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N-glycosylation is a major post-translational modification of proteins and involved in many diseases, however, the state and role of N-glycosylation in cartilage degeneration of osteonecrosis of femoral head (ONFH) remain unclear. The aim of this study is to identify the glycoproteins of ONFH hip cartilage. Cartilage tissues were collected from nine patients with ONFH and nine individuals with traumatic femoral neck fracture. Cartilage glycoproteins were identified by glycoproteomics based on LC-MS/MS. The differentially N-glycoproteins including glycosites were identified in ONFH and controls. A total of 408 N-glycoproteins with 444 N-glycosites were identified in ONFH and control cartilage. Among them, 104 N-glycoproteins with 130 N-glycosites were significantly differential in ONFH and control cartilage, which including matrix-remodeling-associated protein 5, prolow-density lipoprotein receptor-related protein 1, clusterin and lysosome-associated membrane glycoprotein 2. Gene Ontology analysis revealed the significantly differential glycoproteins mainly belonged to protein metabolic process, single-multicellular organism process, proteolysis, biological adhesion and cell adhesion. KEGG pathway and protein-protein interaction analysis suggested that the significantly differential glycoproteins were associated with PI3K-Akt signalling pathway, ECM-receptor interaction, protein processing in the endoplasmic reticulum and N-glycan biosynthesis. This information provides substantial insight into the role of protein glycosylation in the development of cartilage degeneration of ONFH patients.

Fish oil is an important source of lipid in aquafeeds, but is prone to oxidation. In aquatic species, oxidized fish oil may induce oxidative stress. Although the intestine is an essential nutritional and immunological organ in juvenile hybrid grouper (Epinephelus fuscoguttatus × Epinephelus lanceolatus), the cause of its oxidative stress remains unknown. In the current research, a 65-day rearing experiment with diets containing 0% (R), 3% (L), 6% (M), or 9% (H) oxidized fish oil was conducted to explore the effects of oxidized fish oil on the antioxidant ability, histology, and transcriptome of the intestine in juvenile hybrid grouper. The concentrations of vitamin E and antioxidant enzymes including superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-Px) in the foregut, midgut, and hindgut decreased significantly with the level of oxidized fish oil in the diet rose, whereas malondialdehyde (MDA) concentration increased. The oxidative stress damaged intestinal tissue structure. In detail, the goblet cell in H group was significantly reduced and the microvilli was depauperated. Compared with group R, the oxidized stress altered the intestinal transcriptome map in H group. A total of 29,360 unigenes had been annotated in all databases. There were 18 differentially expressed genes (DEGs), with 12 being up-regulated and 6 being down-regulated. According to the GO enrichment, the functions of differential genes were mostly enriched in primary metabolic process, cellular biosynthetic process, organic substance biosynthetic process, cellular macromolecule metabolic process, and protein metabolic process. There were 2 significant pathways including sulfur metabolism pathways and p53 signaling pathways involving cytochrome c (Cyt2c) gene with a high expression state under oxidative stress. In summary, oxidized fish oil induced intestinal oxidative stress. Meanwhile, oxidative stress was destroying the structure of the gut tissue. Furthermore, oxidative stress altered the intestinal transcriptome map.

Saussurea involucrata (S. involucrata) had been reported to have anti-hepatoma function. However, the mechanism is complex and unclear. To evaluate the anti-hepatoma mechanism of S. involucrata comprehensively and make a theoretical basis for the mechanical verification of later research, we carried out this work. In this study, the total phenolic acids from S. involucrata determined by a cell suspension culture (ESPI) was mainly composed of 4,5-dicaffeoylquinic acid, according to the LC-MS analysis. BALB/c nude female mice were injected with HepG2 cells to establish an animal model of liver tumor before being divided into a control group, a low-dose group, a middle-dose group, a high-dose group, and a DDP group. Subsequently, EPSI was used as the intervention drug for mice. Biochemical indicators and differences in protein expression determined by TMT quantitative proteomics were used to resolve the mechanism after the low- (100 mg/kg), middle- (200 mg/kg), and high-dose (400 mg/kg) interventions for 24 days. The results showed that EPSI can not only limit the growth of HepG2 cells in vitro, but also can inhibit liver tumors significantly with no toxicity at high doses in vivo. Proteomics analysis revealed that the upregulated differentially expressed proteins (DE proteins) in the high-dose group were over three times that in the control group. ESPI affected the pathways significantly associated with the protein metabolic process, metabolic process, catalytic activity, hydrolase activity, proteolysis, endopeptidase activity, serine-type endopeptidase activity, etc. The treatment group showed significant differences in the pathways associated with the renin-angiotensin system, hematopoietic cell lineage, etc. In conclusion, ESPI has a significant anti-hepatoma effect and the potential mechanism was revealed.

Type 2 diabetes mellitus (T2DM) is the most common endocrine disorder which poses a serious threat to human health. This investigation aimed to screen the candidate genes differentially expressed in T2DM by bioinformatics analysis. The expression profiling by high throughput sequencing of GSE81608 dataset was retrieved from the gene expression omnibus (GEO) database and analyzed to identify the differentially expressed genes (DEGs) between T2DM and normal controls. Then, Gene Ontology (GO) and pathway enrichment analysis, protein-protein interaction (PPI) network, modules, miRNA-hub gene regulatory network construction and TF-hub gene regulatory network construction, and topological analysis were performed. Receiver operating characteristic curve (ROC) analysis was also performed to verify the diagnostics value and expression of identified hub genes. A total of 927 DEGs (461 were up regulated and 466 down regulated genes) were identified in T2DM. GO and REACTOME results showed that DEGs mainly enriched in protein metabolic process, establishment of localization, metabolism of proteins and metabolism. The top centrality hub genes APP, MYH9, TCTN2, USP7, SYNPO, GRB2, HSP90AB1, UBC, HSPA5 and SQSTM1 were screened out as the critical genes among the DEGs from the PPI network, modules, miRNA-hub gene regulatory network construction and TF-hub gene regulatory network. ROC analysis provide diagnostics value of hub genes. This study identified key genes, signal pathways and therapeutic agents, which might help us, improve our understanding of the mechanisms of HGPS and identify some new therapeutic agents for T2DM.

