Proteomics, Holm Oak (Quercus ilex L.) and Other Recalcitrant and Orphan Forest Tree Species: How do They See Each Other?

Proteomics has had a big impact on plant biology, considered as a valuable tool for several forest species, such as Quercus, Pines, Poplars, and Eucalyptus. This review assesses the potential and limitations of the proteomics approaches and is focused on Quercus ilex as a model species and other forest tree species. Proteomics has been used with Q. ilex since 2003 with the main aim of examining natural variability, developmental processes, and responses to biotic and abiotic stresses as in other species of the genus Quercus or Pinus. As with the progress in techniques in proteomics in other plant species, the research in Q. ilex moved from 2-DE based strategy to the latest gel-free shotgun workflows. Experimental design, protein extraction, mass spectrometric analysis, confidence levels of qualitative and quantitative proteomics data, and their interpretation are a true challenge with relation to forest tree species due to their extreme orphan and recalcitrant (non-orthodox) nature. Implementing a systems biology approach, it is time to validate proteomics data using complementary techniques and integrate it with the -omics and classical approaches. The full potential of the protein field in plant research is quite far from being entirely exploited. However, despite the methodological limitations present in proteomics, there is no doubt that this discipline has contributed to deeper knowledge of plant biology and, currently, is increasingly employed for translational purposes.

Limited proteomic response in the marine snail Melarhaphe neritoides after long-term emersion.

Rocky intertidal organisms are commonly exposed to environmental gradients, promoting adaptations to these conditions. Emersion time varies along the intertidal range and in the supralittoral zone is frequently larger than a single tidal cycle, even lasting for weeks. The planktonic-dispersing gastropod Melarhaphe neritoides is a common species of the high shore, adapted to reduce water loss in order to survive during long-term emersion. In this study, we investigated the molecular response, at the proteome level, of M. neritoides collected in high-shore tide pools to a series of emersion periods, from 8 to 24 days, in laboratory conditions. We compared this response to individuals maintained submerged during this period, because this was their original habitat. We also included a reversion treatment in the study, in which emersed individuals were returned to the submerged conditions. Although we detected an increase in overall protein concentration with longer emersion periods, contrary to general expectation, the two dimensional electrophoresis (2DE)-based proteomic analysis did not show significant differences between the treatments at the level of individual protein spots, even after an emersion period of 24 days. Our results suggest that the metabolism remains unaltered independent of the treatment carried out or the changes are very subtle and therefore difficult to detect with our experimental design. We conclude that M. neritoides could be equally adapted to emersion and submersion without drastic physiological changes.

Tuber proteome comparison of five potato varieties by principal component analysis.

BACKGROUND: Data analysis of omics data should be performed by multivariate analysis such as principal component analysis (PCA). The way data are clustered in PCA is of major importance to develop some classification systems based on multivariate analysis, such as soft independent modeling of class analogy (SIMCA). In a previous study a one-class classifier based on SIMCA was built using microarray data from a set of potatoes. The PCA grouped the transcriptomic data according to varieties. The present work aimed to use PCA to verify the clustering of the proteomic profiles for the same potato varieties. RESULTS: Proteomic profiles of five potato varieties (Biogold, Fontane, Innovator, Lady Rosetta and Maris Piper) were evaluated by two-dimensional gel electrophoresis (2-DE) performed on two immobilized pH gradient (IPG) strip lengths, 13 and 24 cm, both under pH range 4-7. For each strip length, two gels were prepared from each variety; in total there were ten gels per analysis. For 13 cm strips, 199-320 spots were detected per gel, and for 24 cm strips, 365-684 spots. CONCLUSION: All four PCAs performed with these datasets presented clear grouping of samples according to the varieties. The data presented here showed that PCA was applicable for proteomic analysis of potato and was able to separate the samples by varieties. © 2016 Society of Chemical Industry.

Proteomic and Real-Time PCR analyses of Saccharomyces cerevisiae VL3 exposed to microcystin-LR reveals a set of protein alterations transversal to several eukaryotic models.

Some of the most common toxins present in freshwater, in particular microcystins (MCs), are produced by cyanobacteria. These toxins have a negative impact on human health, being associated with episodes of acute hepatotoxicity and being considered potentially carcinogenic to humans. To date the exact mechanisms of MC-induced toxicity and tumor promotion were not completely elucidated. To get new insights underlying microcystin-LR (MCLR) molecular mechanisms of toxicity we have performed the proteomic profiling using two-dimensional electrophoresis and MALDI-TOF/TOF of Saccharomyces cerevisiae cells exposed for 4 h-1 nM and 1 µM of MCLR, and compared them to the control (cells not exposed to MCLR). We identified 14 differentially expressed proteins. The identified proteins are involved in metabolism, genotoxicity, cytotoxicity and stress response. Furthermore, we evaluated the relative expression of yeast's PP1 and PP2A genes and also of genes from the Base Excision Repair (BER) DNA-repair system, and observed that three out of the five genes analyzed displayed dose-dependent responses. Overall, the different proteins and genes affected are related to oxidative stress and apoptosis, thus reinforcing that it is probably the main mechanism of MCLR toxicity transversal to several organisms, especially at lower doses. Notwithstanding these MCLR responsive proteins could be object of further studies to evaluate their suitability as biomarkers of exposure to the toxin.

Proteomic analysis of domestic pig pancreas during development using two-dimensional electrophoresis and matrix-assisted laser desorption/ionization-time of flight mass spectrometry.

Pig pancreas may be a therapeutic resource for human diabetic patients. However, this potential is hindered by a lack of knowledge of the molecular events of pig pancreas development. In this study, the embryonic day 60, neonate and 6-month protein profiles of pig pancreas were ascertained at using two-dimensional gel electrophoresis and matrix assisted laser desorption/ionization-time of flight mass spectrometry. Twenty four proteins were differentially expressed during pig pancreas development. Among them, 12 spots increased and 7 spots decreased according to development. The expression of 5 protein were highest at birth. Expression of digestive enzymes including trypsin, pancreatic triacylglycerol lipase and pancreatic alpha-amylase was elevated in adults, whereas chymotrypsins were highly expressed in neonates. Proteins that were abundantly expressed during gestation were alpha-1-antitrypsin, alpha-fetoprotein and transferrins. Taken together, we found out that several proteins were significantly up- or down- regulated from pig pancreas based on developmental stage. This study will provide basis for understanding development of pig pancreas.