Physiological Functions of the COPI Complex in Higher Plants

COPI vesicles are essential to the retrograde transport of proteins in the early secretory pathway. The COPI coatomer complex consists of seven subunits, termed alpha-, beta-, beta'-, gamma-, delta-, epsilon-, and zeta-COP, in yeast and mammals. Plant genomes have homologs of these subunits, but the essentiality of their cellular functions has hampered the functional characterization of the subunit genes in plants. Here we have employed virus-induced gene silencing (VIGS) and dexamethasone (DEX)-inducible RNAI of the COPI subunit genes to study the in vivo functions of the COPI coatomer complex in plants. The beta'-, gamma-, and delta-COP subunits localized to the Golgi as GFP-fusion proteins and interacted with each other in the Golgi. Silencing of beta'-, gamma-, and delta-COP by VIGS resulted in growth arrest and acute plant death in Nicotiana benthamiana, with the affected leaf cells exhibiting morphological markers of programmed cell death. Depletion of the COPI subunits resulted in disruption of the Golgi structure and accumulation of autolysosome-like structures in earlier stages of gene silencing. In tobacco BY-2 cells, DEX-inducible RNAi of beta'-COP caused aberrant cell plate formation during cytokinesis. Collectively, these results suggest that COPI vesicles are essential to plant growth and survival by maintaining the Golgi apparatus and modulating cell plate formation.1196Ysciescopu

Sphingosine mediates FTY720-induced apoptosis in LLC-PK1 cells

FTY720, a synthetic sphingoid base analog, was examined as a new sphingosine kinase inhibitor, which converts endogenous sphingosine into its phosphate form. With 20 ??M of FTY720, sphingosine accumulated in the LLC-PK1 cells in a time- and dose-dependent manner. The FTY720 treated cells showed a high concentration of fragmented DNA, a high caspase-3 like activity and TUNEL staining cells. It was also found that the sphingosine and sphinganine level increased in a time- and dose-dependent manner within 12 h after the FTY720 treatment. The sphingosine kinase activity was reduced by FTY720 as much as other sphingosine kinase inhibitors, N, N-dimethylsphingosine (DMS), dl-threo-dihydrosphingosine (DHS). The fragmented DNA content as a result of the 20 ??M of FTY720 treatment and by 5 ??M of the exogenously added BSA-sphingosine complex indicated typical apoptosis. Under similar conditions, the accumulated sphingosine concentration in all the cells was almost identical even though the sphingosine distribution inside the cells was somewhat different. These results indicate that the FTY720 induced apoptosis is associated with the inhibition of the sphingosine kinase activity and is strongly associated with the successive accumulation of sphingosine.open172

The Fear-Avoidance Model of Chronic Pain: Assessing the Role of Neuroticism and Negative Affect in Pain Catastrophizing Using Structural Equation Modeling

postprin

Crystal Structure of the Rad3/XPD regulatory domain of Ssl1/p44

The Ssl1/p44 subunit is a core component of the yeast/mammalian general transcription factor TFIIH, which is involved in transcription and DNA repair. Ssl1/p44 binds to and stimulates the Rad3/XPD helicase activity of TFIIH. To understand the helicase stimulatory mechanism of Ssl1/p44, we determined the crystal structure of the N-terminal regulatory domain of Ssl1 from Saccharomyces cerevisiae. Ssl1 forms a von Willebrand factor A fold in which a central six-stranded beta-sheet is sandwiched between three alpha helices on both sides. Structural and biochemical analyses of Ssl1/p44 revealed that the beta 4-alpha 5 loop, which is frequently found at the interface between von Willebrand factor A family proteins and cellular counterparts, is critical for the stimulation of Rad3/XPD. Yeast genetics analyses showed that double mutation of Leu-239 and Ser-240 in the beta 4-alpha 5 loop of Ssl1 leads to lethality of a yeast strain, demonstrating the importance of the Rad3-Ssl1 interactions to cell viability. Here, we provide a structural model for the Rad3/XPD-Ssl1/p44 complex and insights into how the binding of Ssl1/p44 contributes to the helicase activity of Rad3/XPD and cell viability.X1165Ysciescopu

Molecular lens applied to benzene and carbon disulfide molecular beams

A molecular lens of the nonresonant dipole force formed by focusing a nanosecond IR laser pulse has been applied to benzene and CS2 molecular beams. Using the velocity map imaging technique for molecular ray tracing, characteristic molecular lens parameters including the focal length (f ), minimum beam width (W), and distance to the minimum beam width position (D) were determined. The laser intensity dependence of the observed lens parameters was in good agreement with theoretical predictions. W was independent of the laser peak intensity (I-0), whereas f and D varied linearly with 1/I-0. The differences in lens parameters between the molecular species were well correlated with the polarizability per mass values of the molecules. A high chromatographic resolution of Rs = 0.84 was achieved between the images of benzene molecular beams undeflected and deflected by the lens. The possibilities for a new type of chromatography are discussed.open293

Paraoxonase 2 protein is spatially expressed in the human placenta and selectively reduced in labour