Enrofloxacin is widely used for the prevention and control of bacterial diseases in aquaculture. The liver is crucial for enrofloxacin metabolism, but enrofloxacin can induce liver damage. Herein, we explored proteomic changes in the liver of grass carp (Ctenopharyngodon idellus) following treatment with enrofloxacin using isobaric tag for relative and absolute quantitation (iTRAQ) technology. All experiments included two biological replicates and blank controls. Among the 3082 proteins identified, 103 were differentially abundant, comprising 49 up- and 54 downregulated proteins. Gene Ontology (GO) annotation identified macromolecular complex (63.60%), intracellular non-membrane-bound organelle (51.50%), and non-membrane-bound organelle (51.50%) as the most enriched cellular component terms. Structural molecule activity (26.80%), structural constituent of ribosome (17.90%), and calcium ion binding (16.10%) were the top three molecular function terms. Organic substance biosynthetic process (37.80%), biosynthetic process (37.80%), and protein metabolic process (37.80%) were the top three biological process terms. The Kyoto Encyclopaedia of Genes and Genomes (KEGG) pathway analysis found 17 enriched KEGG pathways, with protein digestion and absorption, extracellular matrix (ECM)–receptor interactions, and ribosome and focal adhesion the most significant (p

To investigate the different effects of dietary α-linolenic acid (ALA) and linoleic acid (LA) on the euryhaline fish Japanese seabass, a feeding trial followed by hepatic transcriptome assay was conducted. Two experimental diets containing 10% LA-rich sunflower seed oil (diet LA) or 10% ALA-rich perilla oil (diet ALA) were used in the feeding trial. LA and ALA in diets were characteristically incorporated into fish tissues while no significant difference was observed in growth performance and body proximate composition between groups LA and ALA. Compared to LA, ALA up-regulated transcription of 49 unigenes and down-regulated those of 311 unigenes. Quantitative RT-PCR studies on eight lipid metabolism-related genes and seven randomly selected genes were conducted to validate the transcriptomic results. Lipid metabolism-related genes ApoA1, ApoA4, ApoE, FABP1, FABP3, FABP4, FATP6, and DGAT1, as well as ribosomal proteins L9e, L13e, and S4e, were transcriptionally down-regulated by ALA. The differentially expressed genes (DEGs) were primarily enriched in Gene Ontology terms such as Lipid transport, Protein metabolic process, and Ribosome biogenesis, as well as in KEGG pathways such as Complement and coagulation cascades and Ribosome. The Protein-Protein Interaction (PPI) network based on the peptide biosynthesis-related DEGs showed that ribosomal proteins such as SAe, L4e, S4e, L15e, L9e, and L13Ae had high betweenness centrality in the dietary regulation of peptide biosynthetic processes. In conclusion, under the present experimental conditions, a high level of dietary α-linolenic acid tended to suppress lipid transport and protein biosynthetic processes in the liver of Japanese seabass at the gene expression level.

Allergic bronchopulmonary aspergillosis (ABPA) constantly develops in asthmatics, which has not been fully investigated. This study aimed to investigate serum differentially expressed proteins (DEPs) between ABPA and asthma using the new approach isobaric tags by relative and absolute quantitation (iTRAQ). Each 16 serum samples from ABPA or asthmatic subjects were pooled and screened using iTRAQ. After bioinformatic analysis, five candidate DEPs were validated in the enlarged serum samples from additional 21 ABPA, 31 asthmatic and 20 healthy subjects using ELISA. A receiver operating characteristic (ROC) curve was used to estimate the diagnostic power of carnosine dipeptidase 1 (CNDP1). A total of 29 DEPs were screened out between ABPA and asthmatic groups. Over half of them were enriched in proteolysis and regulation of protein metabolic process. Further verification showed serum levels of immunoglobulin heavy constant gamma 1, α-1-acid glycoprotein 1, corticosteroid-binding globulin and vitronectin were neither differentially altered between ABPA and asthma nor consistent with the proteomic analysis. Only serum CNDP1 was significantly decreased in ABPA patients, compared with asthmatics and healthy controls (P

Proton beam irradiation is a next-generation technique to develop mutant crop varieties. The mutagenic effects and molecular mechanisms of radiation are important multi-disciplinary research subjects. This study was conducted to investigate the types of mutations induced in the soybean genome by proton beam irradiation. In total, 22 plants, including 10 M2 plants treated with proton beam irradiation at 118 and 239 Gy, each, and two wild-type plants (Daepung) were sequenced by genotyping-by-sequencing (GBS). In total, 7453 single nucleotide polymorphisms (SNPs) were detected in the 20 M2 plants, compared with the two wild-type controls. The SNP frequency was 1/36,976 bp with proton beam irradiation at 118 Gy, and 1/32,945 bp at 239 Gy. Of these, 3569 SNPs were detected in genic regions. We observed that proton beam irradiation induced more substitutions than small insertion–deletions (INDELs). Based on the mutagenic effect of proton beam irradiation, the frequency of transition mutations was shown to be higher than that of transversions. The proton beam-induced SNPs were distributed uniformly in most of the chromosomes. Gene ontology (GO) analysis showed that there were many genes involved in protein metabolic process under biological process, intracellular membrane-bounded organelle under cellular component, and nucleic acid binding under molecular function. This study could provide valuable information for investigating the potential mechanisms of mutation, and guidance for developing soybeans cultivars using mutation breeding.

