The Chikungunya virus (CHIKV) is an arthropod borne virus. In the last 50 years, it has been the cause of numerous outbreaks in tropical and temperate regions, worldwide. There is limited understanding regarding the underlying molecular mechanisms involved in CHIKV replication and how the virus interacts with its host. In the present study, comparative proteomics was used to identify secreted host proteins that changed in abundance in response to early CHIKV infection. Two-dimensional gel electrophoresis was used to analyse and compare the secretome profiles of WRL-68 cells infected with CHIKV against mock control WRL-68 cells. The analysis identified 25 regulated proteins in CHIKV infected cells. STRING network analysis was then used to predict biological processes that may be affected by these proteins. The processes predicted to be affected include signal transduction, cellular component and extracellular matrix (ECM) organization, regulation of cytokine stimulus and immune response. These results provide an initial view of CHIKV may affect the secretome of infected cells during early infection. The results presented here will compliment earlier results from the study of late host response. However, functional characterization will be necessary to further enhance our understanding of the roles played by these proteins in the early stages of CHIKV infection in humans.
Haloxylon persicum is an endangered western Asiatic desert plant species, which survives under extreme environmental conditions. In this study, we focused on transcriptome analysis of H. persicum to understand the molecular mechanisms associated with drought tolerance. Two different periods of polyethylene glycol (PEG)-induced drought stress (48 h and 72 h) were imposed on H. persicum under in vitro conditions, which resulted in 18 million reads, subsequently assembled by de novo method with more than 8000 transcripts in each treatment. The N50 values were 1437, 1467, and 1524 for the control sample, 48 h samples, and 72 h samples, respectively. The gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) pathway analysis resulted in enrichment of mitogen-activated protein kinase (MAPK) and plant hormone signal transduction pathways under PEG-induced drought conditions. The differential gene expression analysis (DGEs) revealed significant changes in the expression pattern between the control and the treated samples. The KEGG analysis resulted in mapping transcripts with 138 different pathways reported in plants. The differential expression of drought-responsive transcription factors depicts the possible signaling cascades involved in drought tolerance. The present study provides greater insight into the fundamental transcriptome reprogramming of desert plants under drought.
The hypoxia-inducible factors (HIFs) are key transcription factors in determining cellular responses involving alterations in protein levels in response to limited oxygen availability in animal cells. 2-Oxoglutarate-dependent oxygenases play key roles in regulating levels of HIF and its transcriptional activity. We describe MS-based proteomics studies in which we compared the results of subjecting human breast cancer MCF-7 cells to hypoxia or treating them with a cell-penetrating derivative (dimethyl N-oxalylglycine; DMOG) of the stable 2OG analogue N-oxalylglycine. The proteomic results are consistent with reported transcriptomic analyses and support the proposed key roles of 2OG-dependent HIF prolyl- and asparaginyl-hydroxylases in the hypoxic response. Differences between the data sets for hypoxia and DMOG might reflect context-dependent effects or HIF-independent effects of DMOG.
Adequate genetic information is essential for sustainable crustacean fisheries and aquaculture management. The commercially important orange mud crab, Scylla olivacea, is prevalent in Southeast Asia region and is highly sought after. Although it is a suitable aquaculture candidate, full domestication of this species is hampered by the lack of knowledge about the sexual maturation process and the molecular mechanisms behind it, especially in males. To date, data on its whole genome is yet to be reported for S. olivacea. The available transcriptome data published previously on this species focus primarily on females and the role of central nervous system in reproductive development. De novo transcriptome sequencing for the testes of S. olivacea from immature, maturing and mature stages were performed. A total of approximately 144 million high-quality reads were generated and de novo assembled into 160,569 transcripts with a total length of 142.2 Mb. Approximately 15-23% of the total assembled transcripts were annotated when compared to public protein sequence databases (i.e. UniProt database, Interpro database, Pfam database and Drosophila melanogaster protein database), and GO-categorised with GO Ontology terms. A total of 156,181 high-quality Single-Nucleotide Polymorphisms (SNPs) were mined from the transcriptome data of present study. Transcriptome comparison among the testes of different maturation stages revealed one gene (beta crystallin like gene) with the most significant differential expression-up-regulated in immature stage and down-regulated in maturing and mature stages. This was further validated by qRT-PCR. In conclusion, a comprehensive transcriptome of the testis of orange mud crabs from different maturation stages were obtained. This report provides an invaluable resource for enhancing our understanding of this species' genome structure and biology, as expressed and controlled by their gonads.
