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Background: Chlamydomonas reinhardtii is gaining recognition as a promising expression system for the production of recombinant proteins. However, its performance as a cellular biofactory remains suboptimal, especially with respect to consistent expression of heterologous genes. Gene silencing mechanisms, position effect, and low nuclear transgene expression are major drawbacks for recombinant protein production in this model system. To unveil the molecular changes following transgene insertion, retention, and expression in this species, we genetically engineered C. reinhardtii wild type strain 137c (strain cc-125 mt+) to express the fluorescent protein mVenus and subsequently analysed its intracellular proteome. Results: The obtained transgenic cell lines showed differences in abundance in more than 400 proteins, with multiple pathways altered post-transformation. Proteins involved in chromatin remodelling, translation initiation and elongation, and protein quality control and transport were found in lower abundance. On the other hand, ribosomal proteins showed higher abundance, a signal of ribosomal stress response. Conclusions: These results provide new insights into the modifications of C. reinhardtii proteome after transformation, highlighting possible pathways involved in gene silencing. Moreover, this study identifies multiple protein targets for future genetic engineering approaches to improve the prospective use of C. reinhardtii as cell biofactory for industrial applications. [ABSTRACT FROM AUTHOR]

Background: Chlamydomonas reinhardtii is gaining recognition as a promising expression system for the production of recombinant proteins. However, its performance as a cellular biofactory remains suboptimal, especially with respect to consistent expression of heterologous genes. Gene silencing mechanisms, position effect, and low nuclear transgene expression are major drawbacks for recombinant protein production in this model system. To unveil the molecular changes following transgene insertion, retention, and expression in this species, we genetically engineered C. reinhardtii wild type strain 137c (strain cc-125 mt+) to express the fluorescent protein mVenus and subsequently analysed its intracellular proteome. Results: The obtained transgenic cell lines showed differences in abundance in more than 400 proteins, with multiple pathways altered post-transformation. Proteins involved in chromatin remodelling, translation initiation and elongation, and protein quality control and transport were found in lower abundance. On the other hand, ribosomal proteins showed higher abundance, a signal of ribosomal stress response. Conclusions: These results provide new insights into the modifications of C. reinhardtii proteome after transformation, highlighting possible pathways involved in gene silencing. Moreover, this study identifies multiple protein targets for future genetic engineering approaches to improve the prospective use of C. reinhardtii as cell biofactory for industrial applications. [ABSTRACT FROM AUTHOR]

Phosphorus (P) is an essential macronutrient required for respiration, photosynthesis/carbohydrate metabolism, redox homeostasis, signaling, and synthesis/function of nucleic acids, cellular membranes, enzymes, etc. To cope with P-deficiency, plants reprogram gene expression for necessary alterations in metabolic/signaling pathways. To attain P homeostasis, plants readjust metabolism through transcriptional, post-transcriptional, and/or post-translational machinery involving biochemical, physiological, genomic, epigenomic, proteomic, and metabolomic functions. However, the underlying molecular mechanisms/pathways/genes for P-deficiency tolerance in crop plants remain elusive. To decipher the mechanisms/pathways adopted by rice under P-starvation/deficiency stress, a pair of contrasting rice [Pusa-44 (high-yielding, P-deficiency sensitive) and its near-isogenic line (NIL)-23, P-deficiency tolerant) for Pup1 QTL] genotype was used for comparative analyses. Omics analyses of shoot and root tissues from 45-day-old plants grown hydroponically in P-sufficient (16 ppm Pi), P-deficient (4 ppm Pi) or P-starved (0 ppm Pi) medium revealed important roles of P transporters, transcription factors (TFs), Transposable elements (TEs), auxin-responsive proteins, cell wall modulation, fatty acid metabolism, and chromatin architecture/epigenetic modifications in making NIL-23 tolerant to stress. The proteins involved in photosynthesis, sucrose-/starch-/energy-metabolism, transcription factors, and phytohormone signaling were observed to be differentially expressed in NIL-23. Since only a few coding genes are located on the QTL, modulations in morpho-physio-biochemical and molecular parameters under P-starvation/deficiency stress were attributed to the regulatory functions of Pup1 through TFs, TEs, epigenetic/chromatin architectural changes, etc. introgressed in Pusa-44 genetic background. Thus, the study provides new insights into molecular/regulatory functions of Pup1 under P-starvation/deficiency stress in rice, which might be useful to improve P-use efficiency in rice for better productivity in P-deficient soils. [ABSTRACT FROM AUTHOR]

The iPSC-derived 3D models are considered to be a connective link between 2D culture and in vivo studies. However, the sensitivity of such 3D models is yet to be established. We assessed the sensitivity of the hiPSC-derived 3D spheroids against 2D cultures of neural progenitor cells. The sub-toxic dose of Sodium Arsenite (SA) was used to investigate the alterations in miRNA-proteins in both systems. Though SA exposure induced significant alterations in the proteins in both 2D and 3D systems, these proteins were uncommon except for 20 proteins. The number and magnitude of altered proteins were higher in the 2D system compared to 3D. The association of dysregulated miRNAs with the target proteins showed their involvement primarily in mitochondrial bioenergetics, oxidative and ER stress, transcription and translation mechanism, cytostructure, etc., in both culture systems. Further, the impact of dysregulated miRNAs and associated proteins on these functions and ultrastructural changes was compared in both culture systems. The ultrastructural studies revealed a similar pattern of mitochondrial damage, while the cellular bioenergetics studies confirm a significantly higher energy failure in the 2D system than to 3D. Such a higher magnitude of changes could be correlated with a higher amount of internalization of SA in 2D cultures than in 3D spheroids. Our findings demonstrate that a 2D culture system seems better responsive than a 3D spheroid system against SA exposure. [ABSTRACT FROM AUTHOR]

Background: Boron (B) is an essential micronutrient for plants. Inappropriate B supply detrimentally affects the productivity of numerous crops. Understanding of the molecular responses of plants to different B supply levels would be of significance in crop improvement and cultivation practices to deal with the problem. Results: We conducted a comprehensive analysis of the transcriptome and proteome of tobacco seedlings to investigate the expression changes of genes/proteins in response to different B supply levels, with a particular focus on B deficiency. The global gene and protein expression profiles revealed the potential mechanisms involved in the responses of tobacco to B deficiency, including up-regulation of the NIP5;1-BORs module, complex regulation of genes/proteins related to cell wall metabolism, and up-regulation of the antioxidant machinery. Conclusion: Our results demonstrated that B deficiency caused severe morphological and physiological disorders in tobacco seedlings, and revealed dynamic expression changes of tobacco genes/proteins in response to different B supply levels, especially to B deficiency, thus offering valuable insights into the molecular responses of tobacco to B deficiency. [ABSTRACT FROM AUTHOR]

In the context of proteomic-scale research, it is imperative to automatically analyze numerous species and subspecies to discern distinctive characteristics present in multiple species of the fungi kingdom that contain sequences of interest that could fulfill a specific biological function. To achieve this, complex sequences must be recognized within an organism's entire set of proteomes. Our study presents FungiRegEx, a piece of software that facilitates the identification of regular expressions of proteomes of fungal organisms and uses real-time data retrieval of the different species from the JGI Mycocosm database without the need to download any file. Integrating a graphical user interface that makes it easy to use, the tool offers regular expression searches on 2402 fungal species from the JGI Mycocosm portal. The tool was validated with the AXSXG sequence and the RXRL effector, demonstrating the effectiveness of FungiRegEx in identifying user-defined patterns in the recovered sequences. This tool allows customization and filtering, and it can save results if required, combining speed, adaptability, and ease of use. It provides an experience without a console and programming, displaying the results in a GUI and making them easier to read. Its architecture guarantees optimized use of resources, time consumption, and implementation flexibility, allowing the customization of specific software parameters for resource management. The tool's potential for future research and exploration is emphasized, providing a nuanced perspective on its practical use within the fungal genomics community. The tools are available at the addresses mentioned in the text. [ABSTRACT FROM AUTHOR]

Neutrophil and (alveolar) macrophage immunity is considered crucial for eliminating Aspergillus fumigatus. Data derived from bronchoalveloar lavage (BAL) characterizing the human immuno-pulmonary response to Aspergillus fumigatus are non-existent. To obtain a comprehensive picture of the immune pathways involved in chronic pulmonary aspergillosis (CPA), we performed proteome analysis on AL of 9 CPA patients and 17 patients with interstitial lung disease (ILD). The dihydrorhodamine (DHR) test was also performed on BAL and blood neutrophils from CPA patients and compared to blood neutrophils from healthy controls (HCs). BAL from CPA patients primarily contained neutrophils, while ILD BAL was also characterized by a large fraction of lymphocytes; these differences likely reflecting the different immunological etiologies underlying the two disorders. BAL and blood neutrophils from CPA patients displayed the same oxidative burst capacity as HC blood neutrophils. Hence, immune evasion by Aspergillus involves other mechanisms than impaired neutrophil oxidative burst capacity per se. CPA BAL was enriched by proteins associated with innate immunity, as well as, more specifically, with neutrophil degranulation, Toll-like receptor 4 signaling, and neutrophil-mediated iron chelation. Our data provide the first comprehensive target organ-derived immune data on the human pulmonary immune response to Aspergillus fumigatus. [ABSTRACT FROM AUTHOR]