Inflammatory bowel disease (IBD) is an idiopathic prolonged ailment accountable for inflammatory conditions of the intestine. Moreover, arthritis is responsible for joints' stiffness and painful inflammation. IBD shows certain articular extra-intestinal manifestations associating IBD with arthritis. IBD associated arthritis is found to be linked with ankylosing spondylitis (AS). The present study insights for the potential and putative drug targets and biomarkers of IBD associated with arthritis using in silico approaches. Microarray data analysis of datasets involving IBD affected and AS affected vs controls were done to explore the differentially expressed genes (DEGs). In majority of the datasets, the common DEGs found were sterile alpha motif domain containing 9 like (SAMD9L), inhibin beta A subunit (INHBA), transmembrane protein 45A (TMEM45A) and transmembrane and tetratricopeptide repeat containing 1 (TMTC1). The common functions and pathways found between the DEGs were control of macromolecule metabolism process, control of metabolic process, control of primary metabolic process, and control of protein metabolic process, cell differentiation, organ development, single-organism development process, multicellular organism development process, development of system, single-multicellular organism development process, developmental process, development of anatomical structure, multicellular organismal development process, control of biological process, cell proliferation, hematopoietic progenitor cell differentiation and immune system process. TMTC1 and INBHA were found to be more biologically significant genes according to the topological properties of the network. This study also suggests that TMTC1, INBHA, TMEM45A and SAMD9L DEGs and their accompanying pathways might have the potential to be exploited as drug targets and biomarkers in the diagnosis and/or treatment of IBD linked arthritis and warrants for further experimental validation.

Background: Monocytes are critical components, not only for innate immunity, but also for the activation of the adaptive immune system. Many studies in animals and humans have demonstrated that monocytes may be closely associated with chronic inflammatory diseases and be proved to be pivotal in the association between high-intensity exercise and anti-inflammation response. However, the underlying molecular mechanisms driving this are barely understood. The present study aimed to screen for potential hub genes and candidate signaling pathways associated with the effects of high-intensity exercise on human monocytes through bioinformatics analysis. Materials and Methods: The GSE51835 gene expression dataset was downloaded from the Gene Expression Omnibus database. The dataset consists of 12 monocyte samples from two groups of pre-exercise and post-exercise individuals. Identifying differentially expressed genes (DEGs) with R software, and functional annotation and pathway analyses were then performed with related web databases. Subsequently, a protein–protein interaction (PPI) network which discovers key functional protein and a transcription factors-DEGs network which predicts upstream regulators were constructed. Results: A total of 146 differentially expressed genes were identified, including 95 upregulated and 51 downregulated genes. Gene Ontology analysis indicated that in the biological process functional group, these DEGs were mainly involved in cellular response to hydrogen peroxide, response to unfolded protein, negative regulation of cell proliferation, cellular response to laminar fluid shear stress, and positive regulation of protein metabolic process. The top five enrichment pathways in a Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were the FoxO signaling pathway, protein processing in the endoplasmic reticulum, influenza A, the ErbB signaling pathway, and the MAPK signaling pathway. TNF, DUSP1, ATF3, CXCR4, NR4A1, BHLHE40, CDKN1B, SOCS3, TNFAIP3, and MCL1 were the top 10 potential hub genes. The most important modules obtained in the PPI network were performed KEGG pathway analysis, which showed that these genes were mainly involved in the MAPK signaling pathway, the IL-17 signaling pathway, the TNF signaling pathway, osteoclast differentiation, and apoptosis. A transcription factor (TF) target network illustrated that FOXJ2 was a critical regulatory factor. Conclusions: This study identified the essential genes and pathways associated with exercise and monocytes. Among these, four essential genes (TNF, DUSP1, CXCR4, and NR4A1) and the FoxO signaling pathway play vital roles in the immune function of monocytes. High-intensity exercise may improve the resistance of chronic inflammatory diseases by regulating the expression of these genes.

Low-intensity pulsed ultrasound (LIPUS) as an adjuvant therapy in in vitro and in vivo bone engineering has proven to be extremely useful. The present study aimed at investigating the effect of 30 mW/cm(2) LIPUS stimulation on commercially available human mesenchymal stem cells (hMSCs) cultured in basal or osteogenic medium at different experimental time points (7d, 14d, 21d). The hypothesis was that LIPUS would improve the osteogenic differentiation of hMSC and guarantying the maintenance of osteogenic committed fraction, as demonstrated by cell vitality and proteomic analysis. LIPUS stimulation (a) regulated the balance between osteoblast commitment and differentiation by specific networks (activations of RhoA/ROCK signaling and upregulation of Ribosome constituent/Protein metabolic process, Glycolysis/Gluconeogenesis, RNA metabolic process/Splicing and Tubulins); (b) allowed the maintenance of a few percentage of osteoblast precursors (21d CD73 + /CD90 + : 6%; OCT-3/4 + /NANOG + /SOX2 + : 10%); (c) induced the activation of osteogenic specific pathways shown by gene expression (early: ALPL, COL1A1, late: RUNX2, BGLAP, MAPK1/6) and related protein release (COL1a1, OPN, OC), in particular in the presence of osteogenic soluble factors able to mimic bone microenvironment. To summarize, LIPUS might be able to improve the osteogenic commitment of hMSCs in vitro, and, at the same time, enhance their osteogenic differentiation. This article is protected by copyright. All rights reserved.

Nicotinamide adenine dinucleotide (NAD) is an essential molecule for living organisms and plays a vital role in aging and age-associated diseases. In eukaryotic cells, cellular NAD is mainly generated by the scavenge pathway in which nicotinamide phosphoribosyltransferase (NAMPT) catalyzes the formation of nicotinamide mononucleotide. Inhibition of NAMPT is a therapeutic strategy for cancer treatment. To explore the effects of NAMPT inhibition on cellular processes, cells were treated with 10 nM FK866, an NAMPT inhibitor, resulting in a decrease in the cellular NAD level, a lower growth rate, and enhanced susceptivity to oxidative stress as compared to the untreated cells. Quantitative proteomics revealed that 325 proteins were downregulated in the FK866-treated cells, and were involved in diverse cellular processes including nucleobase-containing compound metabolic process, protein metabolic process, antioxidant and DNA repair processes. Downregulation of 4 selected proteins was confirmed by western blotting and quantitative PCR. Downregulation of antioxidant proteins GRX1 and catalase, and DNA-repair proteins PCNA and PARP1 contributed to the enhanced susceptibility of FK866-treated cells to oxidative stress. FK866 treatment also caused mitochondrial dysfunction through downregulation of mitochondrial ribosomal proteins. Taken together, these results demonstrate that FK866 treatment efficiently decreases the cellular NAD level and induces autonomous changes in proteostasis, leading to cell growth inhibition and increased susceptibility to oxidative stress.