The natural rubber latex extracted from the bark of Hevea brasiliensis plays various important roles in today's modern society. Following ultracentrifugation, the latex can be separated into 3 layers: C-serum, lutoids, and rubber particles. Previous studies have shown that a large number of proteins are present in these 3 layers. However, a complete proteome for this important plant is still unavailable. Protein sequences have been recently translated from the completed draft genome database of H. brasiliensis, leading to the creation of annotated protein databases of the following H. brasiliensis biosynthetic pathways: photosynthesis, latex allergens, rubberwood formation, latex biosynthesis, and disease resistance. This research was conducted to identify the proteins contained within the latex by way of de novo sequencing from mass spectral data obtained from the 3 layers of the latex. Peptides from these proteins were fragmented using collision-induced dissociation, higher-energy collisional dissociation, and electron-transfer dissociation activation methods. A large percentage of proteins from the biosynthetic pathways (63% to 100%) were successfully identified. In addition, a total of 1839 unique proteins were identified from the whole translated draft genome database (AnnHBM).
Chemo-resistance is associated with poor prognosis in colorectal cancer (CRC), with the absence of early biomarker. Exosomes are microvesicles released by body cells for intercellular communication. Circular RNAs (circRNAs) are non-coding RNAs with covalently closed loops and enriched in exosomes. Crosstalk between circRNAs in exosomes and chemo-resistance in CRC remains unknown. This research aims to identify exosomal circRNAs associated with FOLFOX-resistance in CRC. FOLFOX-resistant HCT116 CRC cells (HCT116-R) were generated from parental HCT116 cells (HCT116-P) using periodic drug induction. Exosomes were characterized using transmission electron microscopy (TEM), Zetasizer and Western blot. Our exosomes were translucent cup-shaped structures under TEM with differential expression of TSG101, CD9, and CD63. We performed circRNAs microarray using exosomal RNAs from HCT116-R and HCT116-P cells. We validated our microarray data using serum samples. We performed drug sensitivity assay and cell cycle analysis to characterize selected circRNA after siRNA-knockdown. Using fold change >2 and p
Salmonella typhimurium is an important biofilm-forming bacteria. It is known to be resistant to a wide range of antimicrobials. The present study was carried out to evaluate the effects of dimethyl sulfoxide (DMSO) against S. typhimurium biofilm and investigate whole-cell protein expression by biofilm cells following treatment with DMSO. Antibiofilm activities were assessed using pellicle assay, crystal violet assay, colony-forming unit counting and extracellular polymeric substance (EPS) matrix assay whilst differential protein expression was investigated using a combination of one dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis, tandem mass spectrometry and bioinformatics. Treatment with 32% DMSO inhibited pellicle formation, biofilm viability, biofilm biomass and several important components of EPS matrix. Subtractive protein profiling identified two unique protein bands (25.4 and 51.2 kDa) which were present only in control biofilm and not in 32% DMSO-treated biofilm. In turn, 29 and 46 proteins were successfully identified from the protein bands of 25.4 and 51.2 kDa respectively. Protein interaction network analysis identified several biological pathways to be affected, including glycolysis, PhoP-PhoQ phosphorelay signalling and flagellar biosynthesis. The present study suggests that DMSO may inhibit multiple biological pathways to control biofilm formation.
Rhodococcus jostii RHA1 (RHA1) degrades polychlorinated biphenyl (PCB) via co-metabolism with biphenyl. To identify the novel open reading frames (ORFs) that contribute to PCB/biphenyl metabolism in RHA1, we compared chromatin immunoprecipitation chip and transcriptomic data. Six novel ORFs involved in PCB/biphenyl metabolism were identified. Gene deletion mutants of these 6 ORFs were made and were tested for their ability to grow on biphenyl. Interestingly, only the ro10225 deletion mutant showed deficient growth on biphenyl. Analysis of Ro10225 protein function showed that growth of the ro10225 deletion mutant on biphenyl was recovered when exogenous recombinant Ro10225 protein was added to the culture medium. Although Ro10225 protein has no putative secretion signal sequence, partially degraded Ro10225 protein was detected in conditioned medium from wild-type RHA1 grown on biphenyl. This Ro10225 fragment appeared to form a complex with another PCB/biphenyl oxidation enzyme. These results indicated that Ro10225 protein is essential for the formation of the PCB/biphenyl dioxygenase complex in RHA1.
