Changes in the nuclear proteome of developing wheat (Triticum aestivum L.) grain

Wheat grain end-use value is determined by complex molecular interactions that occur during grain development, including those in the cell nucleus. However, our knowledge of how the nuclear proteome changes during grain development is limited. Here, we analyzed nuclear proteins of developing wheat grains collected during the cellularization, effective grain-filling, and maturation phases of development, respectively. Nuclear proteins were extracted and separated by two-dimensional gel electrophoresis. Image analysis revealed 371 and 299 reproducible spots in gels with first dimension separation along pH 4-7 and pH 6-11 isoelectric gradients, respectively. The relative abundance of 464 (67%) protein spots changed during grain development. Abundance profiles of these proteins clustered in six groups associated with the major phases and phase transitions of grain development. Using nano liquid chromatography tandem mass spectrometry to analyse 387 variant and non-variant protein spots, 114 different proteins were identified that were classified into 16 functional classes. We noted that some proteins involved in the regulation of transcription, like HMG1/2-like protein and histone deacetylase HDAC2, were most abundant before the phase transition from cellularization to grain-filling, suggesting that major transcriptional changes occur during this key developmental phase. The maturation period was characterized by high relative abundance of proteins involved in ribosome biogenesis. Data are available via ProteomeXchange with identifier PXD002999

Reference gene selection for head and neck squamous cell carcinoma gene expression studies

<p>Abstract</p> <p>Background</p> <p>It is no longer adequate to choose reference genes blindly. We present the first study that defines the suitability of 12 reference genes commonly used in cancer studies (<it>ACT, ALAS, B2M, GAPDH, HMBS, HPRT, KALPHA, RPS18, RPL27, RPS29, SHAD </it>and <it>TBP</it>) for the normalization of quantitative expression data in the field of head and neck squamous cell carcinoma (HNSCC).</p> <p>Results</p> <p>Raw expression levels were measured by RT-qPCR in HNSCC and normal matched mucosa of 46 patients. We analyzed the expression stability using geNorm and NormFinder and compared the expression levels between subgroups. In HNSCC and/or normal mucosa, the four best normalization genes were <it>ALAS, GAPDH, RPS18 </it>and <it>SHAD </it>and the most stable combination of two genes was <it>GAPDH-SHAD</it>. We recommend using <it>KALPHA-TBP </it>for the study of T1-T2 tumors, <it>RPL27-SHAD </it>for T3-T4 tumors, <it>KALPHA-SHAD </it>for N0 tumors, and <it>ALAS-TBP </it>for N+ tumors. <it>ACT, B2M, GAPDH, HMBS, HPRT, KALPHA, RPS18, RPS29, SHAD </it>and <it>TBP </it>were slightly misregulated (<1.7-fold) between tumor and normal mucosa but can be used for normalization, depending on the resolution required for the assay.</p> <p>Conclusion</p> <p>In the field of HNSCC, this study will guide researchers in selecting the most appropriate reference genes from among 12 potentially suitable reference genes, depending on the specific setting of their experiments.</p

Intramolecular Cyclization of N-phenyl N'(2-chloroethyl)ureas leads to Active N-phenyl-4,5-dihydrooxazol-2-amines Alkylating β-Tubulin Glu198 and Prohibitin Asp40

International audienceThe cyclization of anticancer drugs into active intermediates has been reported mainly for DNA alkylating molecules including nitrosoureas. We previously defined the original cytotoxic mechanism of anticancerous phenyl '(2-chloroethyl)ureas (CEUs) that involves their reactivity towards cellular proteins and not against DNA; two CEUs subsets have been shown to alkylate β-tubulin and prohibitin leading to inhibition of cell proliferation by G/M or G/S cell cycle arrest. In this study, we demonstrated that cyclic derivatives of CEUs, -phenyl-4,5-dihydrooxazol-2-amines (Oxas) are two to threefold more active than CEUs and share the same cytotoxic properties in B16F0 melanoma cells. Moreover, the CEU original covalent binding by an ester linkage on β-tubulin Glu198 and prohibitin Asp40 was maintained with Oxas. Surprisingly, we observed that Oxas were spontaneously formed from CEUs in the cell culture medium and were also detected within the cells. Our results suggest that the intramolecular cyclization of CEUs leads to active Oxas that should then be considered as the key intermediates for protein alkylation. These results could be useful for the design of new prodrugs for cancer chemotherapy

Gestion des appels d'urgence routiers : contexte et perspectives d'évolution

Face à la diversification des moyens d'appels à disposition des usagers de la route en détresse, se pose la question de l'évolution du RAU (réseau d'appels d'urgence), équipement routier dédié aux urgences. Pour l'évaluer ce document présente :Le diagnostic de fonctionnement du RAU et des appels d'urgence au travers des aspects techniques(RAU, téléphonie mobile. . .), de l'organisation des centres d'appels d'urgence, de l'acheminements des appels, d'enquêtes auprès des acteurs et des usagers.Le fonctionnement dans quatre pays européens, synthèse d'une étude faite en Allemagne,Grande-Bretagne, Italie, Hollande.Une corrélation accidents, appels d'urgence : croisement géographique de l'implantation des PAU sur RN avec des données accidents.Des perspectives d'évolutions sur le plan technique, sur l'organisation et l'acheminement des appels.Une synthèse de propositions et de suites à donner

Impact of tannin supplementation on proteolysis during post-ruminal digestion in wethers using a dynamic in vitro system: a plant (Medicago sativa) digestomic approach

