Proteomic profiling of serum samples from chikungunya-infected patients provides insights into host response

Background

Chikungunya is a highly debilitating febrile illness caused by Chikungunya virus, a single-stranded RNA virus, which is transmitted by Aedes aegypti or Aedes albopictus mosquito species. The pathogenesis and host responses in individuals infected with the chikungunya virus are not well understood at the molecular level. We carried out proteomic profiling of serum samples from chikungunya patients in order to identify molecules associated with the host response to infection by this virus.

Results

Proteomic profiling of serum obtained from the infected individuals resulted in identification of 569 proteins. Of these, 63 proteins were found to be differentially expressed (≥ 2-fold) in patient as compared to control sera. These differentially expressed proteins were involved in various processes such as lipid metabolism, immune response, transport, signal transduction and apoptosis.

Conclusions

This is the first report providing a global proteomic profile of serum samples from individuals infected with the chikungunya virus. Our data provide an insight into the proteins that are involved as host response factors during an infection. These proteins include clusterin, apolipoproteins and S100A family of proteins.     

Prolongation of insulin-induced activation of mitogen-activated protein kinases ERK 1/2 and phosphatidylinositol 3-kinase by vanadyl sulfate, a protein tyrosine phosphatase inhibitor

Vanadium salts such as vanadyl sulfate (VS), potent inhibitors of protein tyrosine phosphatases, have been shown to mimic, augment, and prolong insulin's action. However, the molecular mechanism of responses to these salts is not clear. In the present studies, we examined if VS-induced effects on insulin action are associated with enhancement or augmentation in the activation state of key components of the insulin signaling pathway. Treatment of insulin receptor-overexpressing cells with insulin or VS resulted in a time-dependent transient increase in phosphorylation and activation of extracellular signal-regulated kinases 1 and 2 (ERK 1/2) that peaked at about 5 min, then declined rapidly to about baseline within 30 min. However, when the cells were treated with VS before stimulation with insulin, sustained ERK 1/2 phosphorylation and activation were observed well beyond 60 min. VS treatment also prolonged the insulin-stimulated activation of phosphatidylinositol 3-kinase (PI3-K), which was associated with sustained interaction between insulin receptor substrate-1 (IRS-1) and the p(85 alpha) subunit of phosphatidylinositol 3-kinase (PI3-K) in response to insulin. These data indicate that prolongation of insulin-stimulated ERK 1/2 and PI3-K activation by VS is due to a more stable complex formation of IRS-1 with the p(85 alpha) subunit which may, in turn, be responsible for its ability to enhance and extend the biological effects of insulin.     

Mutational Engineering of the cyanobacterium Nostoc muscorum for resistance to growth-inhibitory action of LiCl and NaCl

The effect of NaCl on two vital processes of cyanobacterial metabolism, viz. N(2) fixation and oxygenic photosynthesis, was studied in the cyanobacterium Nostoc muscorum grown diazotrophically. An increase in NaCl concentration suppressed the formation of heterocyst and adversely affected the nitrogenase activity in the parent, whereas in Li(+)-R and Na(+)-R mutants NaCl stress did not cause any adverse effect. The rate of photosynthetic O(2)-evolution was also adversely affected by the NaCl stress, but the magnitude was less than that of nitrogenase activity. L-Proline, the well-known osmoprotectant, provided protection to the cyanobacterium against NaCl stress. The parent strain utilized L-proline as a nitrogen source and suppressed heterocyst formation and nitrogenase activity, while mutants showed normal heterocyst frequency and nitrogenase activity. Therefore, it may be that the proline metabolism is altered as a result of mutation. The intracellular levels of proline in the parent were enhanced about threefold in the medium containing 1 mol x m(-3) proline, while in mutants there was no significant increase in the intracellular level of proline. In the medium containing both NaCl and proline, the intracellular level of proline was enhanced in the parent as well as in both mutant strains. This suggests that the parent strain possessed both normal proline uptake and salt-induced proline uptake systems, whereas the mutant strains were defective in normal proline uptake and had only salt-induced proline uptake. The over-accumulation of proline in the presence of NaCl stress is due either to the loss of proline oxidase activity or to the accumulation of exogenous proline.     