The endocannabinoid system (ECS) consists particularly of cannabinoid receptors 1 and 2 (CB1 and CB2), their endogenous ligands, and enzymes that synthesize and degrade their ligands. It acts in a variety of organs and disease states ranging from cancer progression over neuropathic pain to neurodegeneration. Protein components engaged in the signaling, trafficking, and homeostasis machinery of the G-protein coupled CB2, are however largely unknown. It is therefore important to identify further interaction partners to better understand CB2 receptor functions in physiology and pathophysiology. For this purpose, we used an affinity purification and mass spectrometry-based proteomics approach of Strep-HA-CB2 receptor in HEK293 cells. After subtraction of background interactions and protein frequency library assessment we could identify 83 proteins that were classified by the identification of minimally 2 unique peptides as highly probable interactors. A functional protein association network analysis obtained an interaction network with a significant enrichment of proteins functionally involved in protein metabolic process, in endoplasmic reticulum, response to stress but also in lipid metabolism and membrane organization. The network especially contains proteins involved in biosynthesis and trafficking like calnexin, Sec61A, tubulin chains TUBA1C and TUBB2B, TMED2, and TMED10. Six proteins that were only expressed in stable CB2 expressing cells were DHC24, DHRS7, GGT7, HECD3, KIAA2013, and PLS1. To exemplify the validity of our approach, we chose a candidate having a relatively low number of edges in the network to increase the likelihood of a direct protein interaction with CB2 and focused on the scaffold/phagosomal protein p62/SQSTM1. Indeed, we independently confirmed the interaction by co-immunoprecipitation and immunocytochemical colocalization studies. 3D reconstruction of confocal images furthermore showed CB2 localization in close proximity to p62 positive vesicles at the cell membrane. In summary, we provide a comprehensive repository of the CB2 interactome in HEK293 cells identified by a systematic unbiased approach, which can be used in future experiments to decipher the signaling and trafficking complex of this cannabinoid receptor. Future studies will have to analyze the exact mechanism of the p62-CB2 interaction as well as its putative role in disease pathophysiology.

Background and Aims Arbuscular mycorrhizal (AM) symbiosis begins with molecular signal communication (MSC) between AM fungi and the roots of the host plant. We aimed to test the hypothesis that the transcriptional profiles of wheat roots can be changed significantly by AM symbiotic signals, without direct contact. Methods Non-mycorrhizal (NM) and MSC treatments involved burying filter membrane bags containing sterilized and un-sterilized inoculum of the AM fungus Rhizophagus irregularis, respectively. The bags physically separated roots and AM structures but allowed molecular signals to pass through. Extracted RNA from wheat roots was sequenced by high-throughput sequencing. Results Shoot total nitrogen and phosphorus content of wheat plants was decreased by the MSC treatment. A total of 2360 differentially expressed genes (DEGs), including 1888 up-regulated DEGs and 472 down-regulated DEGs, were found dominantly distributed on chromosomes 2A, 2B, 2D, 3B, 5B and 5D. The expression of 59 and 121 genes was greatly up- and down-regulated, respectively. Only a portion of DEGs could be enriched into known terms during gene ontology analysis, and were mostly annotated to ‘catalytic activity’, ‘protein metabolic process’ and ‘membrane’ in the molecular function, biological process and cellular component ontology categories, respectively. More than 120 genes that may be involved in key processes during AM symbiosis development were regulated at the pre-physical contact stages. Conclusions The transcriptional profiles of wheat roots can be changed dramatically by MSC. Much of the information provided by our study is of great importance for understanding the mechanisms underlying the development of AM symbiosis.

Topological analysis of large networks, which focus on a specific biological process or on related biological processes, where functional coherence exists among the interacting members, may provide a wealth of insight into cellular functionality. This work presents an unbiased systems approach to analyze genetic, transcriptional regulatory and physical interaction networks of yeast genes possessing such functional coherence to gain novel biological insight. The present analysis identified only a few transcriptional regulators amongst a large gene cohort associated with the protein metabolism and processing in yeast. These transcription factors are not functionally required for the maintenance of these tasks in growing cells. Rather, they are involved in rewiring gene transcription in response to such major challenges as starvation, hypoxia, DNA damage, heat shock or the accumulation of unfolded proteins. Indeed, only a subset of these proteins were captured empirically in the nuclear-enriched fraction of non-stressed yeast cells, suggesting that the transcriptional regulation of protein metabolism and processing in yeast is primarily concerned with maintaining cellular robustness in the face of threat by either internal or external stressors.

Heat stress decreases milk yield and deleteriously alters milk composition. Reduced feed intake partially explains some of the consequences of heat stress, but metabolic changes in the mammary tissue and liver associated with milk synthesis have not been thoroughly evaluated. In the current study, changes of protein abundance in the mammary tissue and liver between heat-stressed cows with ad libitum intake and pair-fed thermal neutral cows were investigated using the iTRAQ proteomic approach. Most of the differentially expressed proteins from mammary tissue and liver between heat-stressed and pair-fed cows were involved in Gene Ontology category of protein metabolic process. Pathway analysis indicated that differentially expressed proteins in the mammary tissue were related to pyruvate, glyoxylate and dicarboxylate metabolism pathways, while those in the liver participated in oxidative phosphorylation and antigen processing and presentation pathways. Several heat shock proteins directly interact with each other and were considered as central "hubs" in the protein interaction network. These findings provide new insights to understand the turnover of protein biosynthesis pathways within hepatic and mammary tissue that likely contribute to changes in milk composition from heat-stressed cows.

Puccinia striiformis f. sp. tritici (Pst), the causal agent of wheat stripe rust, is an obligate pathogen that causes significant yield losses worldwide. In order to profile the change of the proteome during uredospore germination of Pst, proteins from uredospores and germtubes (germlings) were isolated and quantified by iTRAQ (isobaric tag for relative and absolute quantitation) technology. Among the 1548 proteins identified, there were 64 and 54 proteins up- and down-regulated during uredospore germination respectively. Proteins involved in catabolic process, energy production and transport were found to be specially accumulated in germlings, which was an indication of metabolic transition from dormancy to germination. While, proteins involved in protein metabolic process were largely down-regulated, suggesting fewer proteins were needed for uredospore germination. Additionally, a number of down-regulated proteins exhibited increasing mRNA level during germination, suggesting relative less correlation between proteins and mRNAs in dormant uredospores.

