dissection of the transcriptional regulation of grape ASR and response to glucose and abscisic acid

Despite the fact that the precise physiological function of ASRs [abscisic acid (ABA), stress, ripening] remains unknown, they have been suggested to play a dual role in the plant response to environmental cues, as highly hydrophilic proteins for direct protection, as well as transcription factors involved in the regulation of gene expression. To investigate further the biological positioning of grape ASR in the hormonal and metabolic signal network, three promoters corresponding to its cDNA were isolated and submited to a detailed in silico and functional analysis. The results obtained provided evidence for the allelic polymorphism of the grape ASR gene, the organ-preferential expression conferred on the GUS reporter gene, and the specific phloem tissue localization revealed by in situ hybridization. The study of glucose and ABA signalling in its transcriptional control, by transfection of grape protoplasts using the dual luciferase system, revealed the complexity of ASR gene expression regulation. A model was proposed allowing a discussion of the place of ASR in the fine tuning of hormonal and metabolic signalling involved in the integration of environmental cues by the plant organism.

Links between lipid homeostasis, organelle morphodynamics and protein trafficking in eukaryotic and plant secretory pathways.

The role of lipids as molecular actors of protein transport and organelle morphology in plant cells has progressed over the last years through pharmacological and genetic investigations. The manuscript is reviewing the roles of various lipid families in membrane dynamics and trafficking in eukaryotic cells, and summarizes some of the related physicochemical properties of the lipids involved. The article also focuses on the specific requirements of the sphingolipid glucosylceramide (GlcCer) in Golgi morphology and protein transport through the plant secretory pathway. The use of a specific inhibitor of plant glucosylceramide synthase and selected Arabidopsis thaliana RNAi lines stably expressing several markers of the plant secretory pathway, establishes specific steps sensitive to GlcCer biosynthesis. Collectively, data of the literature demonstrate the existence of links between protein trafficking, organelle morphology, and lipid metabolism/homeostasis in eukaryotic cells including plant cells.

Profiling of sugar transporter genes in grapevine coping with water deficit.

The profiling of grapevine (Vitis vinifera L.) genes under water deficit was specifically targeted to sugar transporters. Leaf water status was characterized by physiological parameters and soluble sugars content. The expression analysis provided evidence that VvHT1 hexose transporter gene was strongly down‐regulated by the increased sugar content under mild water‐deficit. The genes of monosaccharide transporter VvHT5, sucrose carrier VvSUC11, vacuolar invertase VvGIN2 and grape ASR (ABA, stress, ripening) were up‐regulated under severe water stress. Their regulation in a drought‐ABA signalling network and possible roles in complex interdependence between sugar subcellular partitioning and cell influx/efflux under Grapevine acclimation to dehydration are discussed.

Characterization of leucine-leucine transport in leaf tissues

Uptake of the dipeptide [(3)H]Leu-Leu into leaf discs from mature broad bean (Vicia faba L.) was characterized. Uptake was maximal at pH 6.0 and appeared to be mediated by three systems with apparent K(m) values of 20 µM, 350 µM and 43 mM, respectively. Leu-Leu uptake was sensitive to N-ethylmaleimide, p-chloromercuribenzenesulphonic acid, diethylpyrocarbonate, and carbonyl-cyanide-m-chlorophenylhydrazone. Nitrate did not compete with peptide uptake, although the peptide transporter and the nitrate transporter have been reported to be homologous. The ability of leaf tissues to take up peptides strongly decreased with leaf age, and the phloem export of peptides as measured by exudation experiments was very low. It is concluded that the leaf tissues contain a peptide transporter that may take up some peptides with a high affinity, but that this transporter is not involved in the long-distance transport of nitrogen under the form of di- or tri-peptides.