Surfing transcriptomic landscapes. A step beyond the annotation of chromosome 16 proteome.

The Spanish team of the Human Proteome Project (SpHPP) marked the annotation of Chr16 and data analysis as one of its priorities. Precise annotation of Chromosome 16 proteins according to C-HPP criteria is presented. Moreover, Human Body Map 2.0 RNA-Seq and Encyclopedia of DNA Elements (ENCODE) data sets were used to obtain further information relative to cell/tissue specific chromosome 16 coding gene expression patterns and to infer the presence of missing proteins. Twenty-four shotgun 2D-LC-MS/MS and gel/LC-MS/MS MIAPE compliant experiments, representing 41% coverage of chromosome 16 proteins, were performed. Furthermore, mapping of large-scale multicenter mass spectrometry data sets from CCD18, MCF7, Jurkat, and Ramos cell lines into RNA-Seq data allowed further insights relative to correlation of chromosome 16 transcripts and proteins. Detection and quantification of chromosome 16 proteins in biological matrices by SRM procedures are also primary goals of the SpHPP. Two strategies were undertaken: one focused on known proteins, taking advantage of MS data already available, and the second, aimed at the detection of the missing proteins, is based on the expression of recombinant proteins to gather MS information and optimize SRM methods that will be used in real biological samples. SRM methods for 49 known proteins and for recombinant forms of 24 missing proteins are reported in this study.

Measuring BVOC emissions released by tomato plants grown in a soilless integrated rooftop greenhouse.

Urban design is currently promoting the inclusion of plants in buildings. However, plants emit biogenic volatile organic compounds (BVOCs), which alone or in combination with other airborne molecules such as CO2, may result in a general increase in tropospheric pollution. Many studies have documented the effects of biotic and abiotic factors on plant BVOC responses, but few have assessed the contribution of typical CO2 levels found in indoor work and meeting spaces. To answer this question, we monitored CO2 and constitutive (MT-limonene) and induced (LOX-cis-3-hexenal) BVOC emissions of a fully developed tomato crop grown hydroponically inside an integrated rooftop greenhouse (i-RTG) in a Mediterranean climate. Two distinctive CO2 assays were performed at the level of the i-RTG by supplying or not CO2. The impact of CO2 on plant physiological emittance was then assessed, and the resulting BVOC rates were compared with reference to EU-LCI values. MT-limonene was ubiquitous among the assays and the most abundant, while LOX-cis-3-hexenal was detected only under controlled CO2 management. The highest levels detected were below the indicated LCIs and were approximately tenfold lower than the corresponding LCI for MT-limonene (50.88 vs. 5000 µg m-3) and eightfold (6.63 µg m-3) higher than the constitutive emission level for LOX-cis-3-hexenal. Over extended sampling (10 min) findings revealed a general emission decrease and significantly different CO2 concentration between the assays. Despite similar decreasing rates of predicted net photosynthesis (Pn) and stomatal conductance (gs) their correlation with decreasing CO2 under uncontrolled condition indirectly suggested a negative CO2 impact on plant emission activity. Conversely, increasing CO2 under the controlled assay showed a positive correlation with induced emissions but not with constitutive ones. Because of significantly higher levels of relative humidity registered under the uncontrolled condition, this factor was considered to affect more than CO2 the emission response and even its collection. This hypothesis was supported by literature findings and attributed to a common issue related with the sampling in static enclosure. Hence, we suggested a careful monitoring of the sampling conditions or further improvements to avoid bias and underestimation of actual emissions. Based on the main outcomes, we observed no evidence of a hazardous effect of registered CO2 rates on the BVOC emissions of tomato plant. Furthermore, because of the low BVOC levels measured in the i-RTG, we assumed as safe the recirculation of this air along building's indoor environments.

Selective extraction of levoglucosan and its isomers from complex matrices using ligand exchange-solid phase extraction for analysis by liquid chromatography-electrospray ionization-tandem mass spectrometry.

The analysis of trace quantities of monosaccharide anhydrides (MAs) in sediments is complicated by the lack of fast and reliable technologies to selectively extract these water-soluble non-ionic compounds from samples of complex composition. Here we describe a solid phase extraction method that takes advantage of the affinity between monosaccharide anhydrides (MAs) and immobilized Na+ ions related to ligand-exchange processes (LE-SPE). The capacity factor of LE-SPE columns was enhanced by using non-aqueous mobile phases such as DCM/MeOH mixtures. We have used the unique properties of LE-SPE columns to selectively extract MAs from lacustrine, coastal, and deep-sea oceanic sediment samples. The analytical procedure produces extracts with low ion suppression effects (0-20%), resulting in ideal conditions for MAs quantification with LC-ESI-MS/MS systems irrespective of the sedimentary matrix and MAs concentration. The analytical method yields repeatable concentration values (RSD of 9-23% for levoglucosan and 15-34% for mannosan and galactosan) and an IS recovery of 45-70%. The instrumental dynamic range is 10-10000 pg injected, but in practice, the methodological lower limit of quantification is constrained by sample contamination during processing. The combination of LE-SPE and LC-ESI-MS/MS has the potential to produce sensitive and reliable technologies to analyze saccharides and amino acids in environmental and biological samples.