Breast cancer (BC) remains one of the leading causes of mortality among women, with triple-negative breast cancer (TNBC) standing out for its aggressive nature and limited treatment options. Metabolic reprogramming, one of cancer's hallmarks, underscores the importance of targeting metabolic vulnerabilities for therapeutic intervention. This study aimed to investigate the impact of de novo serine biosynthetic pathway (SSP) inhibition, specifically targeting phosphoglycerate dehydrogenase (PHGDH) with NCT-503, on three TNBC cell lines: MDA-MB-231, MDA-MB-468 and Hs 578T. First, MS-based proteomics was used to confirm the distinct expression of PHGDH and other SSP enzymes using the intracellular proteome profiles of untreated cells. Furthermore, to characterize the response of the TNBC cell lines to the inhibitor, both in vitro assays and label-free, bottom-up proteomics were employed. NCT-503 exhibited significant cytotoxic effects on all three cell lines, with MDA-MB-468 being the most susceptible (IC50 20.2 ± 2.8 µM), while MDA-MB-231 and Hs 578T showed higher, comparable IC50s. Notably, differentially expressed proteins (DEPs) induced by NCT-503 treatment were mostly cell line-specific, both in terms of the intracellular and secreted proteins. Through overrepresentation and Reactome GSEA analysis, modifications of the intracellular proteins associated with cell cycle pathways were observed in the MDA-MBs following treatment. Distinctive dysregulation of signaling pathways were seen in all TNBC cell lines, while modifications of proteins associated with the extracellular matrix organization characterizing both MDA-MB-231 and Hs 578T cell lines were highlighted through the treatment-induced modifications of the secreted proteins. Lastly, an analysis was conducted on the DEPs that exhibited greater abundance in the NCT-503 treatment groups to evaluate the potential chemo-sensitizing properties of NCT-503 and the druggability of these promising targets. [ABSTRACT FROM AUTHOR]

Macrophages are key immune cells that can adapt their metabolic phenotype in response to different stimuli. Lysine deacetylases are important enzymes regulating inflammatory gene expression and lysine deacetylase inhibitors have been shown to exert anti-inflammatory effects in models of chronic obstructive pulmonary disease. We hypothesized that these anti-inflammatory effects may be associated with metabolic changes in macrophages. To validate this hypothesis, we used an unbiased and a targeted proteomic approach to investigate metabolic enzymes, as well as liquid chromatography–mass spectrometry and gas chromatography–mass spectrometry, to quantify metabolites in combination with the measurement of functional parameters in primary murine alveolar-like macrophages after lipopolysaccharide-induced activation in the presence or absence of lysine deacetylase inhibition. We found that lysine deacetylase inhibition resulted in reduced production of inflammatory mediators such as tumor necrosis factor α and interleukin 1β. However, only minor changes in macrophage metabolism were observed, as only one of the lysine deacetylase inhibitors slightly increased mitochondrial respiration while no changes in metabolite levels were seen. However, lysine deacetylase inhibition specifically enhanced expression of proteins involved in ubiquitination, which may be a driver of the anti-inflammatory effects of lysine deacetylase inhibitors. Our data illustrate that a multiomics approach provides novel insights into how macrophages interact with cues from their environment. More detailed studies investigating ubiquitination as a potential driver of lysine deacetylase inhibition will help developing novel anti-inflammatory drugs for difficult-to-treat diseases such as chronic obstructive pulmonary disease. [ABSTRACT FROM AUTHOR]

Duckweed is an aquatic model plant with tremendous potential in industrial and agricultural applications. Duckweed rarely flowers which significantly hinders the resource collection and heterosis utilization. Salicylic acid (SA) can significantly induce duckweed to flower; however, the underlying regulatory mechanisms remain largely unknown. In this work, transcriptome and proteome were conducted in parallel to examine the expression change of genes and proteins in Lemna gibba under SA treatment. A high-quality reference transcriptome was generated using Iso-Seq strategy, yielding 42,281 full-length transcripts. A total of 422, 423, and 417 differentially expressed genes (DEGs), as well as 213, 51, and 92 differentially expressed proteins (DEPs), were identified at flower induction, flower initiation, and flowering stages by ssRNA-seq and iTRAQ methods. Most DEGs and DEPs were only regulated at either the transcriptomic or proteomic level. Additionally, DEPs exhibited low expression correlations with the corresponding mRNAs, suggesting that post-transcriptional regulation plays a pivotal role in SA-induced flowering in L. gibba. Specifically, the genes related to photosynthesis, stress, and hormone metabolism were mainly regulated at the mRNA level, those associated with mitochondrial electron transport / ATP synthesis, nucleotide synthesis, and secondary metabolism were regulated at the protein level, while those related to redox metabolism were regulated at the mRNA and/or protein levels. The post-transcriptional regulation of genes relevant to hormone synthesis, transcription factors, and flowering was also extensively analyzed and discussed. This is the first study of integrative transcriptomic and proteomic analyses in duckweed, providing novel insights of post-transcriptional regulation in SA-induced flowering of L. gibba. [ABSTRACT FROM AUTHOR]

Background: Amyotrophic lateral sclerosis/Parkinsonism-dementia complex in Kii peninsula, Japan (Kii ALS/PDC), is an endemic neurodegenerative disease whose causes and pathogenesis remain unknown. However, astrocytes in autopsied cases of Kii ALS/PDC show characteristic lesions. In addition, relationships between extracellular vesicles (EVs) and neurodegenerative diseases are increasingly apparent. Therefore, we focused on proteins in EVs derived from Kii ALS/PDC astrocytes in the present study. Methods: Induced pluripotent stem cells (iPSCs) derived from three healthy controls (HCs) and three patients with Kii ALS/PDC were differentiated into astrocytes. EVs in the culture medium of astrocytes were collected and subjected to quantitative proteome analysis. Results: Our proteome analysis reveals that EV-containing proteins derived from astrocytes of patients with Kii ALS/PDC show distinctive patterns compared with those of HCs. Moreover, EVs derived from Kii ALS/PDC astrocytes display increased proteins related to proteostasis and decreased proteins related to anti-inflammation. Discussion: Proteins contained in EVs from astrocytes unveil protective support to neurons and may reflect the molecular pathomechanism of Kii ALS/PDC; accordingly, they may be potential biomarker candidates of Kii ALS/PDC. [ABSTRACT FROM AUTHOR]

Background: Blood–brain barrier (BBB) dysfunction is supposed to be an early event in the development of Alzheimer's disease (AD). This study aimed to investigate the relationship between BBB alterations and AD progression in terms of amyloid-β peptide (Aβ) accumulation in the brains of humanized amyloid precursor protein knock-in (APP-KI) mice. Methods: Brain Aβ accumulation was examined using immunohistochemical analysis. Alterations in differentially expressed proteins were determined using sequential window acquisition of all theoretical fragment ion mass spectroscopy (SWATH-MS)-based quantitative proteomics, and Metascape, STRING, Gene Ontology, and KEGG were used for network analyses of altered biological pathways and processes. Statistical significance was determined using the unpaired two-tailed Student's t-test and Welch's t-test for two groups and one-way analysis of variance followed by Tukey's test for more than two groups. Correlations between two groups were determined using Pearson's correlation analysis. Results: Brain Aβ accumulation in APP-KI mice was detectable at 2 months, increased significantly at 5 months, and remained elevated at 12 months of age. The levels of differentially expressed proteins in isolated brain capillaries were higher in younger mice, whereas those in the brain were higher in older mice. Network analyses indicated changes in basement membrane-associated and ribosomal proteins in the brain capillaries. There were no significant changes in key proteins involved in drug or Aβ transport at the BBB. In contrast, solute carrier transporter levels in astrocytes, microglia, and neurons were altered in the brain of older mice. Moreover, the levels of the lipid transporters Apoe and Apoj were upregulated in both the brain and isolated brain capillaries after Aβ accumulation. Conclusions: Our results suggest that changes in the brain occurred after advanced Aβ accumulation, whereas initial Aβ accumulation was sufficient to cause alterations in the BBB. These findings may help elucidate the role of BBB alterations in AD progression and predict the distribution of drugs across the BBB in the brain of patients with AD. [ABSTRACT FROM AUTHOR]

