Production of a fibronectin-associated lymphokine by cloned mouse T cells

Azobenzenearsonate-specific cloned mouse T cells able to transfer delayed hypersensitivity reactions in vivo produced macrophage agglutination factor (MaggF) after stimulation with mitogen or antigen in vitro. Mitogen (Con A) elicited MAggF production directly from T cells. Responses to Ag were Ag-specific, required syngeneic accessory cells in addition to T cells, and were independent of T cell fine specificity for azobenzenearsonate. Mouse MAggF shared a number of biochemical and immunochemical properties with the fibronectins (FN): 1) high Mr similar to that of plasma FN; 2) binding to gelatin, heparin, and polyclonal antibodies and mAb specific for cellular and plasma FN; 3) inhibition of activity in solution by monoclonal anti-human FN directed against plasma FN gelatin-binding domain; and 4) action on peritoneal exudate macrophages mediated through a FN-receptor cross reactive with one on human monocytes. MAggF production required active protein synthesis and was associated with significant increases in gelatin-binding immunoreactive FN (Mr 440 kDa on immunoblotting) in culture supernatants and T cell lysates. Metabolically labeled peptides could be precipitated by anti-FN from culture supernatants of activated T cells. Stimulated cultures contained significantly more cells with immunohistologically demonstrable cytoplasmic FN than unstimulated control cultures. We suggest that T cell FN is a distinct species of cellular FN which may play an important role in mediating delayed hypersensitivity inflammatory reactions in vivo.     

Larval rearing of fringe-lipped carp,Labeo fimbriatus (Bloch) with low cost diets including green bottle fly Lucilia sericata (Meigen) larvae meal incorporated diet

A study was conducted with non-conventional ingredients to test their efficacy as fishmeal (FM) replacers in the diet of fringe- lipped carp. Labeo fimbriatus first feeding larvae and fry were reared for 30 and 60 days in indoor, 50 L, aerated, circular plastic tanks at 100 and 30 numbers tank⁻¹, respectively. In the first feeding larvae to fry rearing experiment (Exp. 1), the fish were fed with either of the following isonitrogenous and isocaloric diets – live plankton, FM diet, green bottle fly (Lucilia sericata) larvae meal (GBFLM) diet and silkworm pupa (SWP) diet. The fry to fingerling rearing (Exp. 2), was also conducted using the same diets described above except live plankton. All compounded diets were formulated to contain 40% crude protein for the experiment 1 and 35% for experiment 2 and were fed ad libitum. Triplicate tanks were maintained for each treatment in both the experiments. In Exp. 1, the mean final weight of fry was higher with plankton and FM diets, while no difference (p > .05) was observed between FM and GBFLM diets. Weight of fish fed SWP diets was not statistically different from those fed GBFLM diet. No difference (p > .05) in final length, survival and condition factor was recorded. Analysis of digestive enzyme activity of whole fish revealed lower (p < .05) activity of amylase in fish fed plankton. In Exp. 2, no difference (p > .05) was observed between the different diet groups in terms of mean final weight, length, survival and condition factor. Analysis of digestive enzyme activity of whole fish revealed no difference (p > .05) in the activity of digestive enzymes between the treatments except a lower (p < .05) activity of trypsin in FM diet and lipase in FM and GBFLM diets. Since the survival and condition factors of animals is the most important aspect during nursery rearing, similar (p > .05) values recorded in different treatments indicate the possibility of incorporation of these non-conventional protein sources in the diet of L. fimbriatus during first feeding larvae to fry and fry to fingerling rearing.     

Rapid identification of potential drought tolerance genes from Solanum tuberosum by using a yeast functional screening method

Identification of major stress tolerance genes of a crop plant is important for the rapid development of its stress-tolerant cultivar. Here, we used a yeast functional screen method to identify potential drought-tolerance genes from a potato plant. A cDNA expression library was constructed from hyperosmotic stressed potato plants. The yeast transformants expressing different cDNAs were selected for their ability to survive in hyperosmotic stress conditions. The relative tolerances of the selected yeast transformants to multiple abiotic stresses were also studied. Specific potato cDNAs expressed in the tolerant yeast transformants were identified. Sixty-nine genes were found capable of enhancing hyperosmotic stress tolerance of yeast. Based on the relative tolerance data generated, 12 genes were selected, which could be most effective in imparting higher drought tolerance to potato with better survival in salt and high-temperature stresses. Orthologues of few genes identified here are previously known to increase osmotic stress tolerance of yeast and plants; however, specific studies are needed to confirm their role in the osmotic stress tolerance of potato.     

Enhancing the evaluation of PI3K inhibitors through 3D melanoma models

Targeted therapies for mutant BRAF metastatic melanoma are effective but not curative due to acquisition of resistance. PI3K signaling is a common mediator of therapy resistance in melanoma; thus, the need for effective PI3K inhibitors is critical. However, testing PI3K inhibitors in adherent cultures is not always reflective of their potential in vivo. To emphasize this, we compared PI3K inhibitors of different specificity in two‐ and three‐dimensional (2D, 3D) melanoma models and show that drug response predictions gain from evaluation using 3D models. Our results in 3D demonstrate the anti‐invasive potential of PI3K inhibitors and that drugs such as PX‐866 have beneficial activity in physiological models alone and when combined with BRAF inhibition. These assays finally help highlight pathway effectors that could be involved in drug response in different environments (e.g. p4E‐BP1). Our findings show the advantages of 3D melanoma models to enhance our understanding of PI3K inhibitors.     