Assessment of greenhouse emissions of the green bean through the static enclosure technique.

Urban green installations are extensively promoted to increase sustainable and accessible food production and simultaneously improve the environmental performance and liveability of city buildings. In addition to the multiple benefits of plant retrofitting, these installations may lead to a consistent increase in biogenic volatile organic compounds (BVOCs) in the urban environment, especially indoors. Accordingly, health concerns could limit the implementation of building-integrated agriculture. In a building-integrated rooftop greenhouse (i-RTG), throughout the whole hydroponic cycle, green bean emissions were dynamically collected in a static enclosure. Four representative BVOCs, α-pinene (monoterpene), ß-caryophyllene (sesquiterpene), linalool (oxygenated monoterpene) and cis-3-hexenol (LOX derivate), were investigated in the samples collected from two equivalent sections of a static enclosure, one empty and one occupied by the i-RTG plants, to estimate the volatile emission factor (EF). Throughout the season, extremely variable BVOC levels between 0.04 and 5.36 ppb were found with occasional but not significant (P > 0.05) variations between the two sections. The highest emission rates were observed during plant vegetative development, with EFs equivalent to 78.97, 75.85 and 51.34 ng g-1 h-1 for cis-3-hexenol, α-pinene, and linalool, respectively; at plant maturity, all volatiles were either close to the LLOQ (lowest limit of quantitation) or not detected. Consistent with previous studies significant relationships (r ≥ 0.92; P < 0.05) were individuated within volatiles and temperature and relative humidity of the sections. However, correlations were all negative and were mainly attributed to the relevant effect of the enclosure on the final sampling conditions. Overall, levels found were at least 15 folds lower than the given Risk and LCI values of the EU-LCI protocol for indoor environments, suggesting low BVOC exposure in the i-RTG. Statistical outcomes demonstrated the applicability of the static enclosure technique for fast BVOC emissions survey inside green retrofitted spaces. However, providing high sampling performance over entire BVOCs collection is recommended to reduce sampling error and incorrect estimation of the emissions.

Isotope dilution mass spectrometry for absolute quantification in proteomics: concepts and strategies.

Isotope dilution mass spectrometry is a reference technique for quantitative analysis, given that it combines the sensitivity and selectivity of MS instruments with the precision and accuracy associated with the use of internal standards. Isotope-labeled proteins are the optimal internal standards for quantitative proteomics as they closely mimic the behavior of their natural counterparts during the analytical process. A major complication of isotope dilution mass spectrometry proteomics is the technical difficulty of obtaining these internal standards, especially in studies where a high number of proteins have to be quantified simultaneously. In this paper, we review some of the characteristics of the isotope dilution mass spectrometry approach, its benefits in terms of reliability and quality control in targeted proteomic analysis and the different strategies developed for its application in proteomics.

Differential redox proteomics allows identification of proteins reversibly oxidized at cysteine residues in endothelial cells in response to acute hypoxia.

Adaptation to decreased oxygen availability (hypoxia) is crucial for proper cell function and survival. In metazoans, this is partly achieved through gene transcriptional responses mediated by hypoxia-inducible factors (HIFs). There is abundant evidence that production of reactive oxygen species (ROS) increases during hypoxia, which contributes to the activation of the HIF pathway. In addition to altering the cellular redox balance, leading to oxidative stress, ROS can transduce signals by reversibly modifying the redox state of cysteine residues in certain proteins. Using the "redox fluorescence switch" (RFS), a thiol redox proteomic technique that fluorescently labels reversibly oxidized cysteines, we analyzed endothelial cells subjected to acute hypoxia and subsequent reoxygenation. We observed a general increase in cysteine oxidation during hypoxia, which was reversed by reoxygenation, and two-dimensional electrophoresis revealed the differential oxidation of specific proteins. Using complementary derivatization techniques, we confirmed the modification of individual target proteins and identified specific cysteine residues that were oxidized in hypoxic conditions, thereby overcoming several limitations associated with fluorescence derivatization. These findings provide an important basis for future studies of the role of these modifications in HIF activation and in other acute adaptive responses to hypoxia.