Idesia polycarpa is a dioecious tree; in field surveys, there are rare sex conversions in I. polycarpa individuals with bisexual flowers. To identify the molecular mechanisms underlying sex conversion in this species, an integrative analysis of the proteome and transcriptome profiles of I. polycarpa male and bisexual flowers at key developmental stages was conducted in this study using isobaric tags for relative and absolute quantification and RNA-seq technology. A total of 15,003 proteins were identified; the differentially expressed proteins (DEPs) were enriched in metabolic pathways, biosynthesis of secondary metabolites, and flavonoid metabolism pathways in all comparison groups. A total of 290,442 unigenes were obtained; these were compared with seven databases, revealing 196,366 annotated unigenes. In general, the expression of proteins and genes tended to be positively correlated, with Spearman correlation coefficients in the ranges of 0.152–0.262 (all genes and all proteins) and 0.497–0.778 (DEPs and DEGs). The integrative analysis of DEPs and DEGs between male and bisexual flowers revealed that the most significantly enriched pathways were flavonoid pathways, metabolic pathways, and the biosynthesis of secondary metabolites. Finally, four co-expressed proteins and transcripts and one gene associated with the flavonoid biosynthesis pathway were screened out. The proteins identified were p-coumaroyl shikimate 3′-hydroxylase, and shikimate/quinate hydroxycinnamoyl transferase, and the gene was caffeoyl-CoA O-methyltransferase. The analysis has revealed key potential proteins and genes involved in sex conversion at the molecular level and has provided a basis for future investigations of artificial regulation of sex differentiation in I. polycarpa. [ABSTRACT FROM AUTHOR]

Obesity is mostly associated with adverse health consequences, but may also elicit favorable effects under chronic conditions. This “obesity paradox” is under debate for pulmonary diseases. As confounding factors complicate conclusions from human studies, this study used a controlled animal model combining diet-induced obesity and chronic hypoxia as a model for pulmonary hypertension and chronic obstructive pulmonary disease. Male C57BL/6 mice were fed control or high-fat diet for 30 wk, and half of the animals were exposed to chronic hypoxia (13% O2) for 3 wk. Hypoxia induced right ventricular hypertrophy, thickening of pulmonary arterial and capillary walls, higher lung volumes, and increased hemoglobin concentrations irrespective of the body weight. In contrast, lung proteomes differed substantially between lean- and obese-hypoxic mice. Many of the observed changes were linked to vascular and extracellular matrix (ECM) proteins. In lean-hypoxic animals, circulating platelets were reduced and abundances of various clotting-related proteins were altered, indicating a hypercoagulable phenotype. Moreover, the septal ECM composition was changed, and airspaces were significantly distended pointing to lung hyperinflation. These differences were mostly absent in the obese-hypoxic group. However, the obesity-hypoxia combination induced the lowest blood CO2 concentrations, indicating hyperventilation for sufficient oxygen supply. Moreover, endothelial surface areas were increased in obese-hypoxic mice. Thus, obesity exerts differential effects on lung adaptation to hypoxia, which paradoxically include not only adverse but also rather protective changes. These differences have a molecular basis in the lung proteome and may influence the pathogenesis of lung diseases. This should be taken into account for future individualized prevention and therapy. NEW & NOTEWORTHY An “obesity paradox” is discussed for pulmonary diseases. By linking lung proteome analyses to pulmonary structure and function, we demonstrate that diet-induced obesity affects lung adaptation to chronic hypoxia in various ways. The observed changes include not only adverse but also protective effects and are associated with altered abundances of vascular and extracellular matrix proteins. These results highlight the existence of relevant differences in individuals with obesity that may influence the pathogenesis of lung diseases. [ABSTRACT FROM AUTHOR]

Background: Effective diagnostic biomarkers for aortic aneurysm (AA) that are detectable in blood tests are required because early detection and rupture risk assessment of AA can provide insights into medical therapy and preventive treatments. However, known biomarkers for AA lack specificity and reliability for clinical diagnosis. Methods: We performed proteome analysis of serum samples from patients with atherosclerotic thoracic AA (TAA) and healthy control (HC) subjects to identify diagnostic biomarkers for AA. Serum samples were separated into low-density lipoprotein, high-density lipoprotein, and protein fractions, and the major proteins were depleted. From the proteins identified in the three fractions, we narrowed down biomarker candidates to proteins uniformly altered in all fractions between patients with TAA and HC subjects and evaluated their capability to discriminate patients with TAA and those with abdominal AA (AAA) from HC subjects using receiver operating characteristic (ROC) analysis. For the clinical validation, serum concentrations of biomarker candidates were measured in patients with TAA and AAA registered in the biobank of the same institute, and their capability for the diagnosis was evaluated. Results: Profilin 1 (PFN1) and complement factor D (CFD) showed the most contrasting profiles in all three fractions between patients with TAA and HC subjects and were selected as biomarker candidates. The PFN1 concentration decreased, whereas the CFD concentration increased in the sera of patients with TAA and AAA when compared with those of HC subjects. The ROC analysis showed that these proteins could discriminate patients with TAA and AAA from HC subjects. In the validation study, these candidates showed significant concentration differences between patients with TAA or AAA and controls. PFN1 and CFD showed sufficient area under the curve (AUC) in the ROC analysis, and their combination further increased the AUC. The serum concentrations of PFN1 and CFD also showed significant differences between patients with aortic dissection and controls in the validation study. Conclusion: PFN1 and CFD are potential diagnostic biomarkers for TAA and AAA and measurable in blood samples; their diagnostic performance can be augmented by their combination. These biomarkers may facilitate the development of diagnostic systems to identify patients with AA. [ABSTRACT FROM AUTHOR]

The nuclear envelope (NE) is a double-membrane structure consisting of inner and outer membranes that spatially separate the nucleus from the cytoplasm, and its function is critical for cellular functions such as genome maintenance. In the fission yeast, Schizosaccharomyces pombe , the inner nuclear membrane proteins, Lem2 and Bqt4, play pivotal roles in maintaining the NE structure. We previously found that the double deletion of lem2 + and bqt4 + causes a synthetic lethal defect associated with severe NE rupture, and overexpression of Elo2, a solo very-long-chain fatty acid elongase, suppresses this defect by restoring the NE. However, the molecular basis of this restoration remains elusive. To address this, we identified Lem2- and Bqt4-binding proteins via immunoprecipitation and mass spectrometry in this study. Forty-five and 23 proteins were identified as Lem2- and Bqt4-binding proteins, respectively. Although these binding proteins partially overlapped, Lem2 and Bqt4 interacted with different types of lipid metabolic enzymes: Cho2, Ole1 and Erg11 for Lem2 and Cwh43 for Bqt4. These enzymes are known to be involved in various lipid synthesis processes, suggesting that Lem2 and Bqt4 may contribute to the regulation of lipid synthesis by binding to these enzymes. [ABSTRACT FROM AUTHOR]

In this experiment, the effect of salt stress on the root proteome pattern of Arg (tolerant to salinity) and Moghan3 (sensitive to salinity) cultivars of wheat was investigated by two-dimensional polyacrylamide gel electrophoresis. Salinity was applied at two levels of 0 and 150 mM in the third-leaf stage for three weeks under greenhouse conditions. The proteome analysis of the roots revealed 120 reproducible protein spots, among which 15 spots showed significant changes in expression. In Moghan3, four protein spots with a significant reduction in expression were identified. In the tolerant cultivar of Arg, 11 protein spots showed significant changes in expression, among which five protein spots had an increased expression and six protein spots had a decreased expression. These proteins were classified based on their function into several groups such as the proteins involved in metabolism and energy, reactive oxygen species scavenging and detoxification, the proteins associated with cell walls, and the proteins involved in folding and degradation. Probably, the tolerance to salt stress in the Arg cultivar was related to the increased expression of pyruvate dehydrogenase, phosphoglycerate mutase, catalase, and malate dehydrogenase proteins. While in the sensitive variety of Moghan3, the decrease in the expression of enolase, peroxidase, and glutathione peroxidase proteins under salinity stress conditions probably caused oxidative stress in the plants due to increased production of reactive oxygen species. These findings can be useful for improving the salt tolerance in the breeding programs of wheat. [ABSTRACT FROM AUTHOR]

Background: Thickening of the ligamentum flavum is considered to be the main factor for lumbar spinal canal stenosis (LSCS). Although some mechanisms have been speculated in the thickening of the ligamentum flavum, there are only a few comprehensive approaches to investigate its pathology. The objective of this study was to investigate the pathology of thickened ligamentum flavum in patients with LSCS based on protein expression levels using shotgun proteome analysis. Methods: Ligamentum flavum samples were collected from four patients with LSCS (LSCS group) and four patients with lumbar disc herniation (LDH) as controls (LDH group). Protein mixtures were digested and analyzed by liquid chromatography/mass spectrometry analysis. To compare protein expression levels between the LSCS and LDH groups, the mean Mascot score was compared. Biological processes were assessed using Gene Ontology analysis. Results: A total of 1151 proteins were identified in some samples of ligamentum flavum. Among these, 145 proteins were detected only in the LSCS group, 315 in the LDH group, and 691 in both groups. The demonstrated biological processes occurring in the LSCS group included: extracellular matrix organization, regulation of peptidase activity, extracellular matrix disassembly, and negative regulation of cell growth. Proteins related to fibrosis, chondrometaplasia, and amyloid deposition were found highly expressed in the LSCS group compared with those in the LDH group. Conclusions: Tissue repair via fibrosis, chondrometaplasia, and amyloid deposits may be important pathologies that occur in the thickened ligamentum flavum of patients with LSCS. [ABSTRACT FROM AUTHOR]