An efficient algorithm for protein structure comparison using elastic shape analysis

Background

Protein structure comparison play important role in in silico functional prediction of a new protein. It is also used for understanding the evolutionary relationships among proteins. A variety of methods have been proposed in literature for comparing protein structures but they have their own limitations in terms of accuracy and complexity with respect to computational time and space. There is a need to improve the computational complexity in comparison/alignment of proteins through incorporation of important biological and structural properties in the existing techniques.

Results

An efficient algorithm has been developed for comparing protein structures using elastic shape analysis in which the sequence of 3D coordinates atoms of protein structures supplemented by additional auxiliary information from side-chain properties are incorporated. The protein structure is represented by a special function called square-root velocity function. Furthermore, singular value decomposition and dynamic programming have been employed for optimal rotation and optimal matching of the proteins, respectively. Also, geodesic distance has been calculated and used as the dissimilarity score between two protein structures. The performance of the developed algorithm is tested and found to be more efficient, i.e., running time reduced by 80–90 % without compromising accuracy of comparison when compared with the existing methods. Source codes for different functions have been developed in R. Also, user friendly web-based application called ProtSComp has been developed using above algorithm for comparing protein 3D structures and is accessible free.

Conclusions

The methodology and algorithm developed in this study is taking considerably less computational time without loss of accuracy (Table 2). The proposed algorithm is considering different criteria of representing protein structures using 3D coordinates of atoms and inclusion of residue wise molecular properties as auxiliary information.

     

Natural products from marine invertebrates against <Emphasis Type="Italic">Leishmania</Emphasis> parasites: a comprehensive review

Parasitic infections by Leishmania parasites remains a severe public health problem, especially in developing countries where it is highly endemic. Chemotherapy still remains a first option for the treatment of those diseases, despite the fact that available drugs exhibit a variety of shortcomings. Thus, innovative, less toxic more affordable and effective antileishmanial agents are urgently needed. The marine environment holds an immeasurable bio- and chemical diversity, being a valuable source of natural products with therapeutic potential. As invertebrates comprise about 60 % of all marine organisms, bioprospecting this class of organisms for antileishmanial properties may unravel unique and selective hit molecules. In this context, this review covers results on the literature of marine invertebrate extracts and pure compounds evaluated against Leishmania parasites mainly by in vitro methods. It comprises results obtained from the phyla Porifera, Cnidaria, Bryozoa (Ectoprota), Mollusca, Echinodermata, Annelida, Cetnophora, Platyhelminthes, sub phyla Crustacea (phylum Arthropoda) and Tunicata (phylum Chordata). Moreover, structure–activity relationships and possible mechanisms of action are mentioned, whenever available information is provided. About 70 species of marine invertebrates belonging to seven different phyla are included in this work. Besides a variety of crude extracts, a total of 140 pure compounds was tested against different Leishmania species. Although the research on the antileishmanial potential of marine invertebrates is in its early beginnings, promising results have been achieved that encourage further research. As more extracts and compounds are being screened, the possibility of finding active and selective antileishmanial molecules increases, rising new hope in the search for new treatments against leishmaniases.     

Real‐time PCR Detection of Sorghum Ergot Pathogens Claviceps africana,Claviceps sorghi and Claviceps sorghicola

Sorghum ergot is a serious disease that has caused major losses in sorghum growing regions worldwide. Claviceps africana, originally reported from Zimbabwe, is now the most widely distributed species causing ergot in many countries including the United States of America, whereas both C. africana and Claviceps sorghi exist in India. A third species (Claviceps sorghicola) has been described causing sorghum ergot in Japan. As the three species show morphological similarities, a DNA‐based assay is desirable for rapid identification in cases where ergot‐infected sorghum is found by regulatory authorities. We designed PCR primers and probes from the intron 3 region of the β‐tubulin gene (for C. africana and C. sorghi) and the intron 4 region of EF‐1α (for C. sorghicola) and tested them by real‐time PCR with purified DNA and ergot samples from the field and greenhouse. The primer and probe sets specifically amplified DNA from the respective species with a detection limit of c. 1 pg DNA. Genomic DNA from six other Claviceps species did not amplify in any of the three ergot species‐specific assays. The assays we describe will provide useful tools for detecting sorghum ergot pathogens in seed and grain shipments and for determining which species are present in the samples, thereby aiding in the regulatory decision‐making process.     

Evidence that a protein-protein interaction 'hot spot' on heterotrimeric G protein betagamma subunits is used for recognition of a subclass of effectors

To understand the requirements for binding to G protein betagamma subunits, phage-displayed random peptide libraries were screened using immobilized biotinylated betagamma as the target. Selected peptides were grouped into four different families based on their sequence characteristics. One group (group I) had a clear conserved motif that has significant homology to peptides derived from phospholipase C beta (PLC beta) and to a short motif in phosducin that binds to G protein beta subunits. The other groups had weaker sequence homologies or no homology to the group I sequences. A synthetic peptide from the strongest consensus group blocked activation of PLC by G protein betagamma subunits. The peptide did not block betagamma-mediated inhibition of voltage-gated calcium channels and had little effect on betagamma-mediated inhibition of Gs-stimulated type I adenylate cyclase. Competition experiments indicated that peptides from all four families bound to a single site on betagamma. These peptides may bind to a protein-protein interaction 'hot spot' on the surface of betagamma subunits that is used by a subclass of effectors.