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1.
New Phytol ; 214(1): 233-244, 2017 Apr.
Article in English | MEDLINE | ID: mdl-27935038

ABSTRACT

This work investigates the extent of translational regulation during seed germination. The polysome occupancy of each gene is determined by genome-wide profiling of total mRNA and polysome-associated mRNA. This reveals extensive translational regulation during Arabidopsis thaliana seed germination. The polysome occupancy of thousands of individual mRNAs changes to a large extent during the germination process. Intriguingly, these changes are restricted to two temporal phases (shifts) during germination, seed hydration and germination. Sequence features, such as upstream open reading frame number, transcript length, mRNA stability, secondary structures, and the presence and location of specific motifs correlated with this translational regulation. These features differed significantly between the two shifts, indicating that independent mechanisms regulate translation during seed germination. This study reveals substantial translational dynamics during seed germination and identifies development-dependent sequence features and cis elements that correlate with the translation control, uncovering a novel and important layer of gene regulation during seed germination.


Subject(s)
Arabidopsis/embryology , Arabidopsis/genetics , Germination/genetics , Polyribosomes/metabolism , Protein Biosynthesis/genetics , Seeds/genetics , Base Sequence , Gene Expression Regulation, Plant , Nucleic Acid Conformation , RNA, Messenger/chemistry , RNA, Messenger/genetics , RNA, Messenger/metabolism , RNA, Plant/chemistry , RNA, Plant/genetics , RNA, Plant/metabolism
2.
Mol Plant Microbe Interact ; 28(11): 1227-36, 2015 Nov.
Article in English | MEDLINE | ID: mdl-26106900

ABSTRACT

Stomata remain abnormally opened and unresponsive to abscisic acid in grapevine leaves infected by downy mildew. This deregulation occurs from 3 days postinoculation and increases concomitantly with leaf colonization by the pathogen. Using epidermal peels, we demonstrated that the active compound involved in this deregulation is located in the apoplast. Biochemical assays showed that the active compound present in the apoplastic fluids isolated from Plasmopara viticola-infected grapevine leaves (IAF) is a CysCys bridge-independent, thermostable and glycosylated protein. Fractionation guided assays based on chromatography coupled to stomatal response and proteomic analysis allowed the identification of both plant and pathogen proteins in the active fraction obtained from IAF. Further in silico analysis and discriminant filtrations based on the comparison between predictions and experimental indications lead to the identification of two Vitis vinifera proteins as candidates for the observed stomatal deregulation.


Subject(s)
Glycoproteins/metabolism , Oomycetes/metabolism , Plant Leaves/metabolism , Plant Proteins/metabolism , Plant Stomata/metabolism , Vitis/metabolism , Amino Acid Sequence , Cell Wall/genetics , Cell Wall/metabolism , Cell Wall/microbiology , Chromatography, Ion Exchange , Computer Simulation , Fungal Proteins/metabolism , Glycoproteins/classification , Glycoproteins/genetics , Host-Pathogen Interactions , Molecular Sequence Data , Oomycetes/physiology , Phylogeny , Plant Diseases/genetics , Plant Diseases/microbiology , Plant Epidermis/genetics , Plant Epidermis/metabolism , Plant Epidermis/microbiology , Plant Leaves/genetics , Plant Leaves/microbiology , Plant Proteins/classification , Plant Proteins/genetics , Plant Stomata/genetics , Plant Stomata/microbiology , Proteomics/methods , Sequence Homology, Amino Acid , Vitis/genetics , Vitis/microbiology
3.
BMC Plant Biol ; 14: 306, 2014 Nov 18.
Article in English | MEDLINE | ID: mdl-25403240

ABSTRACT

BACKGROUND: Protein synthesis is a highly energy demanding process and is regulated according to cellular energy levels. Light and sugar availability affect mRNA translation in plant cells but the specific roles of these factors remain unclear. In this study, sucrose was applied to Arabidopsis seedlings kept in the light or in the dark, in order to distinguish sucrose and light effects on transcription and translation. These were studied using microarray analysis of steady-state mRNA and mRNA bound to translating ribosomes. RESULTS: Steady-state mRNA levels were affected differently by sucrose in the light and in the dark but general translation increased to a similar extent in both conditions. For a majority of the transcripts changes of the transcript levels were followed by changes in polysomal mRNA levels. However, for 243 mRNAs, a change in polysomal occupancy (defined as polysomal levels related to steady-state levels of the mRNA) was observed after sucrose treatment in the light, but not in the dark condition. Many of these mRNAs are annotated as encoding ribosomal proteins, supporting specific translational regulation of this group of transcripts. Unexpectedly, the numbers of ribosomes bound to each mRNA decreased for mRNAs with increased polysomal occupancy. CONCLUSIONS: Our results suggest that sucrose regulate translation of these 243 mRNAs specifically in the light, through a novel regulatory mechanism. Our data shows that increased polysomal occupancy is not necessarily leading to more ribosomes per transcript, suggesting a mechanism of translational induction not solely dependent on increased translation initiation rates.


Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/metabolism , Gene Expression Regulation, Plant , Protein Biosynthesis , Signal Transduction , Sucrose/metabolism , Arabidopsis/genetics , Arabidopsis/radiation effects , Arabidopsis Proteins/genetics , Gene Expression Profiling , Light , Oligonucleotide Array Sequence Analysis , Polyribosomes/metabolism , Ribosomal Proteins/genetics , Ribosomal Proteins/metabolism , Ribosomes/metabolism , Seedlings/genetics , Seedlings/metabolism , Seedlings/radiation effects
4.
Front Plant Sci ; 5: 353, 2014.
Article in English | MEDLINE | ID: mdl-25101105

ABSTRACT

Stress impacts negatively on plant growth and crop productivity, caicultural production worldwide. Throughout their life, plants are often confronted with multiple types of stress that affect overall cellular energy status and activate energy-saving responses. The resulting low energy syndrome (LES) includes transcriptional, translational, and metabolic reprogramming and is essential for stress adaptation. The conserved kinases sucrose-non-fermenting-1-related protein kinase-1 (SnRK1) and target of rapamycin (TOR) play central roles in the regulation of LES in response to stress conditions, affecting cellular processes and leading to growth arrest and metabolic reprogramming. We review the current understanding of how TOR and SnRK1 are involved in regulating the response of plants to low energy conditions. The central role in the regulation of cellular processes, the reprogramming of metabolism, and the phenotypic consequences of these two kinases will be discussed in light of current knowledge and potential future developments.

5.
Mol Plant Microbe Interact ; 24(9): 1061-73, 2011 Sep.
Article in English | MEDLINE | ID: mdl-21649510

ABSTRACT

The oomycete Plasmopara viticola is responsible for downy mildew, a severe grapevine disease. In infected grapevine leaves, we have observed an abnormal starch accumulation at the end of the dark period, suggesting modifications in starch metabolism. Therefore, several complementary approaches, including transcriptomic analyses, measurements of enzyme activities, and sugar quantification, were performed in order to investigate and to understand the effects of P. viticola infection on leaf starch and-to a larger extent-carbohydrate metabolism. Our results indicate that starch accumulation is associated with an increase in ADP-glucose pyrophosphorylase (AGPase) activity and modifications in the starch degradation pathway, especially an increased α-amylase activity. Together with these alterations in starch metabolism, we have observed an accumulation of hexoses, an increase in invertase activity, and a reduction of photosynthesis, indicating a source-to-sink transition in infected leaf tissue. Additionally, we have measured an accumulation of the disaccharide trehalose correlated to an increased trehalase gene expression and enzyme activity. Altogether, these results highlight a dramatic alteration of carbohydrate metabolism correlated with later stages of P. viticola development in leaves.


Subject(s)
Enzymes/metabolism , Oomycetes/growth & development , Plant Diseases/microbiology , Starch/metabolism , Vitis/physiology , Carbohydrate Metabolism , Chlorophyll/metabolism , Enzymes/genetics , Gene Expression Regulation, Plant , Glucose-1-Phosphate Adenylyltransferase/genetics , Glucose-1-Phosphate Adenylyltransferase/metabolism , Hexoses/analysis , Hexoses/metabolism , Oligonucleotide Array Sequence Analysis , Oomycetes/pathogenicity , Photosynthesis/physiology , Plant Leaves/metabolism , Plant Leaves/microbiology , Polysaccharides/analysis , Polysaccharides/metabolism , RNA, Plant/genetics , Starch/analysis , Trehalose/metabolism , Vitis/enzymology , Vitis/genetics , Vitis/microbiology , alpha-Amylases/genetics , alpha-Amylases/metabolism , beta-Amylase/genetics , beta-Amylase/metabolism
6.
Mol Genet Genomics ; 285(4): 273-85, 2011 Apr.
Article in English | MEDLINE | ID: mdl-21340517

ABSTRACT

The recent publication of the grapevine genome sequence facilitates the use of qRT-PCR to study gene expression changes. For this approach, reference genes are commonly used to normalize data and their stability of expression should be systematically validated. Among grapevine defenses is the production of the antimicrobial stilbenic phytoalexins, notably the highly fungitoxic pterostilbene, which plays a crucial role in grapevine interaction with Plasmopara viticola and Botrytis cinerea. As a resveratrol O-methyltransferase (ROMT) gene involved in pterostilbene synthesis was recently identified, we investigated the accumulation of the corresponding transcripts to those of two other stilbene biosynthesis related genes phenylalanine ammonia lyase (PAL) and stilbene synthase (STS) in response to pathogen infection. Using three computer-based statistical methods and C(t) values or LRE method generated values as input data, we have first identified two reference genes (VATP16 and 60SRP) suitable for normalization of qPCR expression data obtained in grapevine leaves and berries infected by P. viticola and B. cinerea, respectively. Next, we have highlighted that the expression of ROMT is induced in P. viticola-infected leaves and also in B. cinerea-infected berries, confirming the involvement of pterostilbene in grapevine defenses.


Subject(s)
Gene Expression Regulation, Plant , Genes, Plant/genetics , Reverse Transcriptase Polymerase Chain Reaction/methods , Reverse Transcriptase Polymerase Chain Reaction/standards , Stilbenes/metabolism , Vitis/genetics , DNA Primers/metabolism , Reference Standards , Reproducibility of Results , Software , Statistics as Topic , Vitis/enzymology
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