De novo transcriptome sequencing, assembly, and gene expression profiling of a salt-stressed halophyte (Salsola drummondii) from a saline habitat.

Salsola drummondii is a perennial habitat-indifferent halophyte growing in saline and nonsaline habitats of the Arabian hyperarid deserts. It offers an invaluable opportunity to examine the molecular mechanisms of salt tolerance. The present study was conducted to elucidate these mechanisms through transcriptome profiling of seedlings grown from seeds collected in a saline habitat. The Illumina Hiseq 2500 platform was employed to sequence cDNA libraries prepared from shoots and roots of nonsaline-treated plants (controls) and plants treated with 1200 mM NaCl. Transcriptomic comparison between salt-treated and control samples resulted in 17,363 differentially expressed genes (DEGs), including 12,000 upregulated genes (7870 in roots, 4130 in shoots) and 5363 downregulated genes (4258 in roots and 1105 in shoots). The majority of identified DEGs are known to be involved in transcription regulation (79), signal transduction (82), defense metabolism (101), transportation (410), cell wall metabolism (27), regulatory processes (392), respiration (85), chaperoning (9), and ubiquitination (98) during salt tolerance. This study identified potential genes associated with the salt tolerance of S. drummondii and demonstrated that this tolerance may depend on the induction of certain genes in shoot and root tissues. These gene expressions were validated using reverse-transcription quantitative PCR, the results of which were consistent with transcriptomics results. To the best of our knowledge, this is the first study providing genetic information on salt tolerance mechanisms in S. drummondii.

Phytotoxic and Genotoxic Effects of Copper Nanoparticles in Coriander (Coriandrum sativum-Apiaceae).

Engineered metal nanoparticles have been widely used in several applications that may lead to increased exposure to the environment. In this study, we assessed the phytotoxic effect of various concentrations of copper nanoparticles CuNP, (200, 400 and 800 mg/L) on coriander (Coriandrum sativum) plants grown hydroponically. C. sativum plants treated with CuNP demonstrated decreased biomass and root length in comparison to control untreated plants. Additionally, decreased levels of photosynthetic pigments (chlorophyll a and b) were also seen in C. sativum plants treated with CuNP, as well as damage to the C. sativum root plasma membrane as demonstrated by Evan's blue dye and increased electrolyte leakage. Moreover, our results exhibited increased levels of H2O2 and MDA on C. Sativum plants treated with CuNP. X-Ray Fluorescence (XRF) analysis confirmed that C. sativum treated with CuNP accumulated the latter in plant root tissues. Random amplified polymorphic DNA (RAPD) analysis confirmed the genotoxic effect of CuNP, which altered the C. sativum genome. This was shown by the different banding pattern of RAPD. Overall, our results exhibited that CuNP is toxic to C. sativum plants.

Copper Nanoparticles Induced Genotoxicty, Oxidative Stress, and Changes in Superoxide Dismutase (SOD) Gene Expression in Cucumber (Cucumis sativus) Plants.

With the increased use of metal nanoparticles (NPs), their access to the food chain has become a main concern to scientists and holds controversial social implications. This research particularly sheds light on copper nanoparticles (CuNP), as they have been commonly used in several industries nowadays. In this study, we investigated the phytotoxicity of CuNP on cucumber (Cucumis sativus) plants grown hydroponically. Atomic Absorption Spectroscopy (AAS), X-Ray Fluorescence (XRF), and Scanning Electron Microscopy (SEM) analysis confirmed that C. sativus treated with CuNP accumulated CuNP in the plant tissues, with higher levels in roots, with amounts that were concentration dependent. Furthermore, genotoxicity was assessed using Random amplified polymorphic DNA (RAPD) technique, and our results showed that CuNP caused genomic alterations in C. sativus. Phenotypical, physiological, and biochemical changes were assessed by determining the CuNP treated plant's total biomass, chlorophyll, H2O2 and MDA contents, and electrolyte leakage percentage. The results revealed notable adverse phenotypical changes along with decreased biomass and decreased levels of the photosynthetic pigments (Chlorophyll a and b) in a concentration-dependent manner. Moreover, CuNP induced damage to the root plasma membrane as determined by the increased electrolyte leakage. A significant increase in H2O2 and MDA contents were detected in C. sativus CuNP treated plants. Additionally, copper-zinc superoxide dismutase (Cu-Zn SOD) gene expression was induced under CuNP treatment. Overall, our results demonstrated that CuNP of 10-30 nm size were toxic to C. sativus plants. This finding will encourage the safe production and disposal NPs. Thus, reducing nano-metallic bioaccumulation into our food chain through crop plants; that possesses a threat to the ecological system.

Analysis of human blood plasma proteome from ten healthy volunteers from Indian population.