Many studies have been conducted to elucidate the role of Type VI collagen in muscle and tendon, however, its role in oral tissues remains unclear. In this study, an α2(VI) deficient mouse (Col6α2-KO) model was used to examine the role of Type VI collagen in oral tissues. Tissue volume and mineral density were measured in oral tissues by µCT. Proteome analysis was performed using protein extracted from alveolar bone. In addition, alveolar bone was evaluated with a periodontitis induced model. µCT analysis showed the Col6α2-KO mice had less volume of alveolar bone, dentin and dental pulp, while the width of periodontal ligament (PDL) was greater than WT. The mineral density in alveolar bone and dentin were elevated in Col6α2-KO mice compared with WT. Our proteome analysis showed significant changes in proteins related to ECM organization and elevation of proteins associated with biomineralization in the Col6α2-KO mice. In induced periodontitis, Col6α2-KO mice had greater alveolar bone loss compared with WT. In conclusion, Type VI collagen has a role in controlling biomineralization in alveolar bone and that changes in the ECM of alveolar bone could be associated with greater bone loss due to periodontitis. [ABSTRACT FROM AUTHOR]

While wheat (Triticum aestivum L.) is a widely grown and enjoyed crop, the diverse and complex global situation and climate are exacerbating the instability of its supply. In particular, pre-harvest sprouting (PHS) is one of the major abiotic stresses that frequently occurs due to irregular climate conditions, causing serious damage to wheat and its quality. In this study, transcriptomic analysis with RNA-seq and proteomic analysis with LC-MS/MS were performed in PHS-treated spikes from two wheat cultivars presenting PHS sensitivity and tolerance, respectively. A total of 13,154 differentially expressed genes (DEGs) and 706 differentially expressed proteins (DEPs) were identified in four comparison groups between the susceptible/tolerant cultivars. Gene function and correlation analysis were performed to determine the co-profiled genes and proteins affected by PHS treatment. In the functional annotation of each comparative group, similar functions were confirmed in each cultivar under PHS treatment; however, in Keumgang PHS+7 (K7) vs. Woori PHS+7 (W7), functional annotations presented clear differences in the "spliceosome" and "proteasome" pathways. In addition, our results indicate that alternative splicing and ubiquitin–proteasome support the regulation of germination and seed dormancy. This study provides an advanced understanding of the functions involved in transcription and translation related to PHS mechanisms, thus enabling specific proposals for the further analysis of germination and seed dormancy mechanisms and pathways in wheat. [ABSTRACT FROM AUTHOR]

Enzalutamide (ENZA) is a frequently used therapy in metastatic castration‐resistant prostate cancer (mCRPC). Baseline or acquired resistance to ENZA have been observed, but the molecular mechanisms of resistance are poorly understood. We aimed to identify proteins involved in ENZA resistance and to find therapy‐predictive serum markers. We performed comparative proteome analyses on ENZA‐sensitive parental (LAPC4, DuCaP) and ‐resistant prostate cancer cell lines (LAPC4‐ENZA, DuCaP‐ENZA) using liquid chromatography tandem mass spectrometry (LC‐MS/MS). The top four most promising candidate markers were selected using bioinformatic approaches. Serum concentrations of selected markers (ALCAM, AGR2, NDRG1, IDH1) were measured in pretreatment samples of 72 ENZA‐treated mCRPC patients using ELISA. In addition, ALCAM serum levels were measured in 101 Abiraterone (ABI) and 100 Docetaxel (DOC)‐treated mCRPC patients' baseline samples. Results were correlated with clinical and follow‐up data. The functional role of ALCAM in ENZA resistance was assessed in vitro using siRNA. Our proteome analyses revealed 731 significantly differentially abundant proteins between ENZA‐sensitive and ‐resistant cells and our filtering methods identified four biomarker candidates. Serum analyses of these proteins revealed only ALCAM to be associated with poor patient survival. Furthermore, higher baseline ALCAM levels were associated with poor survival in ABI‐ but not in DOC‐treated patients. In LAPC4‐ENZA resistant cells, ALCAM silencing by siRNA knockdown resulted in significantly enhanced ENZA sensitivity. Our analyses revealed that ALCAM serum levels may help to identify ENZA‐ and ABI‐resistant patients and may thereby help to optimize future clinical decision‐making. Our functional analyses suggest the possible involvement of ALCAM in ENZA resistance. [ABSTRACT FROM AUTHOR]

Previous studies have shown that early therapeutic events of neural precursor cells (NPCs) transplantation to animals with acute ischemic stroke readily protected neuronal cell damage and improved behavioral recovery through paracrine mechanisms. In this study, we tested the hypothesis that administration of conditioned medium from NPCs (NPC-CMs) could recapitulate the beneficial effects of cell transplantation. Rats with permanent middle cerebral artery occlusion (pMCAO) were randomly assigned to one of the following groups: PBS control, Vehicle (medium) controls, single (NPC-CM(S)) or multiple injections of NPC-CM(NPC-CM(M)) groups. A single intravenous injection of NPC-CM exhibited strong neuroregenerative potential to induce behavioral recovery, and multiple injections enhanced this activity further by suppressing inflammatory damage and inducing endogenous neurogenesis leading to histopathological and functional recovery. Proteome analysis of NPC-CM identified a number of proteins that are known to be associated with nervous system development, neurogenesis, and angiogenesis. In addition, transcriptome analysis revealed the importance of the inflammatory response during stroke recovery and some of the key hub genes in the interaction network were validated. Thus, our findings demonstrated that NPC-CM promoted functional recovery and reduced cerebral infarct and inflammation with enhanced endogenous neurogenesis, and the results highlighted the potency of NPC-CM in stroke therapy. [ABSTRACT FROM AUTHOR]

High expression of carbonyl reductase 1 (CBR1) protein in ovarian cancer cells inhibits tumor growth and metastasis. However, the underlying mechanism is unknown. To investigate the mechanism by which CBR1 suppresses tumor growth, the present study generated ovarian cancer cells that constitutively overexpress human CBR1 (hCBR1) protein. Ovarian cancer cell lines (OVCAR-3 and SK-OV-3) were transfected with a plasmid encoding hCBR1, followed by selection with G418 to isolate hCBR1-overexpressing lines. The proliferation rates of hCBR1-overexpressing cells were then compared with those of negative control and wild-type cells. Overexpression of hCBR1 led to significant inhibition of proliferation (P<0.05). Subsequently, to investigate changes in intracellular signaling pathways, cellular proteins were extracted and subjected to proteome analysis using liquid chromatography followed by mass spectrometry. There was an inverse correlation between CBR1 protein expression and cell proliferation. In addition, Ingenuity Pathway Analysis of hCBR1-overexpressing cell lines was performed, which revealed changes in the expression of proteins involved in signaling pathways related to growth regulation. Of these, the eukaryotic translation initiation factor 2 (eIF2) signaling pathway was upregulated most prominently. Thus, alterations in multiple tumor-related signaling pathways, including eIF2 signaling, may lead to growth suppression. Taken together, the present data may lead to the development of new drugs that target CBR1 and related signaling pathways, thereby improving outcomes for patients with ovarian cancer. [ABSTRACT FROM AUTHOR]

Salivary gland‐specific transcriptomes of nine heteropteran insects with distinct feeding strategies (predaceous, hematophagous, and phytophagous) were analyzed and annotated to compare and identify the venom components as well as their expression profiles. The transcriptional abundance of venom genes was verified via quantitative real‐time PCR. Hierarchical clustering of 30 representative differentially expressed venom genes from the nine heteropteran species revealed unique groups of salivary gland‐specific genes depending on their feeding strategy. The commonly transcribed genes included a paralytic neurotoxin (arginine kinase), digestive enzymes (cathepsin and serine protease), an anti‐inflammatory protein (cystatin), hexamerin, and an odorant binding protein. Both predaceous and hematophagous (bed bug) heteropteran species showed relatively higher transcription levels of genes encoding proteins involved in proteolysis and cytolysis, whereas phytophagous heteropterans exhibited little or no expression of these genes, but had a high expression of vitellogenin, a multifunctional allergen. Saliva proteomes from four representative species were also analyzed. All venom proteins identified via saliva proteome analysis were annotated using salivary gland transcriptome data. The proteomic expression profiles of venom proteins were in good agreement with the salivary gland‐specific transcriptomic profiles. Our results indicate that profiling of the salivary gland transcriptome provides important information on the composition and evolutionary features of venoms depending on their feeding strategy. [ABSTRACT FROM AUTHOR]