Utilization of heterosis is one of the most outstanding advancements in plant breeding. However, its genetic basis is not well understood. We used digital gene expression technology to investigate the transcriptomes of the mature maize embryo of an elite maize hybrid Zhengdan958 and its parental lines to screen differentially-expressed genes and to study their relationship to heterosis. In the study, we found five expression patterns, including co-silence expression of parental lines (754 genes), parental-specific expression (2,200 genes), hybrid-specific expression (704 genes), single-parental consistent expression (2,431 genes), and co-expression of the hybrid and parental lines (15,628 genes). Genes showing hybrid-specific expression pattern were enriched in photosynthesis related pathways. Those over-dominance expressed genes in co-expression pattern were striking enriched in protein metabolic process, and those under-dominance expressed genes in co-expression pattern were signally enriched in biosynthetic process. Those results indicated that mature embryo of Zhengdan958 has higher photosynthetic efficiency and better regulation ability in protein metabolic and biosynthetic process for better preparation for growth recover, and those enriched genes can be related with heterosis of maize mature embryos.

Increasing studies have shown that mature spermatozoa contain many transcripts including mRNAs and miRNAs. However, the expression profile of long non-coding RNAs (lncRNAs) in mammalian sperm has not been systematically investigated. Here, we used highly purified RNA to investigate lncRNA expression profiles in mouse mature sperm by stranded-specific RNA-seq. We identified 20,907 known and 4,088 novel lncRNAs transcripts, and the existence of intact lncRNAs was confirmed by RT-PCR and fluorescence in situ hybridization on two representative lncRNAs. Compared to round spermatids, 1,794 upregulated and 165 downregulated lncRNAs and 4,435 upregulated and 3,920 downregulated mRNAs were identified in sperm. Based on the "Cis and Trans" RNA-RNA interaction principle, we found 14,259 targeted coding genes of differently expressed lncRNAs. In terms of Gene ontology (GO) analysis, differentially expressed lncRNAs targeted genes mainly related to nucleic acid metabolic, protein modification, chromatin and histone modification, heterocycle compound metabolic, sperm function, spermatogenesis and other processes. In contrast, differentially expressed transcripts of mRNAs were highly enriched for protein metabolic process and RNA metabolic, spermatogenesis, sperm motility, cell cycle, chromatin organization, heterocycle and aromatic compound metabolic processes. Kyoto encyclopedia of genes and genomes (KEGG) pathway analysis showed that the differentially expressed lncRNAs were involved in RNA transport, mRNA surveillance pathway, PI3K-Akt signaling pathway, AMPK signaling pathway, protein processing in endoplasmic reticulum. Metabolic pathways, mRNA surveillance pathway, AMPK signaling pathway, cell cycle, RNA transport splicesome and endocytosis incorporated with the differentially expressed mRNA. Furthermore, many lncRNAs were specifically expressed in testis/sperm, and 880 lncRNAs were conserved between human and mouse. In summary, this study provides a preliminary database valuable for identifying lncRNAs critical in the late stage of spermatogenesis or important for sperm function regulation, fertilization and early embryo development.

The banana is one of the most important fruits in the world. Bananas undergo a rapid ripening process after harvest, resulting in a short shelf. In this study, the mechanism underlying pulp ripening of harvested bananas was investigated using integrated transcriptomic, proteomic, and metabolomic analysis. Ribonucleic acid sequencing (RNA-Seq) revealed that a great number of genes related to transcriptional regulation, signal transduction, cell wall modification, and secondary metabolism were up-regulated during pulp ripening. At the protein level, 84 proteins were differentially expressed during pulp ripening, most of which were associated with energy metabolism, oxidation-reduction, cell wall metabolism, and starch degradation. According to partial least squares discriminant analysis, 33 proteins were identified as potential markers for separating different ripening stages of the fruit. In addition to ethylene&rsquo, s central role, auxin signal transduction might be involved in regulating pulp ripening. Moreover, secondary metabolism, energy metabolism, and the protein metabolic process also played an important role in pulp ripening. In all, this study provided a better understanding of pulp ripening of harvested bananas.

Duchenne muscular dystrophy (DMD) is a lethal X-linked disorder caused by null mutations in the dystrophin gene. Although the primary defect is the deficiency of muscle dystrophin, secondary events, including chronic inflammation, fibrosis, and muscle regeneration failure are thought to actively contribute to disease progression. Despite several advances, there is still no effective therapy for DMD. Therefore, the potential regenerative capacities, and immune-privileged properties of mesenchymal stromal cells (MSCs), have been the focus of intense investigation in different animal models aiming the treatment of these disorders. However, these studies have shown different outcomes according to the sources from which MSCs were obtained, which raise the question whether stem cells from distinct sources have comparable clinical effects. Here, we analyzed the protein content of the secretome of MSCs, isolated from three different sources (adipose tissue, skeletal muscle, and uterine tubes), obtained from five donors and evaluated their in vitro properties when cocultured with DMD myoblasts. All MSC lineages showed pathways enrichment related to protein metabolic process, oxidation-reduction process, cell proliferation, and regulation of apoptosis. We found that MSCs secretome proteins and their effect in vitro vary significantly according to the tissue and donors, including opposite effects in apoptosis assay, indicating the importance of characterizing MSC secretome profile before its use in animal and clinical trials. Despite the individual differences a pool of conditioned media from all MSCs lineages was able to delay apoptosis and enhance migration when in contact with DMD myoblasts.