Analysis of any mammalian plasma proteome is a challenge, particularly by mass spectrometry, due to the presence of albumin and other abundant proteins which can mask the detection of low abundant proteins. As detection of human plasma proteins is valuable in diagnostics, exploring various workflows with minimal fractionation prior to mass spectral analysis, is required in order to study population diversity involving analysis in a large cohort of samples. Here, we used 'reference plasma sample', a pool of plasma from 10 healthy individuals from Indian population in the age group of 25-60 yrs including 5 males and 5 females. The 14 abundant proteins were immunodepleted from plasma and then evaluated by three different workflows for proteome analysis using a nanoflow reverse phase liquid chromatography system coupled to a LTQ Orbitrap Velos mass spectrometer. The analysis of reference plasma sample a) without prefractionation, b) after prefractionation at peptide level by strong cation exchange chromatography and c) after prefractionation at protein level by sodium dodecyl sulfate polyacrylamide gel electrophoresis, led to the identification of 194, 251 and 342 proteins respectively. Together, a comprehensive dataset of 517 unique proteins was achieved from all the three workflows, including 271 proteins with high confidence identified by ≥ 2 unique peptides in any of the workflows or identified by single peptide in any of the two workflows. A total of 70 proteins were common in all the three workflows. Some of the proteins were unique to our study and could be specific to Indian population. The high-confidence dataset obtained from our study may be useful for studying the population diversity, in discovery and validation process for biomarker identification.

Secretome analysis of Glioblastoma cell line--HNGC-2.

Glioblastoma multiforme (GBM) is the most common and aggressive type of primary malignant tumor of the central nervous system. We have carried out a deep analysis of the secretome of a rapidly proliferating and tumorigenic cell line HNGC-2, representing GBM, in an effort to identify proteins, which may be targeted in the plasma of GBM patients as markers for diagnosis and disease surveillance. Prefractionation of the proteins from the conditioned medium of HNGC-2 cells in SDS gels followed by LC-MS/MS analysis using an ESI-IT mass spectrometer (LTQ) led to a total of 996 protein identifications with ≥2 peptides each. Of them, 664 proteins were observed in the transcriptome of HNGC-2 cells. The dataset of 996 proteins was mapped to important functional groups, such as cellular assembly and organisation, DNA recombination and repair, and other classes. Actin cytoskeleton signalling, phosphatidyl inositol 3 kinase (PI3K/AKT) and integrin linked kinase (ILK) signalling pathways were seen as enriched pathways. Comparisons with the published secretome of cell lines from 12 different cancers, including GBM, revealed that 348 proteins shared a commonality with a secretome of at least one other cell line, 321 of which were found to contain signal sequences or transmembrane domains and 335 could be linked to a plasma membrane or extracellular localization. Through intergration of this data we arrived at a non-redundant list of 597 protein identifications with the potential for secretion either by classical secretory pathways or by non-secretory processes; 233 of them have been detected in cerebrospinal fluid or plasma as per the published literature, and 172 have been implicated in GBM or other cancers. The HNGC-2 secretome dataset could serve as a useful resource for designing a targeted investigation of GBM biomarkers in plasma.

Glioblastoma cell secretome: analysis of three glioblastoma cell lines reveal 148 non-redundant proteins.

Glioblastoma multiforme (GBM) is the most aggressive among human gliomas with poor prognosis. Study of tumor cell secretome - proteins secreted by cancer cell lines, is a powerful approach to discover potential diagnostic or prognostic biomarkers. Here we report, for the first time, proteins secreted by three GBM cell lines, HNGC2, LN229 and U87MG. Analysis of the conditioned media of these cell lines by LC-MS/MS using ESI-IT mass spectrometer (LTQ) resulted in the confident identification of 102, 119 and 64 proteins, respectively. Integration of the results from all the three cell lines lead to a dataset of 148 non-redundant proteins. Subcellular classification using Genome Ontology indicated that 42% of the proteins identified belonged to extracellular or membrane proteins, viz. Vinculin, Tenascin XB, SERPIN F1 and TIMP-1. 52 proteins matched with the secretomes of 11 major cancer types reported earlier whereas remaining 96 are unique to our study. 25 protein identifications from the dataset represent proteins related to GBM or other cancer tissues as per Human Protein Atlas; at least 22 are detectable in plasma, 11 of them being reported even in cerebrospinal fluid. Our study thus provides a valuable resource of GBM cell secretome with potential for further investigation as GBM biomarkers.

Convergence pattern studies of folate receptor.

Gene patterns and sequences of folic acid synthesizing genes that are converged as meaningful patterns during evolution in the higher eukaryotes has been identified using sequence alignment and pattern analysis. Based on the finding, we are postulating that part of genes that are involved in synthesis of folic acid in lower eukaryotes, are converged into meaningful similar functional domains of folate receptor in higher eukaryotes.

Insilco analysis of functionally important residues in folate receptors.

Lack of crystal structure data of folate binding proteins has left so many questions unanswered (for example, important residues in active site, binding domain, important amino acid residues involved in interactions between ligand and receptor). With sequence alignment and PROSITE motif identification, we attempted to answer evolutionarily significant residues that are of functional importance for ligand binding and that form catalytic sites. We have analyzed 46 different FRs and FBP sequences of various organisms obtained from Genbank. Multiple sequence alignment identified 44 highly conserved identical amino acid residues with 10 cysteine residues and 12 motifs including ECSPNLGPW (which might help in the structural stability of FR).