Amyloidosis is induced by extracellular deposition of certain proteins. Thirty-six proteins have so far been identified as amyloidogenic proteins in humans. Although it is very important to determine the specific amyloid protein type for the choice of therapy for amyloidosis patient, it might be difficult to identify specific proteins from amyloid-deposited tissue. Apolipoprotein A-IV is known as an amyloid-associated protein, but there have been few reports of apolipoprotein A-IV amyloidosis. Here we report a case of systemic apolipoprotein A-IV-associated amyloidosis that was confirmed by proteome analysis using formalin-fixed paraffin-embedded tissue and an immunohistochemical technique. [ABSTRACT FROM AUTHOR]

Attenuating the Taxol biosynthesis by fungi with storage and subculturing is the major challenge that limits their further industrial applications. Aspergillus flavipes has been reported as a potent Taxol producer, with plausible increasing to its Taxol yield upon coculturing with the microbiome of Podocarpus gracilior (El-Sayed et al., Process Biochemistry 76:55–67, 2019a; Scientific Reports 9, 2019b; Enzyme and Microbial Technology 131, 2019c); however, the identity of these microbial inducers remains ambiguous. Thus, this study was to assess the potency of individual microbes to trigger the Taxol biosynthesis by A. flavipes and to unravel the differentially expressed protein in response to bacterial interaction. Among the 25 bacterial endophytes of P. gracilior, Bacillus subtilis was the potent isolate enhancing the Taxol yield of A. flavipes by ~1.6-fold. Strikingly, this bacterial elicitor displayed a reliable inhibition to the growth of A. flavipes, so the released antifungal compound by B. subtilis could be the same signals for triggering the expression of A. flavipes Taxol synthesis. The highest Taxol yield by A. flavipes was obtained with the viable cells of B. subtilis, ensuring the pivotality of physical intimate bacterial-fungal interaction. Differential proteome of the cocultures A. flavipes and B. subtilis as well as the axenic A. flavipes was conducted by LC-MS/MS. From the total of 106 identified proteins, 50 proteins were significantly expressed, 47 were upregulated ones, and 59 were downregulated ones for the cocultures normalizing to the axenic one. From the Gene Ontology (GO) and KEGG enrichment analyses, the cellular process, primary metabolic process, and nitrogen compound metabolic process were significantly changed in the coculture normalizing to monoculture of A. flavipes. The molecular function terms (histones H2B, H2A, peptidyl-prolyl cis-trans isomerase, and nucleoside-diphosphate kinase (NDPK)) were the highly significantly expressed proteins of A. flavipes in response to B. subtilis, with strong correlation to triggering of Taxol biosynthesis. The intimate interaction of A. flavipes with B. subtilis strongly modulates the Taxol biosynthetic machinery of A. flavipes by modulating the chromatin remodeling. [ABSTRACT FROM AUTHOR]

Autoantibodies have been identified as key players in several neuroimmunological disorders such as autoimmune encephalitis, CNS demyelinating diseases, neuropathies, and neuromuscular junction disorders. Recent advances of proteomic technologies enable scientists to determine the target protein of autoantibodies. These methods are divided into three categories, namely, (1) mass spectrometry (MS)‐based method, (2) gene expression library screening, and (3) protein microarray. MS‐based methods can analyze posttranslationally modified proteins or conformational proteins, while gene expression library screening cannot. However, in MS‐based methods, protein‐abundant bias could prevent the identification of the target protein, and DNA sequencing could be more easily performed than MS‐based methods. Therefore, a sound knowledge of the advantages and limitations of each technique is required for selection of the optimal methodology for antigen identification in neuroimmunological disorders. [ABSTRACT FROM AUTHOR]

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a rapidly growing infectious disease, widely spread with high mortality rates. Since the release of the SARS-CoV-2 genome sequence in March 2020, there has been an international focus on developing target-based drug discovery, which also requires knowledge of the 3D structure of the proteome. Where there are no experimentally solved structures, our group has created 3D models with coverage of 97.5% and characterized them using state-of-the-art computational approaches. Models of protomers and oligomers, together with predictions of substrate and allosteric binding sites, protein-ligand docking, SARS-CoV-2 protein interactions with human proteins, impacts of mutations, and mapped solved experimental structures are freely available for download. These are implemented in SARS CoV-2 3D, a comprehensive and user-friendly database, available at https://sars3d.com/. This provides essential information for drug discovery, both to evaluate targets and design new potential therapeutics. [ABSTRACT FROM AUTHOR]

Purpose: Metastasis is an unavoidable event happened among almost all small cell lung cancer (SCLC) patients. However, the molecular driven factors have not been elucidated. Recently, a novel hydrolase called cell migration inducing hyaluronidase (CEMIP) triggered both migration and invasion in many tumors but not SCLC. Therefore, in this study, we verified that CEMIP promoted migration and invasion in SCLC and applied proteomics analysis to screen out potential target profiles and the signaling pathway related to CEMIP regulation. Method: Immunofluorescence was conducted to exam the expression of CEMIP on SCLC and paired adjacent normal tissues among enrollment. RT-qPCR and Western blot (WB) assays were conducted to valuate cellular protein and mRNA expression of CEMIP and EMT markers. Lentivirus-CEMIP-shRNAs and CEMIP plasmid were used for expression manipulating. Changes of cellular migration and invasion were tested through transwell assays. Tandem Mass Tag (TMT) peptide labeling coupled with LC–MS/MS was used for quantifying proteins affected by reducing expression of CEMIP on H446 cells. Results: The expression of CEMIP showed 1.64 ± 0.16-fold higher in SCLC tissues than their normal counterpart. Decreasing the expression of CEMIP on SCLC cells H446 regressed both cellular migration and invasion ability, whereas the promoting cellular migration and invasion was investigated through over-expressing CEMIP on H1688. Proteomic and bioinformatics analysis revealed that total 215 differentially expressed proteins (DEPs) that either their increasing or decreasing relative expression met threshold of 1.2-fold changes with p value ≤ 0.05. The dramatic up-regulated DEPs included an unidentified peptide sequence (encoded by cDNA FLJ52096) SPICE1 and CRYAB, while the expression of S100A6 was largely down-regulated. DEPs mainly enriched on caveolae of cellular component, calcium ion binding of biological process and epithelial cell migration of molecular function. KEGG enrichment indicated that DEPs mainly exerted their function on TGF-β, GABAergic synapse and MAPK signaling pathway. Conclusion: It is the first report illustrating that CEMIP might be one of the metastatic triggers in SCLC. And also, it provided possible molecular mechanism cue and potential downstream target on CEMIP-induced cellular migration and invasion on SCLC. [ABSTRACT FROM AUTHOR]

This study describes the optimization of a culture medium to improve protease production by Janthinobacterium sp. using response surface methodology (RSM) and comparative proteomic analysis. Using a one-variable-at-a-time approach, dextrose, Triton X-100, peptone, and pH were identified as significant factors affecting protease production. The optimized medium resulted in a 6.25-fold increase in protease activity when compared to that in the original medium. To investigate the metabolic changes in the optimized medium, extracellular and intracellular proteins were analyzed by two-dimensional gel electrophoresis. Some proteins essential to cell viability were highly expressed in the optimized medium and the outer membrane protein A was secreted at a higher level. These results can be useful for the industrial application of psychrophiles that produce cold-adapted enzymes. [ABSTRACT FROM AUTHOR]

One and two dimensional electrophoresis is a powerful technique for separation of proteins from complex biological sources. These are effective techniques of gene expression analysis at translational level especially for the species of which genome is not yet sequenced. High resolution electrophoresis separation of proteins from complex biological systems is crucial in protein science. Three protein extraction methods, such as Phenol, Multi detergent and TCA acetone were analyzed in the present investigation to find out appropriate protein extraction method amenable to one dimensional (1-D) and two dimensional (2-D) protein profiling of Haloxylon recurvum and H. salicornicum. Plant species selected for the present investigation are major components of saline and arid ecosystem and valuable source of food additives, nutritional ingredients, herbal medicines and fuel in the region. Phenolic components present in these plants are major hindrances for extraction of pure proteins. High concentrations of good quality proteins represented by clear, intense and more bands/spots in 1-D and 2-D gels were extracted with Phenol extraction method in comparison to TCA acetone and Multi detergent methods. This is possibly the first report on extraction and purification of proteins amenable to 1-D and 2-D proteome profiling of Haloxylon species of Indian arid zone, and forms a basis for further comparative proteome analysis of these two contrasting species with respect to salinity and drought tolerant ability for prospects. [ABSTRACT FROM AUTHOR]