Background A combined supplement of vitamins B9 and B12 was reported to increase milk and milk component yields of dairy cows without effect on feed intake. The present study was undertaken to verify whether this supplementation positively modifies the pathways involved in milk and milk component synthesis. Thus, by studying the transcriptome activity in these tissues, the effect of supplements of both vitamins on the metabolism of both liver and mammary gland, was investigated. For this study, 24 multiparous Holstein dairy cows were assigned to 6 blocks of 4 animals each according to previous 305-day milk production. Within each block, cows were randomly assigned to weekly intramuscular injections of 5 mL of either saline 0.9 % NaCl, 320 mg of vitamin B9, 10 mg of vitamin B12 or a combination of both vitamins (B9 + B12). The experimental period began 3 weeks before the expected calving date and lasted 9 weeks of lactation. Liver and mammary biopsies were performed on lactating dairy cows 64 ± 3 days after calving. Samples from both tissues were analyzed by microarray and qPCR to identify genes differentially expressed in hepatic and mammary tissues. Results Microarray analysis identified 47 genes in hepatic tissue and 16 genes in the mammary gland whose expression was modified by the vitamin supplements. Gene ontology (GO) categorizes genes in non-overlapping domains of molecular biology. Panther is one of the online GO resources used for gene function classification. It classifies the 63 genes according to Molecular Function, Biological Process and Protein Class. Most of the biological processes modulated by the vitamin supplements were associated to developmental process, protein metabolic process, transport and response to inflammation. In the liver, most of the genes modulated by the vitamin treatments involved protein metabolic process while developmental process appeared to be more affected by the treatments in mammary gland. Out of 25 genes analysed by qPCR, 7 were validated. Conclusion The results indicate that several metabolic processes were modulated by the supplementation of vitamins in early-lactating dairy cows. In addition, the results suggest that the vitamin supplements promoted liver regeneration and reduced catabolism of lipids in early lactation. Electronic supplementary material The online version of this article (doi:10.1186/s12864-016-2872-2) contains supplementary material, which is available to authorized users.

Background The Sapsaree (Canis familiaris) is a Korean native dog that is very friendly, protective, and loyal to its owner, and is registered as a natural monument in Korea (number: 368). To investigate large-scale gene expression profiles and identify the genes related to exercise-induced stress in the Sapsaree, we performed whole-transcriptome RNA sequencing and analyzed gene expression patterns before and after exercise performance. Results We identified 525 differentially expressed genes in ten dogs before and after exercise. Gene Ontology classification and KEGG pathway analysis revealed that the genes were mainly involved in metabolic processes, such as programmed cell death, protein metabolic process, phosphatidylinositol signaling system, and cation binding in cytoplasm. The ten Sapsarees could be divided into two groups based on the gene expression patterns before and after exercise. The two groups were significantly different in terms of their basic body type (p ≤ 0.05). Seven representative genes with significantly different expression patterns before and after exercise between the two groups were chosen and characterized. Conclusions Body type had a significant effect on the patterns of differential gene expression induced by exercise. Whole-transcriptome sequencing is a useful method for investigating the biological characteristics of the Sapsaree and the large-scale genomic differences of canines in general. Electronic supplementary material The online version of this article (doi:10.1186/s40781-016-0097-1) contains supplementary material, which is available to authorized users.

Berberine is one of the main alkaloids found in the Chinese herb Huang lian (Rhizoma Coptidis), which has been reported to have multiple pharmacological activities. This study aimed to analyze the molecular targets of berberine based on literature data followed by a pathway analysis using the PANTHER program. PANTHER analysis of berberine targets showed that the most classes of molecular functions include receptor binding, kinase activity, protein binding, transcription activity, DNA binding, and kinase regulator activity. Based on the biological process classification ofin vitroberberine targets, those targets related to signal transduction, intracellular signalling cascade, cell surface receptor-linked signal transduction, cell motion, cell cycle control, immunity system process, and protein metabolic process are most frequently involved. In addition, berberine was found to interact with a mixture of biological pathways, such as Alzheimer’s disease-presenilin and -secretase pathways, angiogenesis, apoptosis signalling pathway, FAS signalling pathway, Hungtington disease, inflammation mediated by chemokine and cytokine signalling pathways, interleukin signalling pathway, and p53 pathways. We also explored the possible mechanism of action for the anti-diabetic effect of berberine. Further studies are warranted to elucidate the mechanisms of action of berberine using systems biology approach.

Proteomic analysis of body fluids can be used to identify proteins associated with malignant transformation, and tumor fluid samples have emerged as a rich source for the identification of ovarian cancer in this context. To uncover differences among benign and malignant ovarian samples, we performed a high-throughput proteomic analysis consisting of albumin immunodepletion, isotope labeling with acrylamide, and in-depth proteomic profiling by LC-MS/MS in a pool of 10 samples of each histologic type (high-grade ovarian serous carcinoma and benign serous cystadenoma). After, we developed a panel for targeted proteomics using the multiple reaction monitoring (MRM) method for validation of 10 relevant proteins (ACTN4 (Alpha-actinin-4), GSTP1 (Glutathione S-transferase P), LDHA (L-lactate dehydrogenase A chain), LDHB (L-lactate dehydrogenase B chain), PRDX5 (Peroxiredoxin-5), TIMP1 (Metalloproteinase inhibitor 1), ACTG1 (Actin), ALDOA (Fructose-bisphosphate aldolase A), PKM (Pyruvate kinase PKM), and ENO1 (Alpha-enolase)). A total of 1135 proteins were identified in the high-throughput proteomic analysis, corresponding to 593 known genes, where 505 proteins (44%) were detected only in the malignant pool and 272 proteins (24%) were detected only in the benign pool. The protein-protein interaction analysis (STRING-DB) showed strong association of proteins detected in the malignant pool, according to biological process and cellular component, with response to stress, protein metabolic process, immune system process, positive regulation of cellular metabolic process, biogenesis, regulation of cellular organization, transport, protein trafficking, extracellular exosome, extracellular region part, and extracellular region. Differently, the set of proteins detected in the benign pool showed strong association with single organism signaling, cell communication, regulation of signal transduction, system development, response to stimulus, extracellular region part, extracellular exosome, cell periphery, cytosol, cell projection, endomembrane system, and cytoskeleton. Using targeted proteomic analysis, we verified that ACTN4, GSTP1, LDHB, PRDX5, TIMP1, ACTG1, ALDOA, LDHA, PKM, and ENO1 are significantly more abundant in malignant tumor fluid pool when compared to benign samples, confirming the association of proteins detected in the malignant pool with metabolic process, extracellular exosome, and extracellular region part. These results are in accordance with the fact that cancer cells must adjust cellular metabolism to support the demands of growth and remodel their cytoskeleton to allow migration and metastasis. Our findings contribute to show the differences among histologic types of ovarian tumor fluids. We emphasize the contribution of robust proteomic strategies to uncover relevant candidate tumor markers and consistent molecular signatures that can discriminate between benign and malignant tumors. Financial support: FAPESP (Proc. 2011-0947-1, 16/03809-3), CNPq (479934/2011-8, 454703/2014-7), CAPES, CISBi, NAP/USP (grant 12.1.17598.1.3) and Center for Cell Based Therapy - CTC-CEPID (Proc. FAPESP 2013/08135-2). Citation Format: Aline Poersch, Mariana L. Grassi, Camila S. Palma, Vinícius P. Carvalho, Guilherme P. Lanfredi, Helio H. A. Carrara, Francisco J. Candido dos Reis, Vitor M. Faça. Profile of ovarian cancer tumor fluid using high-throughput and targeted proteomic analysis [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr A191.