The natural rubber of Para rubber tree, Hevea brasiliensis, is the main crop involved in industrial rubber production due to its superior quality. The Hevea bark is commercially exploited to obtain latex, which is produced from the articulated secondary laticifer. The laticifer is well defined in the aspect of morphology; however, only some genes associated with its development have been reported. We successfully induced secondary laticifer in the jasmonic acid (JA)-treated and linolenic acid (LA)-treated Hevea bark but secondary laticifer is not observed in the ethephon (ET)-treated and untreated Hevea bark. In this study, we analysed 27,195 gene models using NimbleGen microarrays based on the Hevea draft genome. 491 filtered differentially expressed (FDE) transcripts that are common to both JA- and LA-treated bark samples but not ET-treated bark samples were identified. In the Eukaryotic Orthologous Group (KOG) analysis, 491 FDE transcripts belong to different functional categories that reflect the diverse processes and pathways involved in laticifer differentiation. In the Kyoto Encyclopedia of Genes and Genomes (KEGG) and KOG analysis, the profile of the FDE transcripts suggest that JA- and LA-treated bark samples have a sufficient molecular basis for secondary laticifer differentiation, especially regarding secondary metabolites metabolism. FDE genes in this category are from the cytochrome (CYP) P450 family, ATP-binding cassette (ABC) transporter family, short-chain dehydrogenase/reductase (SDR) family, or cinnamyl alcohol dehydrogenase (CAD) family. The data includes many genes involved in cell division, cell wall synthesis, and cell differentiation. The most abundant transcript in FDE list was SDR65C, reflecting its importance in laticifer differentiation. Using the Basic Local Alignment Search Tool (BLAST) as part of annotation and functional prediction, several characterised as well as uncharacterized transcription factors and genes were found in the dataset. Hence, the further characterization of these genes is necessary to unveil their role in laticifer differentiation. This study provides a platform for the further characterization and identification of the key genes involved in secondary laticifer differentiation.
Burkholderia pseudomallei, the etiological agent for melioidosis, is known to secrete a type III secretion system (TTSS) protein into the host's internal milieu. One of the TTSS effector protein, BipC, has been shown to play an important role in the B. pseudomallei pathogenesis. To identify the host response profile that was directly or indirectly regulated by this protein, genome-wide transcriptome approach was used to examine the gene expression profiles of infected mice. The transcriptome analysis of the liver and spleen revealed that a total of approximately 1,000 genes were transcriptionally affected by BipC. Genes involved in bacterial invasion, regulation of actin cytoskeleton, and MAPK signalling pathway were over-expressed and may be specifically regulated by BipC in vivo. These results suggest that BipC mainly targets pathways related to the cellular processes which could modulate the cellular trafficking processes. The host transcriptional response exhibited remarkable differences with and without the presence of the BipC protein. Overall, the detailed picture of this study provides new insights that BipC may have evolved to efficiently manipulate host-cell pathways which is crucial in the intracellular lifecycle of B. pseudomallei.
Entamoeba histolytica is a protozoan parasite that causes amebiasis and poses a significant health risk for populations in endemic areas. The molecular mechanisms involved in the pathogenesis and regulation of the parasite are not well characterized. We aimed to identify and quantify the differentially abundant membrane proteins by comparing the membrane proteins of virulent and avirulent variants of E. histolytica HM-1:IMSS, and to investigate the potential associations among the differentially abundant membrane proteins. We performed quantitative proteomics analysis using isobaric tags for relative and absolute quantitation labeling, in combination with two mass spectrometry instruments, that is, nano-liquid chromatography (nanoLC)-matrix-assisted laser desorption/ionization-mass spectrometry/mass spectrometry and nanoLC-electrospray ionization tandem mass spectrometry. Overall, 37 membrane proteins were found to be differentially abundant, whereby 19 and 18 membrane proteins of the virulent variant of E. histolytica increased and decreased in abundance, respectively. Proteins that were differentially abundant include Rho family GTPase, calreticulin, a 70-kDa heat shock protein, and hypothetical proteins. Analysis by Protein ANalysis THrough Evolutionary Relationships database revealed that the differentially abundant membrane proteins were mainly involved in catalytic activities (29.7%) and metabolic processes (32.4%). Differentially abundant membrane proteins that were found to be involved mainly in the catalytic activities and the metabolic processes were highlighted together with their putative roles in relation to the virulence. Further investigations should be performed to elucidate the roles of these proteins in E. histolytica pathogenesis.
Infectious bursal disease is a highly contagious disease in the poultry industry and causes immunosuppression in chickens. Genome-wide regulations of immune response genes of inbred chickens with different genetic backgrounds, following very virulent infectious bursal disease virus (vvIBDV) infection are poorly characterized. Therefore, this study aims to analyse the bursal tissue transcriptome of six inbred chicken lines 6, 7, 15, N, O and P following infection with vvIBDV strain UK661 using strand-specific next-generation sequencing, by highlighting important genes and pathways involved in the infected chicken during peak infection at 3 days post-infection. All infected chickens succumbed to the infection without major variations among the different lines. However, based on the viral loads and bursal lesion scoring, lines P and 6 can be considered as the most susceptible lines, while lines 15 and N were regarded as the least affected lines. Transcriptome profiling of the bursa identified 4588 genes to be differentially expressed, with 2985 upregulated and 1642 downregulated genes, in which these genes were commonly or uniquely detected in all or several infected lines. Genes that were upregulated are primarily pro-inflammatory cytokines, chemokines and IFN-related. Various genes that are associated with B-cell functions and genes related to apoptosis were downregulated, together with the genes involved in p53 signalling. In conclusion, bursal transcriptome profiles of different inbred lines showed differential expressions of pro-inflammatory cytokines and chemokines, Th1 cytokines, JAK-STAT signalling genes, MAPK signalling genes, and their related pathways following vvIBDV infection.