Background: Modularity is important for evolutionary innovation. The recombination of existing units to form larger complexes with new functionalities spares the need to create novel elements from scratch. In proteins, this principle can be observed at the level of protein domains, functional subunits which are regularly rearranged to acquire new functions. Results: In this study we analyse the mechanisms leading to new domain arrangements in five major eukaryotic clades (vertebrates, insects, fungi, monocots and eudicots) at unprecedented depth and breadth. This allows, for the first time, to directly compare rates of rearrangements between different clades and identify both lineage specific and general patterns of evolution in the context of domain rearrangements. We analyse arrangement changes along phylogenetic trees by reconstructing ancestral domain content in combination with feasible single step events, such as fusion or fission. Using this approach we explain up to 70% of all rearrangements by tracing them back to their precursors. We find that rates in general and the ratio between these rates for a given clade in particular, are highly consistent across all clades. In agreement with previous studies, fusions are the most frequent event leading to new domain arrangements. A lineage specific pattern in fungi reveals exceptionally high loss rates compared to other clades, supporting recent studies highlighting the importance of loss for evolutionary innovation. Furthermore, our methodology allows us to link domain emergences at specific nodes in the phylogenetic tree to important functional developments, such as the origin of hair in mammals. Conclusions: Our results demonstrate that domain rearrangements are based on a canonical set of mutational events with rates which lie within a relatively narrow and consistent range. In addition, gained knowledge about these rates provides a basis for advanced domain-based methodologies for phylogenetics and homology analysis which complement current sequence-based methods. [ABSTRACT FROM AUTHOR]

Membrane lipid remodelling is one of the strategies that plants have developed to combat abiotic stress. In this study, physiological, lipidomic and proteome analyses were conducted to investigate the changes in glycerolipid and phospholipid concentrations in the wheat (Triticum aestivum L.) cultivars CIGM90.863 and Seri M82 under hypoxia treatment. The growth of CIGM90.863 remained unaffected, whereas Seri M82 was significantly stunted after 8 days of hypoxia treatment. The concentrations of all lipids except lysophosphatidylglycerol were significantly higher in the leaves of Seri M82 than in CIGM90.863 under normal growth conditions. The lipid profile changed significantly under hypoxia stress and varied between genotypes for some of the lipids. Phosphatidic acids remained unchanged in Seri M82 but they were gradually induced in CIGM90.863 in response to hypoxia stress because of the higher phospholipaseDexpression and lower expression of diglycerol kinase and phosphatidate phosphatases. In contrast, digalactosyldiacylglycerol content was highly stable in CIGM90.863 following hypoxia treatment, although it decreased significantly in Seri M82. Phosphatidylglycerol and lipoxygenase showed a stronger and faster response in CIGM90.863 than in Seri M82 under hypoxia stress. Different membrane lipid adjustments in wheat under oxygen deficiency conditions could be partly responsible for the differing tolerance of SeriM82and CIGM90.863. This study will help us to better understand howwheat tolerates hypoxia stress by regulating lipid remodelling. [ABSTRACT FROM AUTHOR]

Phytophthora capsica is a destructive oomycete plant pathogen that causes huge losses to crop production worldwide. However, the novel fungicide SYP-14288 has shown excellent activity against various stages of the oomycete life cycle as well against fungal plant pathogens. The current study utilized isobaric tags for relative and absolute quantitation technology to generate proteome profiles of P. capsici in the presence or absence of SYP-14288 in order to gain a greater understanding of the SYP-14288 mode of action. A total of 1,443 individual proteins were identified during the investigation, of which 599 were considered to have significantly altered expression. Further investigation using Cluster of Orthologous Groups of proteins analysis and Kyoto Encyclopedia of Genes and Genomes pathway analysis indicated most of the proteins with altered expression were associated with carbohydratemetabolism, energy metabolism and their downstream biological functions, especially with regard to oxidoreductase activity and subsequent adenosine triphosphate (ATP) production associated pathways. Quantitative expression analysis using qRT-PCR validated the proteomic data. These results seem to indicate that SYP-14288 treatment caused a shift in energy metabolism that resulted in the activation of compensatory mechanisms affecting carbohydrate and lipid metabolism. The study also found evidence that the up-regulation of transmembrane transporters and proteins associated with stress response might also be coopted to compensate for the disrupted proton gradient and other downstream effects. Taken together these results provide strong evidence that SYP-14288 has a similar mode of action to the oxidative phosphorylation uncoupler Fluazinam but further investigation, including molecular studies, is required to completely characterize the SYP-14288 mode of action in P. capsici. However, the proteomic data collected in the current study does provide important insight into the overall effect of SYP-14288 in P. capsici, which could be useful for the registration and application of this novel fungicide. [ABSTRACT FROM AUTHOR]

Herpes simplex virus 1 (HSV-1) is a predominant cause of herpes simplex encephalitis (HSE), leading to a high mortality rate and severe neurological sequelae worldwide. HSE is typically accompanied by the blood-brain barrier (BBB) disruption, but the underlying mechanisms are unclear. To explore the disruption mechanisms of the BBB, quantitative analysis of the cellular proteome was carried out to investigate the proteomic changes that occur after infection. In this study, bEnd.3 cells were infected with HSV-1, followed by liquid chromatography-tandem mass spectrometry. A total of 6761 proteins were identified in three independent mass spectrometry analyses. Compared to the uninfected cells, 386 and 293 differentially expressed proteins were markedly upregulated or downregulated, respectively. Bioinformatic analysis showed that the activator protein-1 factor, including Fos, Jun, and ATF family proteins and cell adhesion molecules were significantly changed. Further validation of the changes observed for these proteins was carried out by western blotting and quantitative real-time PCR. Transendothelial electrical resistance (TEER) studies were performed to explore the effects of ATF3, Fra1, or JunB overexpression on the function of bEnd.3 cells. Characterization of the differential expression of these proteins in bEnd.3 cells will facilitate further exploration of BBB disruption upon HSV-1 infection. [ABSTRACT FROM AUTHOR]

We aimed to understand the molecular-level changes occurring in the photosynthetic metabolic network in mutant chloroplasts. We performed comparative liquid chromatography-mass spectrometry protein profiling of wild-type (WT) and chlorophyll-deficient melon (Cucumis melo L.) mutants. We identified 390 differentially expressed proteins and 81 shared proteins varied significantly in abundance, of which 76 were upregulated and 5 were downregulated. Differentially expressed proteins were involved in the following biological processes: binding, catalytic, structural, transporter, and antioxidant activities. The mutant had 6.08-fold higher expression of glutamate-1-semialdehyde 2,1-aminomutase (GSAM), an enzyme that synthesizes the chlorophyll precursor, 5-aminolevulinic acid, and a 5.02-fold higher expression of pyridoxal biosynthesis protein, a GSAM coenzyme. An RNA recognition motif-containing protein (RRM) decreased in expression by 5.22-fold. This suggests that GSAM and RRM are particularly relevant to chlorophyll deficiency. [ABSTRACT FROM AUTHOR]

A proteomic analysis of the venom of males and females of the Naja kaouthia monocled cobra specimens kept in captivity was carried out. Using the amino acid sequences of proteins of the taxonomic group Serpentes from the UniProt KB database, 875 proteins were identified in the venom samples and the relative content of about 190 of them was determined in each venom sample by the method of label-free quantitative proteomics. The total content of 35 major protein components of the venom of each individual was shown to comprise about 98% of the total amount of venom proteins. Analysis of relative content of the venom proteins in males and females showed a significantly (p < 0.05) higher content of nerve growth factor and natriuretic peptide in the venom of females. Analysis of the profile of posttranslational modifications of N. kaouthia toxins revealed previously unknown sites for phosphorylation, acetylation, and formylation. [ABSTRACT FROM AUTHOR]

Typically, nitrification is a two-stage microbial process and is key in wastewater treatment and nutrient recovery from waste streams. Changes in salinity represent a major stress factor that can trigger response mechanisms, impacting the activity and the physiology of bacteria. Despite its pivotal biotechnological role, little information is available on the specific response of nitrifying bacteria to varying levels of salinity. In this study, synthetic communities of ammonia-oxidizing bacteria (AOB Nitrosomonas europaea and/or Nitrosomonas ureae) and nitrite-oxidizing bacteria (NOB Nitrobacter winogradskyi and/or Nitrobacter vulgaris) were tested at 5, 10, and 30 mS cm-1 by adding sodium chloride to the mineral medium (0, 40, and 200 mM NaCl, respectively). Ammonia oxidation activity was less affected by salinity than nitrite oxidation. AOB, on their own or in combination with NOB, showed no significant difference in the ammonia oxidation rate among the three conditions. However, N. winogradskyi improved the absolute ammonia oxidation rate of both N. europaea and N. ureae. N. winogradskyi 's nitrite oxidation rate decreased to 42% residual activity upon exposure to 30 mS cm-1, also showing a similar behavior when tested with Nitrosomonas spp. The nitrite oxidation rate of N. vulgaris , as a single species, was not affected when adding sodium chloride up to 30 mS cm-1, however, its activity was completely inhibited when combined with Nitrosomonas spp. in the presence of ammonium/ammonia. The proteomic analysis of a co-culture of N. europaea and N. winogradskyi revealed the production of osmolytes, regulation of cell permeability and an oxidative stress response in N. europaea and an oxidative stress response in N. winogradskyi , as a result of increasing the salt concentration from 5 to 30 mS cm-1. A specific metabolic response observed in N. europaea suggests the role of carbon metabolism in the production of reducing power, possibly to meet the energy demands of the stress response mechanisms, induced by high salinity. For the first time, metabolic modifications and response mechanisms caused by the exposure to salinity were described, serving as a tool toward controllability and predictability of nitrifying systems exposed to salt fluctuations. [ABSTRACT FROM AUTHOR]