Humans parturition involves interaction of hormonal, neurological, mechanical stretch and inflammatory pathways and the placenta plays a crucial role. The paraoxonases (PONs 1–3) protect against oxidative damage and lipid peroxidation, modulation of endoplasmic reticulum stress and regulation of apoptosis. Nothing is known about the role of PON2 in the placenta and labour. Since PON2 plays a role in oxidative stress and inflammation, both features of labour, we hypothesised that placental PON2 expression would alter during labour. PON2 was examined in placentas obtained from women who delivered by cesarean section and were not in labour and compared to the equivalent zone of placentas obtained from women who delivered vaginally following an uncomplicated labour. Samples were obtained from 12 sites within each placenta: 4 equally spaced apart pieces were sampled from the inner, middle and outer placental regions. PON2 expression was investigated by Western blotting and real time PCR. Two PON2 forms, one at 62 kDa and one at 43 kDa were found in all samples. No difference in protein expression of either isoform was found between the three sites in either the labour or non-labour group. At the middle site there was a highly significant decrease in PON2 expression in the labour group when compared to the non-labour group for both the 62 kDa form (p = 0.02) and the 43 kDa form (p = 0.006). No spatial differences were found within placentas at the mRNA level in either labour or non-labour. There was, paradoxically, an increase in PON2 mRNA in the labour group at the middle site only. This is the first report to describe changes in PON2 in the placenta in labour. The physiological and pathological significance of these remains to be elucidated but since PON2 is anti-inflammatory further studies are warranted to understand its role

Influence of real-world characteristics on outcomes for patients with methicillin-resistant Staphylococcal skin and soft tissue infections:a multi-country medical chart review in Europe

BACKGROUND: Patient-related (demographic/disease) and treatment-related (drug/clinician/hospital) characteristics were evaluated as potential predictors of healthcare resource use and opportunities for early switch (ES) from intravenous (IV)-to-oral methicillin-resistant Staphylococcus aureus (MRSA)-active antibiotic therapy and early hospital discharge (ED). METHODS: This retrospective observational medical chart study analyzed patients (across 12 European countries) with microbiologically confirmed MRSA complicated skin and soft tissue infections (cSSTI), ≥3 days of IV anti-MRSA antibiotics during hospitalization (July 1, 2010-June 30, 2011), and discharged alive by July 31, 2011. Logistic/linear regression models evaluated characteristics potentially associated with actual resource use (length of IV therapy, length of hospital stay [LOS], IV-to-oral antibiotic switch), and ES and ED (using literature-based and expert-verified criteria) outcomes. RESULTS: 1542 patients (mean ± SD age 60.8 ± 16.5 years; 61.5% males) were assessed with 81.0% hospitalized for MRSA cSSTI as the primary reason. Several patient demographic, infection, complication, treatment, and hospital characteristics were predictive of length of IV therapy, LOS, IV-to-oral antibiotic switch, or ES and ED opportunities. Outcomes and ES and ED opportunities varied across countries. Length of IV therapy and LOS (r = 0.66, p < 0.0001) and eligibilities for ES and ED (r = 0.44, p < 0.0001) showed relatively strong correlations. IV-to-oral antibiotic switch patients had significantly shorter length of IV therapy (−5.19 days, p < 0.001) and non-significantly shorter LOS (−1.86 days, p > 0.05). Certain patient and treatment characteristics were associated with increased odds of ES (healthcare-associated/ hospital-acquired infection) and ED (patient living arrangements, healthcare-associated/ hospital-acquired infection, initiating MRSA-active treatment 1–2 days post cSSTI index date, existing ED protocol), while other factors decreased the odds of ES (no documented MRSA culture, ≥4 days from admission to cSSTI index date, IV-to-oral switch, IV line infection) and ED (dementia, no documented MRSA culture, initiating MRSA-active treatment ≥3 days post cSSTI index date, existing ES protocol). CONCLUSIONS: Practice patterns and opportunity for further ES and ED were affected by several infection, treatment, hospital, and geographical characteristics, which should be considered in identifying ES and ED opportunities and designing interventions for MRSA cSSTI to reduce IV days and LOS while maintaining the quality of care. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/1471-2334-14-476) contains supplementary material, which is available to authorized users

Site-specific incorporation of phosphotyrosine using an expanded genetic code.

Access to phosphoproteins with stoichiometric and site-specific phosphorylation status is key to understanding the role of protein phosphorylation. Here we report an efficient method to generate pure, active phosphotyrosine-containing proteins by genetically encoding a stable phosphotyrosine analog that is convertible to native phosphotyrosine. We demonstrate its general compatibility with proteins of various sizes, phosphotyrosine sites and functions, and reveal a possible role of tyrosine phosphorylation in negative regulation of ubiquitination

Relationships between CYP2D6 phenotype, breast cancer and hot flushes in women at high risk of breast cancer receiving prophylactic tamoxifen: results from the IBIS-I trial

Licensed under a Creative Commons Attribution Non-Commercial Share Alike Licens