La3+-induced fusion of phosphatidylserine liposomes. Close approach, intermembrane intermediates, and the electrostatic surface potential.

The fusion of large unilamellar phosphatidylserine liposomes (PS LUV) induced by La3+ has been monitored using the 1-aminoapthalene-3,6,8-trisulfonic acid/p-xylenebis(pyridinium bromide) (ANTS/DPX) fluorescence assay for the mixing of aqueous contents. The fusion event is extensive and nonleaky, with up to 95% mixing of contents in the fused liposomes. However, addition of excess EDTA leads to disruption of the fusion products in a way that implies the existence of metastable intermembrane contact sites. The maximal fusion activity occurs between 10 and 100 microM La3+ and fusion can be terminated rapidly, without loss of contents, by the addition of excess La3+, e.g., 1 mM La3+ at pH 7.4. This observation is explained by the very large intrinsic binding constant (approximately 10(5) M-1) of La3+ to the PS headgroup, as measured by microelectrophoresis. Addition of 1 mM La3+ causes charge reversal of the membrane and a large positive surface potential. La3+ binding to PS causes the release of a proton. These data can be explained if La3+ can chelate to PS at two sites, with one of the sites being the primary amino group. This binding model successfully predicts that at pH 4.5 fusion occurs up to 2 mM La3+, due to reduced La3+ binding at low pH. We conclude that the general mechanism of membrane fusion includes three kinetic steps. In addition to (a) aggregation, there is (b) the close approach of the surfaces, or thinning of the hydration layer, and (c) the formation of intermembrane intermediates which determine the extent to which membrane destabilization leads to fusion (mixing of aqueous contents), as opposed to lysis. The lifetime of these intermembrane intermediates appears to depend upon La3+ binding to both PS sites.     

DINAMO: interactive protein alignment and model building.

MOTIVATION: To facilitate the process of structure prediction by both comparative modeling and fold recognition, we describe DINAMO, an interactive protein alignment building and model evaluation tool that dynamically couples a multiple sequence alignment editor to a molecular graphics display. DINAMO allows the user to optimize the alignment and model to satisfy the known heuristics of protein structure by means of a set of analysis tools. The analysis tools return information to both the alignment editor and graphics model in the form of visual cues (color, shape), allowing for rapid evaluation. Several analysis tools may be employed, including residue conservation, residue properties (charge, hydrophobicity, volume), residue environmental preference, and secondary structure propensity. RESULTS: We demonstrate DINAMO by building a model for submission in the 3rd annual Critical Assessment of Techniques for Protein Structure Prediction (CASP3) contest. AVAILABILITY: DINAMO is freely available as a local application or Web-based Java applet at http://tito.ucsc.edu/dinamo     

Cartilage-inducing factor-A. Apparent identity to transforming growth factor-beta

Comparison of the sequence of the N-terminal 30 amino acids of cartilage-inducing factor-A (CIF-A) from bovine demineralized bone with the corresponding sequence of human transforming growth factor-beta (TGF-beta) revealed 100% identity. Furthermore, CIF-A stimulated normal rat kidney fibroblasts to become anchorage-independent and form colonies in soft agar (in the presence of epidermal growth factor) in a manner similar to TGF-beta. Similarly, TGF-beta from human platelets induced rat muscle mesenchymal cells to differentiate and synthesize cartilage-specific macromolecules in a manner equivalent to CIF-A. These data show that CIF-A and TGF-beta are closely related or identical molecules and that these factors may be involved in cell differentiation including cartilage formation as the first step in endochondral bone formation.     

Gastrointestinal and immunological responses of senior dogs to chicory and mannan-oligosaccharides

Thirty-four senior dogs (pointers 8 - 11 years, beagles 9 - 11 years) were used to evaluate the effects of oligosaccharides on nutritional and immunological characteristics. Dogs were randomly allotted to treatments [1% chicory (CH), 1% mannan-oligosaccharide (MOS), 1% chicory + 1% MOS (CM), or no supplementation (control, CON)] in a parallel design with a 4 week baseline period followed by a 4 week treatment period. Dietary supplementation with MOS or CM tended (P = 0.07) to increase food intake due, in part, to an increase in fermentable fibre and a decrease in energy content of the diet. Although wet faecal output increased (P < 0.05) for dogs supplemented with MOS or CM, when corrected for food intake, no differences were noted. The CM treatment increased (P < 0.05) faecal score (1 = hard and dry, 5 = watery liquid), although these scores remained in a desirable range (3 to 3.5). Chicory supplementation increased (P = 0.07) fat digestibility. Chicory or MOS increased (P  0.05) faecal bifidobacteria concentrations 0.4 and 0.5 log₁₀ cfu/g DM, respectively, compared to the CON, while MOS decreased (P < 0.05) faecal E. coli concentrations. Oligosaccharides did not affect white blood cell (WBC) concentrations, but CH and CM tended to increase (P = 0.10) neutrophil concentrations compared to control dogs. Peripheral lymphocyte concentrations were decreased in dogs supplemented with MOS (P = 0.06) and CM (P < 0.05). Chicory and MOS alter faecal microbial populations and certain indices of the immune system of senior dogs.     

The Quilon Limestone,Kerala Basin,India: an archive for Miocene Indo‐Pacific seagrass beds

Reuter, M., Piller, W.E., Harzhauser, M., Kroh, A., Rögl, F. & ?ori?, S. 2010: The Quilon Limestone, Kerala Basin, India: an archive for Miocene Indo‐Pacific seagrass beds. Lethaia, Vol. 44, pp. 76–86. The facies of the fossiliferous Quilon Limestone in SW India is described for the first time in detail at the Padappakkara‐type locality. Facies (fossiliferous, micrite‐rich, bioturbated sediment with intercalated sand pockets) and faunal composition (epiphytic foraminifers, seagrass feeding Smaragdia gastropods, bioimmuration of celleporiform bryozoan colonies) indicate a seagrass environment. The large discoidal archaiasin foraminifer Pseudotaberina malabarica, in particular, is considered as a proxy for seagrass communities. Recent seagrasses have their centre of generic richness in the Indo‐Pacific where they cover wide areas in the tidal and shallow sub‐tidal zones. However, their geological record is only fragmentary and their palaeobiogeographic distribution has a big stratigraphical gap in the Miocene Western Indo‐Pacific region. The described nannoplankton flora and planktonic foraminifers from the Quilon Formation demonstrate that the deposition of the studied seagrass bed occurred in nannoplankton biozone NN3. This timing suggests formation during the closure of the Tethyan Seaway. The Quilon Limestone is thus an early Western Indo‐Pacific seagrass bed and an important step in reconstructing the history of seagrass communities. □Quilon Formation, Pseudotaberina malabarica, seagrass facies, Burdigalian, Indo‐Pacific.