Present study was conducted to explore the role of exogenous salicylic acid (SA), sodium nitroprusside (SNP), abscisic acid (ABA) and proline (PRO) in mitigating high-temperature (HT) induced oxidative stress in different Lablab purpureus L. cultivars. The attempt was made to examine whether these phytohormones, when applied exogenously, were able to regulate plant morpho-physiological behavior by modulating genes and proteins involved in antioxidant defense system. The HT stress induced membrane damage, degraded chlorophyll, generated redox metabolites and significantly reduced growth and biomass in all the cultivars. Among all the four treatments, foliar application of SA and SNP were most effective in the regulation of growth and physiological processes of the cultivars compared to ABA and PRO applications. Thus, signifying the protective role of SA and SNP in mitigation of HT induced oxidative stress and conferring HT stress tolerance in the cultivars. Gene expression and leaf proteome analysis revealed that these phytohormones were also involved in regulation of defense related gene expression, stress inducible proteins and de novo synthesis of specific proteins under HT stress. The experimental findings depict that foliar applications of SA and SNP enhances HT stress tolerance in lablab cultivars by modulating antioxidant defense system and by regulating bio-physical growth more effectively as compared to ABA and PRO application. [ABSTRACT FROM AUTHOR]

Finding the keys to understanding the infectious process of Yersinia ruckeri was not a priority for many years due to the prompt development of an effective biotype 1 vaccine which was used mainly in Europe and USA. However, the gradual emergence of outbreaks in vaccinated fish, which have been reported since 2003, has awakened interest in the mechanism of virulence in this pathogen. Thus, during the last two decades, a large number of studies have considerably enriched our knowledge of many aspects of the pathogen and its interaction with the host. By means of both conventional and a variety of novel strategies, such as cell GFP tagging, bioluminescence imaging and optical projection tomography, it has been possible to determine three putative Y. ruckeri infection routes, the main point of entry for the bacterium being the gill lamellae. Moreover, a wide range of potential virulence factors have been highlighted by specific gene mutagenesis strategies or genome-wide transposon/plasmid insertion-based screening approaches, such us in vivo expression technology (IVET) and signature tagged mutagenesis (STM). Finally, recent proteomic and whole genomic analyses have allowed many of the genes and systems that are potentially implicated in the organism's pathogenicity and its adaptation to the host environmental conditions to be elucidated. Altogether, these studies contribute to a better understanding of the infectious process of Y. ruckeri in fish, which is crucial for the development of more effective strategies for preventing or treating enteric redmouth disease (ERM). [ABSTRACT FROM AUTHOR]

Although the physiological consequences of plant growth under saline conditions have been well described, understanding the core mechanisms conferring plant salt adaptation has only started. We target the root plasma membrane proteomes of two barley varieties, cvs. Steptoe and Morex, with contrasting salinity tolerance. In total, 588 plasma membrane proteins were identified by mass spectrometry, of which 182 were either cultivar or salinity stress responsive. Three candidate proteins with increased abundance in the tolerant cv. Morex were involved either in sterol binding (a GTPase-activating protein for the adenosine diphosphate ribosylation factor [ZIGA2], and a membrane steroid binding protein [MSBP]) or in phospholipid synthesis (phosphoethanolamine methyltransferase [PEAMT]). Overexpression of barley MSBP conferred salinity tolerance to yeast cells, whereas the knock-out of the heterologous AtMSBP1 increased salt sensitivity in Arabidopsis. Atmsbp1 plants showed a reduced number of lateral roots under salinity, and root-tip-specific expression of barley MSBP in Atmsbp1 complemented this phenotype. In barley, an increased abundance of MSBP correlates with reduced root length and lateral root formation as well as increased levels of auxin under salinity being stronger in the tolerant cv. Morex. Hence, we concluded the involvement of MSBP in phytohormone-directed adaptation of root architecture in response to salinity. [ABSTRACT FROM AUTHOR]

Ulcerative colitis (UC) is a chronic, relapsing, inflammatory bowel disease, and patients with long-standing UC are at high risk of developing colorectal cancer as a typical case of the organ-specific chronic inflammation-carcinoma sequence. Interactions between epithelial and stromal cells and alterations in a variety of stromal microenvironments have been demonstrated to have important roles in the carcinogenesis of UC-associated carcinoma. Therefore, the identification of proteins in the inflammatory microenvironment is important not only in the epithelium, however also in the stroma of UC inflammatory foci. To identify proteins associated with UC-associated carcinoma, the present study used proteomic analysis with two-dimensional electrophoresis and mass spectrometry. Differentially expressed proteins were assessed between active and inactive UC biopsy specimens. Results were verified by immunohistochemistry. Peroxiredoxin 1 (PRDX1) was among the proteins identified to have increased expression in active compared with inactive UC. Immunohistochemical analysis indicated that the expression of both PRDX1 and thioredoxin (TRX) increased with increasing inflammation grade in epithelial cells in UC mucosal crypts. PRDX1-positive stromal cells in the lower half of the lamina propria increased along with colitis severity. Furthermore, the expression of both PRDX1 and TRX proteins was increased in UC-associated neoplastic lesions compared with normal mucosa. A stepwise increase in PRDX1 expression was clear with increasing tumor progression in UC-associated tumorigenesis. Since PRDX1 and TRX overexpression was a unique characteristic of UC activity and UC-associated neoplastic lesions, PRDX1 and TRX expression may reflect oxidative stress along with the severity of colitis activity and UC-associated tumorigenesis in patients with UC. [ABSTRACT FROM AUTHOR]

Background. In spite of its invasive nature and risks, kidney biopsy is currently required for precise diagnosis of many chronic kidney diseases (CKDs). Here, we explored the hypothesis that analysis of the urinary proteome can discriminate different types of CKD irrespective of the underlying mechanism of disease. Methods. We used data from the proteome analyses of 1180 urine samples from patients with different types of CKD, generated by capillary electrophoresis coupled to mass spectrometry. A set of 706 samples served as the discovery cohort, and 474 samples were used for independent validation. For each CKD type, peptide biomarkers were defined using statistical analysis adjusted for multiple testing. Potential biomarkers of statistical significance were combined in support vector machine (SVM)- based classifiers. Results. For seven different types of CKD, several potential urinary biomarker peptides (ranging from 116 to 619 peptides) were defined and combined into SVM-based classifiers specific for each CKD. These classifiers were validated in an independent cohort and showed good to excellent accuracy for discrimination of one CKD type from the others (area under the receiver operating characteristic curve ranged from 0.77 to 0.95). Sequence analysis of the biomarkers provided further information that may clarify the underlying pathophysiology. Conclusions. Our data indicate that urinary proteome analysis has the potential to identify various types of CKD defined by pathological assessment of renal biopsies and current clinical practice in general. Moreover, these approaches may provide information to modelmolecular changes per CKD. [ABSTRACT FROM AUTHOR]

Background. Chronic kidney disease (CKD) progression is currently assessed by a decline in estimated glomerular filtration rate (eGFR) and/or an increase in urinary albumin excretion (UAE). However, these markers are considered either to be latestage markers or to have low sensitivity or specificity. In this study, we investigated the performance of the urinary proteome-based classifier CKD273, compared with UAE, in a number of different narrow ranges of CKD severity, with each range separated by an eGFR of 10 mL/min/1.73 m2. Methods. A total of 2672 patients with different CKD stages were included in the study. Of these, 394 individuals displayed a decline in eGFR of >5 mL/min/1.73 m2/year (progressors) and the remaining individuals were considered non-progressors. For all samples, UAE values and CKD273 classification scores were obtained. To assess UAE values and CKD273 scores at different disease stages, the cohort was divided according to baseline eGFRs of ≥80, 70-79, 60-69, 50-59, 40-49, 30-39 and <29 mL/min/1.73 m2. In addition, areas under the curve for CKD273 and UAE were calculated. Results. In early stage CKD, the urinary proteome-based classifier performed significantly better than UAE in detecting progressors. In contrast, UAE performed better in patients with late-stage CKD. No significant difference in performance was found between CKD273 and UAE in patients with moderately reduced renal function. Conclusions. These results suggest that urinary peptides, as combined in the CKD273 classifier, allow the detection of progressive CKD at early stages, a point where therapeutic intervention is more likely to be effective. However, late-stage disease, where irreversible damage of the kidney is already present, is better detected by UAE. [ABSTRACT FROM AUTHOR]