Association mapping of agro-morphological characters among the global collection of finger millet genotypes using genomic SSR markers

Identification of alleles responsible for various agro-morphological characters is a major concern to further improve the finger millet germplasm. Forty-six genomic SSRs were used for genetic analysis and population structure analysis of a global collection of 190 finger millet genotypes and fifteen agro-morphological characters were evaluated. The overall results showed that Asian genotypes were smaller in height, smaller flag leaf length, less basal tiller number, early flowering and early maturity nature, small ear head length, and smaller in length of longest finger. The 46 SSRs yielded 90 scorable alleles and the polymorphism information content values varied from 0.292 to 0.703 at an average of 0.442. The gene diversity was in the range of 0.355 to 0.750 with an average value of 0.528. The 46 genomic SSR loci grouped the 190 finger millet genotypes into two major clusters based on their geographical origin by the both phylogenetic clustering and population structure analysis by STRUCTURE software. Association mapping of QTLs for 15 agro-morphological characters with 46 genomic SSRs resulted in identification of five markers were linked to QTLs of four traits at a significant threshold (P) level of ≤0.01 and ≤0.001. The QTL for basal tiller number was strongly associated with the locus UGEP81 at a P value of 0.001 by explaining the phenotypic variance (R ²) of 10.8 %. The QTL for days to 50 % flowering was linked by two SSR loci UGEP77 and UGEP90, explained 10 and 8.7 % of R ² respectively at a P value of 0.01. The SSR marker, FM9 found to have strong association to two agro-morphological traits, flag leaf width (P—0.001, R ²—14.1 %) and plant height (P—0.001, R ²—11.2 %). The markers linked to the QTLs for above agro-morphological characters found in the present study can be further used for cloning of the full length gene, fine mapping and their further use in the marker assisted breeding programmes for introgression of alleles into locally well adapted germplasm.     

Parameter determination and validation for a mechanistic model of the enzymatic saccharification of cellulose‐Iβ

Cost‐effective production of fuels and chemicals from lignocellulosic biomass often involves enzymatic saccharification, which has been the subject of intense research and development. Recently, a mechanistic model for the enzymatic saccharification of cellulose has been developed that accounts for distribution of cellulose chain lengths, the accessibility of insoluble cellulose to enzymes, and the distinct modes of action of the component cellulases [Griggs et al. (2012) Biotechnol. Bioeng., 109(3):665–675; Griggs et al. (2012) Biotechnol. Bioeng., 109(3):676–685]. However, determining appropriate values for the adsorption, inhibition, and rate parameters required further experimental investigation. In this work, we performed several sets of experiments to aid in parameter estimation and to quantitatively validate the model. Cellulosic materials differing in degrees of polymerization and crystallinity (α‐cellulose‐I_β and highly crystalline cellulose‐I_β) were digested by component enzymes (EG_I/CBH_I/ ) and by mixtures of these enzymes. Based on information from the literature and the results from these experiments, a single set of model parameters was determined, and the model simulation results using this set of parameters were compared with the experimental data of total glucan conversion, chain‐length distribution, and crystallinity. Model simulations show significant agreement with the experimentally derived glucan conversion and chain‐length distribution curves and provide interesting insights into multiple complex and interacting physico‐chemical phenomena involved in enzymatic hydrolysis, including enzyme synergism, substrate accessibility, cellulose chain length distribution and crystallinity, and inhibition of cellulases by soluble sugars. © 2015 American Institute of Chemical Engineers Biotechnol. Prog., 31:1237–1248, 2015     

Exploratory algorithm to devise multi-epitope subunit vaccine by investigating Leishmania donovani membrane proteins

Visceral leishmaniasis (VL) is a deadly parasitic infection which affects poorest to poor population living in the endemic countries. Increasing resistant to existing drugs, disease burden and a significant number of deaths, necessitates the need for an effective vaccine to prevent the VL infection. This study employed a combinatorial approach to develop a multi-epitope subunit vaccine by exploiting Leishmania donovani membrane proteins. Cytotoxic T- and helper T-lymphocyte binding epitopes along with suitable adjuvant and linkers were joined together in a sequential manner to design the subunit vaccine. The occurrence of B-cell and IFN-γ inducing epitopes approves the ability of subunit vaccine to develop humoral and cell-mediated immune response. Physiochemical parameters of vaccine protein were also assessed followed by homology modeling, model refinement and validation. Moreover, disulfide engineering was performed for the increasing stability of the designed vaccine and molecular dynamics simulation was performed for the comparative stability purposes and to conform the geometric conformations. Further, molecular docking and molecular dynamics simulation study of a mutated and non-mutated subunit vaccine against TLR-4 immune receptor were performed and respective complex stability was determined. In silico cloning ensures the expression of designed vaccine in pET28a(+) expression vector. This study offers a cost-effective and time-saving way to design a novel immunogenic vaccine that could be used to prevent VL infection.

Communicated by Ramaswamy H. Sarma