The aim of this study was to characterize the effects of tannins on plant protein during sheep digestion, using a digestomic approach combining in vivo (rumen) conditions and an in vitro digestive system (abomasum and small intestine). Ruminal fluid from wethers infused with a tannin solution or water (control) was introduced into the digester, and protein degradation was followed by LC-MS/MS. Tannin infusion in the rumen led to a clear decrease in protein degradation-related fermentation end-products, whereas RuBisCo protein was more abundant than in control wethers. In the simulated abomasum, peptidomic analysis showed more degradation products of RuBisCo in the presence of tannins. The effect of RuBisCo protection by tannins continued to impact Rubisco digestion into early-stage intestinal digestion, but was no longer detectable in late-stage intestinal digestion. The peptidomics approach proved a potent tool for identifying and quantifying the type of protein hydrolyzed throughout the gastrointestinal tract

Long-distance frequency dissemination with a resolution of 10-17

We use a new technique to disseminate microwave reference signals along ordinary optical fiber. The fractional frequency resolution of a link of 86 km in length is 10-17 for a one day integration time, a resolution higher than the stability of the best microwave or optical clocks. We use the link to compare the microwave reference and a CO2/OsO4 frequency standard that stabilizes a femtosecond laser frequency comb. This demonstrates a resolution of 3.10-14 at 1 s. An upper value of the instability introduced by the femtosecond laser-based synthesizer is estimated as 1.10-14 at 1 s

Dystrophy-associated caveolin-3 mutations reveal that caveolae couple IL6/STAT3 signaling with mechanosensing in human muscle cells

Caveolin-3 is the major structural protein of caveolae in muscle. Mutations in the CAV3 gene cause different types of myopathies with altered membrane integrity and repair, expression of muscle proteins, and regulation of signaling pathways. We show here that myotubes from patients bearing the CAV3P28L and R26Q mutations present a dramatic decrease of caveolae at the plasma membrane, resulting in abnormal response to mechanical stress. Mutant myotubes are unable to buffer the increase in membrane tension induced by mechanical stress. This results in impaired regulation of the IL6/STAT3 signaling pathway leading to its constitutive hyperactivation and increased expression of muscle genes. These defects are fully reversed by reassembling functional caveolae through expression ofcaveolin-3. Our study reveals that under mechanical stress the regulation of mechan-oprotection by caveolae is directly coupled with the regulation of IL6/STAT3 signaling inmuscle cells and that this regulation is absent in Cav3-associated dystrophic patients

Small-scale study of Debris-Flows Interactions with a Lateral Debris Basin and Crossings: The Manival Torrent case study

Small-scale models are useful tools to study the interactions between debris flows and structures and channels. Small-scale modelling of debris flows remains however complicated because of the complex rheology and scaling challenges of these geophysical processes. An on-going study of a debris basin and the downstream channel where two fords and a bridge are located is presented in this extended abstract. The studied torrent is the Manival catchment, located near Grenoble in France. We present the catchment features, the scientific questions studied, some preliminary calibration results describing the mixtures used to model debris flows as well as results from three debris-flood and two debris-flow runs. In essence, the model highlighted that the structure enable a large share of the bedload transport to pass downstream. Debris flows can be more or equally trapped depending on their rheology which controls the surges dynamics and the deposition slope in the debris basin

The p300/CBP-associated factor (PCAF) is a cofactor of ATF4 for amino acid-regulated transcription of CHOP

When an essential amino acid is limited, a signaling cascade is triggered that leads to increased translation of the ‘master regulator’, activating transcription factor 4 (ATF4), and resulting in the induction of specific target genes. Binding of ATF4 to the amino acid response element (AARE) is an essential step in the transcriptional activation of CHOP (a CCAAT/enhancer-binding protein-related gene) by amino acid deprivation. We set out to identify proteins that interact with ATF4 and that play a role in the transcriptional activation of CHOP. Using a tandem affinity purification (TAP) tag approach, we identified p300/CBP-associated factor (PCAF) as a novel interaction partner of ATF4 in leucine-starved cells. We show that the N-terminal region of ATF4 is required for a direct interaction with PCAF and demonstrate that PCAF is involved in the full transcriptional response of CHOP by amino acid starvation. Chromatin immunoprecipitation analysis revealed that PCAF is engaged on the CHOP AARE in response to amino acid starvation and that ATF4 is essential for its recruitment. We also show that PCAF stimulates ATF4-driven transcription via its histone acetyltransferase domain. Thus PCAF acts as a coactivator of ATF4 and is involved in the enhancement of CHOP transcription following amino acid starvation

EHD2 is a mechanotransducer connecting caveolae dynamics with gene transcription

Caveolae are small invaginated pits that function as dynamic mechanosensors to buffer tension variations at the plasma membrane. Here we show that under mechanical stress, the EHD2 ATPase is rapidly released from caveolae, SUMOylated, and translocated to the nucleus, where it regulates the transcription of several genes including those coding for caveolae constituents. We also found that EHD2 is required to maintain the caveolae reservoir at the plasma membrane during the variations of membrane tension induced by mechanical stress. Metal-replica electron microscopy of breast cancer cells lacking EHD2 revealed a complete absence of caveolae and a lack of gene regulation under mechanical stress. Expressing EHD2 was sufficient to restore both functions in these cells. Our findings therefore define EHD2 as a central player in mechanotransduction connecting the disassembly of the caveolae reservoir with the regulation of gene transcription under mechanical stress