Conjugation of the small ubiquitin-related modifier ( SUMO) to protein substrates has an impact on stress responses and on development. We analyzed the proteome and phosphoproteome of mutants in this pathway. The mutants chosen had defects in SUMO ligase SIZ1, which catalyzes attachment of single SUMO moieties onto substrates, and in ligases PIAL1 and PIAL2, which are known to form SUMO chains. A total of 2657 proteins and 550 phosphopeptides were identified and quantified. Approximately 40% of the proteins and 20% of the phosphopeptides showed differences in abundance in at least one of the analyzed genotypes, demonstrating the influence of SUMO conjugation on protein abundance and phosphorylation. The data show that PIAL1 and PIAL2 are integral parts of the SUMO conjugation system with an impact on stress response, and confirm the involvement of SIZ1 in plant defense. We find a high abundance of predicted SUMO attachment sites in phosphoproteins (70% versus 40% in the total proteome), suggesting convergence of phosphorylation and sumoylation signals onto a set of common targets. [ABSTRACT FROM AUTHOR]

Key message: For the first time, a comprehensive proteome analysis was conducted on Brachypodium leaves under drought stress. Gradual changes in response to drought stress were monitored. Abstract: Drought is one of the major stress factors that dramatically affect the agricultural productivity worldwide. Improving the yield under drought is an urgent challenge in agriculture. Brachypodium distachyon is a model species for monocot plants such as wheat, barley and several potential biofuel grasses. In the current study, a comprehensive proteome analysis was conducted on Brachypodium leaves under different levels of drought application. To screen gradual changes upon drought, Brachypodium leaves subjected to drought for 4, 8 and 12 days were collected for each treatment day and relative water content of the leaves was measured for each time point. Cellular responses of Brachypodium were investigated through a proteomic approach involving two dimensional difference gel electrophoresis (2D-DIGE) and mass spectrometry (MS). Among 497 distinct spots in Brachypodium protein repertoire, a total of 13 differentially expressed proteins (DEPs) were identified as responsive to drought by mass spectrometry and classified according to their functions using bioinformatics tools. The biological functions of DEPs included roles in photosynthesis, protein folding, antioxidant mechanism and metabolic processes, which responded differentially at each time point of drought treatment. To examine further transcriptional expression of the genes that code identified protein, quantitative real time PCR (qRT-PCR) was performed. Identified proteins will contribute to the studies involving development of drought-resistant crop species and lead to the delineation of molecular mechanisms in drought response. [ABSTRACT FROM AUTHOR]

Leaf variegation is often the focus of plant breeding. Here, we studied a variegated mutant of Phalaenopsis aphrodite subsp. formosana, which is usually used as a parent of horticultural breeding, to understand its anatomic and genetic regulatory mechanisms in variegation. Chloroplasts with well-organized thylakoids and starch grains were found only in the mesophyll cells of green sectors but not of yellow sectors, confirming that the variegation belongs to the chlorophyll type. The two-dimensional electrophoresis and LC/MS/MS also reveal differential expressions of PsbP and PsbO between the green and yellow leaf sectors. Full-length cDNA sequencing revealed that mutant transcripts were caused by intron retention. When conditioning on the total RNA expression, we found that the functional transcript of PsbO and mutant transcript of PsbP are higher expressed in the yellow sector than in the green sector, suggesting that the posttranscriptional regulation of PsbO and PsbP differentiates the performance between green and yellow sectors. Because PsbP plays an important role in the stability of thylakoid folding, we suggest that the negative regulation of PsbP may inhibit thylakoid development in the yellow sectors. This causes chlorophyll deficiency in the yellow sectors and results in leaf variegation. We also provide evidence of the link of virus CymMV and the formation of variegation according to the differential expression of CymMV between green and yellow sectors. [ABSTRACT FROM AUTHOR]

Sphingomonas elodeais a Gram-negative bacterium capable of producing ‘gellan gum’ exopolysaccharide that is the most extensively studied expolysaccharides of microbial origin. In this study, we investigated the phenotypic and proteomic alterations inS. elodeaby homogeneously expressing bothgelAandgelNinvolved in positive regulation and extracellular secretion of metabolites in gellan biosynthesis, respectively. Expression of six histidine-tagged GelA and GelN was determined by Western blot analysis. Successful expression of GelA and GelN resulted in both morphological changes of colonies and enhanced secretion of gellan into the growth medium (GelA, 21.2% more and GelN, 48.3% more) overexpressed compared to the wile-type. Comparative two-dimensional gel electrophoresis analysis revealed a differential proteome expression inS. elodeaoverexpressing GelA and GelN. Proteins up- or down-regulated by GelA and GelN overexpression were found to be mainly sugar transportation proteins, two-component regulatory proteins, and proteins involved in secretion pathways. The results suggest that the effect of GelA and GelN overexpression on gellan biosynthesis might be mainly caused by increased transportation of sugar units or enhanced exportation of gellan. [ABSTRACT FROM PUBLISHER]

Background: Chromium (Cr) being multifarious industrial used element, is considered a potential environmental threat. Cr found to be a prospective water and soil pollutant, and thus it is a current area of concern. Oilseed rape (Brassica napus L.) is well known as a major source of edible oil around the globe. Due to its higher growth, larger biomass and capability to uptake toxic materials B. napus is considered a potential candidate plant against unfavorable conditions. To date, no study has been done that described the Cr and GSH mechanism at RNA-Seq level. Results: Both digital gene expression (DGE) and transcriptome profile analysis (TPA) approaches had opened new insights to uncover the several number of genes related to Cr stress and GSH alleviating mechanism in two leading cultivars (ZS 758 and Zheda 622) of B. napus plants. Data showed that Cr inhibited KEGG pathways i.e. stilbenoid, diarlyheptanoid and gingerol biosynthesis; limonene and pentose degradation and glutathione metabolism in ZS 758; and ribosome and glucosinolate biosynthesis in Zheda-622. On the other hand, vitamin B6, tryptophan, sulfur, nitrogen and fructose and manose metabolisms were induced in ZS 758, and zeatin biosynthesis, linoleic acid metabolism, arginine and proline metabolism, and alanine, asparate and glutamate metabolism pathways in Zheda 622. Cr increased the TFs that were related to hydralase activity, antioxidant activity, catalytic activity phosphatase and pyrophosphatase activity in ZS 758, and vitamin binding and oxidoreductase activity in Zheda 622. Cr also up-regulated the promising proteins related to intracellular membrane bounded organelles, nitrile hyrdatase activity, cytoskeleton protein binding and stress response. It also uncovered, a novel Cr-responsive protein (CL2535.Contig1_All) that was statistically increased as compared to control and GSH treated plants. Exogenously applied GSH successfully not only recovered the changes in metabolic pathways but also induced cysteine and methionine metabolism in ZS 758 and ubiquinone and other terpenoid-quinone biosynthesis pathways in Zheda 622. Furthermore, GSH increased the level of TFs i.e. the gene expression of antioxidant and catalytic activities, iron ion binding and hydrolase activity as compared with Cr. Moreover, results pointed out a novel GSH responsive protein (CL827.Contig3_All) whose expression was found to be significantly increased when compared than Cr stress. Results further delineated that GSH induced TFs such as glutathione disulphide oxidoreducatse and aminoacyl-tRNA ligase activity, and beta glucosidase activity in ZS 758. Similarly in Zheda 622, GSH induced the TFs for instance DNA binding and protein dimerization activity. GSH also highlighted the proteins that were involved in transportation, photosynthesis process, RNA polymerase activity, and against the metal toxicity. These results indicated that cultivar ZS 758 had better metabolism and showed higher tolerance against Cr toxicity.Conclusion: The responses of ZS 758 and Zheda 622 differed considerably at both physiological and transcriptional level. Moreover, RNA-Seq method explored the hazardous behavior of Cr as well as GSH up-regulating mechanism by activating plant metabolism, stress responsive genes, TFs and protein encyclopedia. [ABSTRACT FROM AUTHOR]

Although the specific function of SCO2127 remains elusive, it has been assumed that this hypothetical protein plays an important role in carbon catabolite regulation and therefore in antibiotic biosynthesis in Streptomyces coelicolor. To shed light on the functional relationship of SCO2127 to the biosynthesis of actinorhodin, a detailed analysis of the proteins differentially produced between the strain M145 and the Δsco2127 mutant of S. coelicolor was performed. The delayed morphological differentiation and impaired production of actinorhodin showed by the deletion strain were accompanied by increased abundance of gluconeogenic enzymes, as well as downregulation of both glycolysis and acetyl-CoA carboxylase. Repression of mycothiol biosynthetic enzymes was further observed in the absence of SCO2127, in addition to upregulation of hydroxyectoine biosynthetic enzymes and SCO0204, which controls nitrite formation. The data generated in this study reveal that the response regulator SCO0204 greatly contributes to prevent the formation of actinorhodin in the ∆sco2127 mutant, likely through the activation of some proteins associated with oxidative stress that include the nitrite producer SCO0216. [ABSTRACT FROM AUTHOR]