Dermal papillae (DPs) control the formation of hair shafts. In clinical settings, colchicine (CLC) induces patients’ hair shedding. Compared to the control, the ex vivo hair fiber elongation of organ cultured vibrissa hair follicles (HFs) declined significantly after seven days of CLC treatment. The cultured DP cells (DPCs) were used as the experimental model to study the influence of CLC on the protein dynamics of DPs. CLC could alter the morphology and down-regulate the expression of alkaline phosphatase (ALP), the marker of DPC activity, and induce IκBα phosphorylation of DPCs. The proteomic results showed that CLC modulated the expression patterns (fold &gt, 2) of 24 identified proteins, seven down-regulated and 17 up-regulated. Most of these proteins were presumably associated with protein turnover, metabolism, structure and signal transduction. Protein-protein interactions (PPI) among these proteins, established by Search Tool for the Retrieval of Interacting Genes/Proteins (STRING) database, revealed that they participate in protein metabolic process, translation, and energy production. Furthermore, ubiquitin C (UbC) was predicted to be the controlling hub, suggesting the involvement of ubiquitin-proteasome system in modulating the pathogenic effect of CLC on DPC.

This study investigated global gene expression in the small yellow follicles (6-8 mm diameter) of broiler-type B strain Taiwan country chickens (TCCs) in response to acute heat stress. Twelve 30-wk-old TCC hens were divided into four groups: control hens maintained at 25°C and hens subjected to 38°C acute heat stress for 2 h without recovery (H2R0), with 2-h recovery (H2R2), and with 6-h recovery (H2R6). Small yellow follicles were collected for RNA isolation and microarray analysis at the end of each time point. Results showed that 69, 51, and 76 genes were upregulated and 58, 15, 56 genes were downregulated after heat treatment of H2R0, H2R2, and H2R6, respectively, using a cutoff value of two-fold or higher. Gene ontology analysis revealed that these differentially expressed genes are associated with the biological processes of cell communication, developmental process, protein metabolic process, immune system process, and response to stimuli. Upregulation of heat shock protein 25, interleukin 6, metallopeptidase 1, and metalloproteinase 13, and downregulation of type II alpha 1 collagen, discoidin domain receptor tyrosine kinase 2, and Kruppel-like factor 2 suggested that acute heat stress induces proteolytic disintegration of the structural matrix and inflamed damage and adaptive responses of gene expression in the follicle cells. These suggestions were validated through gene expression, using quantitative real-time polymerase chain reaction. Functional annotation clarified that interleukin 6-related pathways play a critical role in regulating acute heat stress responses in the small yellow follicles of TCC hens.

Garlic is widely used as a spice throughout the world. In this study, transcriptional profilings of garlic shoot apex were performed by using the Illumina technology. A total of 45,363 significantly changed expressed transcripts were detected between the dormant and sprouting garlic shoot apex libraries. The expression of 22,836 unigenes was increased by more than 2-fold in sprouting garlic shoot apex as compared with dormant shoot apex (up-regulated unigene), and 22,526 unigenes were identified as having been down-regulated. Gene ontology (GO) annotations indicated that the differentially expressed genes were mainly played role in nucleotide binding, plastid, hydrolase activity, transferase activity, protein metabolic process, nucleic acid binding, protein binding, mitochondrion. A total of 8,725 differentially expressed genes were assigned to five Kyoto Encyclopedia of Genes and Genomes (KEGG) biochemical pathways, including metabolism, genetic information processing, organism system, cellular processes, and environmental information processing. Real-time quantitative RT-PCR (qRT-PCR) was performed to dissect shoot apex sprouting. The differential expression of genes, such as ENHYDROUS, DAG1, DAM, DTH8, indicate that they play a critical role in shoot apex sprouting. These differentially expressed genes comprise related candidates for shoot apex sprouting regulation in Allium species.

Plants are endowed with a sophisticated defense mechanism that gives signals to plant cells about the immediate danger from surroundings and protects them from pathogen invasion. In the search for the particular proteins involved in fruit defense responses, we report here a comparative analysis of tomato fruit (Solanum lycopersicum cv. Ailsa Craig) infected by Rhizopus nigricans Ehrenb, which is a significant contributor to postharvest rot disease in fresh tomato fruits. In total, four hundred forty-five tomato proteins were detected in common between the non-infected group and infected tomato fruit of mature green. Forty-nine differentially expressed spots in 2-D gels were identified, and were sorted into fifteen functional groups. Most of these proteins participate directly in the stress response process, while others were found to be involved in several equally important biological processes: protein metabolic process, carbohydrate metabolic process, ethylene biosynthesis, and cell death and so on. These responses occur in different cellular components, both intra- and extracellular spaces. The differentially expressed proteins were integrated into several pathways to show the regulation style existing in tomato fruit host. The composition of the collected proteins populations and the putative functions of the identified proteins argue for their roles in pathogen-plant interactions. Collectively results provide evidence that several regulatory pathways contribute to the resistance of tomato fruit to pathogen.

PURPOSE To investigate the endothelial gene expression profile in a Col8a2 Q455K mutant knock-in mouse model of early-onset Fuchs' endothelial corneal dystrophy (FECD) and identify potential targets that can be correlated to human late-onset FECD. METHODS Diseased or normal endothelial phenotypes were verified in 12-month-old homozygous Col8a2(Q455K/Q455K) mutant and wild-type mice by clinical confocal microscopy. An endothelial whole genome expression profile was generated by microarray-based analysis. Result validation was performed by real-time PCR. Endothelial COX2 and JUN expression was further studied in human late-onset FECD compared to normal samples. RESULTS Microarray analysis demonstrated endothelial expression of 24,538 genes (162 up-regulated and 172 down-regulated targets) and identified affected gene ontology terms including Response to Stress, Protein Metabolic Process, Protein Folding, Regulation of Apoptosis, and Transporter Activity. Real-time PCR assessment confirmed increased Cox2 (P = 0.001) and Jun mRNA (P = 0.03) levels in Col8a2(Q455K/Q455K) mutant compared to wild-type mice. In human FECD samples, real-time PCR demonstrated a statistically significant increase in COX2 mRNA (P < 0.0001) and JUN mRNA (P = 0.002) and tissue microarray analysis showed increased endothelial COX2 (P = 0.02) and JUN protein (P = 0.04). CONCLUSIONS The present study provides the first endothelial whole genome expression analysis in an animal model of FECD and represents a useful resource for future studies of the disease. In particular endothelial COX2 up-regulation warrants further investigation of its role in FECD.

Climate change substantially affects biodiversity around the world, especially in the Amazon region, which is home to a significant portion of the world's biodiversity. Freshwater fishes are susceptible to increases in water temperature and variations in the concentrations of dissolved gases, especially oxygen and carbon dioxide. It is important to understand the mechanisms underlying the physiological and biochemical abilities of fishes to survive such environmental changes. In the present study, we applied RNA-Seq and de novo transcriptome sequencing to evaluate transcriptome alterations in tambaqui when exposed to five or fifteen days of the B1, A1B and A2 climate scenarios foreseen by the IPCC. The generated ESTs were assembled into 54,206 contigs. Gene ontology analysis and the STRING tool were then used to identify candidate protein domains, genes and gene families potentially responsible for the adaptation of tambaqui to climate changes. After sequencing eight RNA-Seq libraries, 32,512 genes were identified and mapped using the Danio rerio genome as a reference. In total, 236 and 209 genes were differentially expressed at five and fifteen days, respectively, including chaperones, energetic metabolism-related genes, translation initiation factors and ribosomal genes. Gene ontology enrichment analysis revealed that mitochondrion, protein binding, protein metabolic process, metabolic processes, gene expression, structural constituent of ribosome and translation were the most represented terms. In addition, 1,202 simple sequence repeats were detected, 88 of which qualified for primer design. These results show that cellular response to climate change in tambaqui is complex, involving many genes, and it may be controlled by different cues and transcription/translation regulation mechanisms. The data generated from this study provide a valuable resource for further studies on the molecular mechanisms involved in the adaptation of tambaqui and other closely related teleost species to climate change.

Parthenogenetic embryos are one attractive alternative as a source of embryonic stem cells, although many aspects related to the biology of parthenogenetic embryos and parthenogenetically derived cell lines still need to be elucidated. The present work was conducted to investigate the gene expression profile of rabbit parthenote embryos cultured under in vivo conditions using microarray analysis. Transcriptomic profiles indicate 2541 differentially expressed genes between parthenotes and normal in vivo fertilised blastocysts, of which 76 genes were upregulated and 16 genes downregulated in in vivo cultured parthenote blastocyst, using 3 fold-changes as a cut-off. While differentially upregulated expressed genes are related to transport and protein metabolic process, downregulated expressed genes are related to DNA and RNA binding. Using microarray data, 6 imprinted genes were identified as conserved among rabbits, humans and mice: GRB10, ATP10A, ZNF215, NDN, IMPACT and SFMBT2. We also found that 26 putative genes have at least one member of that gene family imprinted in other species. These data strengthen the view that a large fraction of genes is differentially expressed between parthenogenetic and normal embryos cultured under the same conditions and offer a new approach to the identification of imprinted genes in rabbit. © 2012 Naturil-Alfonso et al., This work was supported by Generalitat Valenciana research programme (Prometeo 2009/125). Carmen Naturil was supported by Generalitat Valenciana research programme (Prometeo 2009/125). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Major depression might originate from both environmental and genetic risk factors. The environmental chronic mild stress (CMS) model mimics some environmental factors contributing to human depression and induces anhedonia and helplessness. Mice heterozygous for the synaptic vesicle protein (SVP) vesicular glutamate transporter 1 (VGLUT1) have been proposed as a genetic model of deficient glutamate function linked to depressive-like behaviour. Here, we aimed to identify, in these two experimental models, gene expression changes in the frontal cortex, common to stress and impaired glutamate function. Both VGLUT1+/- and CMS mice showed helpless and anhedonic-like behavior. Microarray studies in VGLUT1+/- mice revealed regulation of genes involved in apoptosis, neurogenesis, synaptic transmission, protein metabolic process or learning and memory. In addition, RT-PCR studies confirmed gene expression changes in several glutamate, GABA, dopamine and serotonin neurotransmitter receptors. On the other hand, CMS affected the regulation of 147 transcripts, some of them involved in response to stress and oxidoreductase activity. Interestingly, 52 genes were similarly regulated in both models. Specifically, a dowregulation in genes that promote cell proliferation (Anapc7), cell growth (CsnK1g1), cell survival (Hdac3), inhibition of apoptosis (Dido1) was observed. Genes linked to cytoskeleton (Hspg2, Invs), psychiatric disorders (Grin1, MapK12) or an antioxidant enzyme (Gpx2) were also downregulated. Moreover, genes that inhibit the MAPK pathways (Dusp14), stimulate oxidative metabolism (Eif4a2) and enhance glutamate transmission (Rab8b) were upregulated. We suggest that these genes could form part of the altered “molecular context” underlying depressive-like behaviour in animal models. The clinical relevance of these findings is discussed.