Nucleotide sequence of a cDNA from Onchocerca gibsoni encoding a novel repetitive antigen.

mRNA from uterine microfilariae of the cattle parasite Onchocerca gibsoni was used for the construction of cDNA libraries. A cDNA clone encoding an antigen recognized by serum from human individuals infected with O. volvulus was found to contain five copies of an 87 bp unit. These 87 bp units were present in the genome in high copy number as long tandem arrays. These are the first cDNA sequence data obtained directly from larvae of any Onchocerca species.     

Identification and characterization of the murine cell surface receptor for the urokinase-type plasminogen activator.

Cell-binding experiments have indicated that murine cells on their surface have specific binding sites for mouse urokinase-type plasminogen activator (u-PA). In contrast to the human system, chemical cross-linking studies with an iodinated ligand did not yield any covalent adducts in the murine system, but in ligand-blotting analysis, two mouse u-PA-binding proteins could be visualized. To confirm that these proteins are the murine counterpart of the human u-PA receptor (u-PAR), a peptide was derived from the murine cDNA clone assigned to represent the murine u-PAR due to cross-hybridization and pronounced sequence similarity with human u-PAR cDNA [Kristensen, P., Eriksen, J., Blasi, F. & Dan?, K. (1991) J. Cell Biol. 115, 1763-1771]. A rabbit antiserum raised against this peptide specifically recognized two polypeptide bands with electrophoretic mobilities identical to those identified by ligand-blotting analysis. Binding of mouse u-PA to its receptor showed species specificity in ligand-blotting analysis, since mouse u-PA did not bind to human u-PAR and human u-PA did not bind to mouse u-PAR. The apparent M(r) of mouse u-PAR varied between different mouse cell lines and ranged over M(r) 45,000-60,000. In four of the cell lines, mouse u-PA bound to two mouse u-PAR variant proteins, whereas in the other two cell lines studied, there was only one mouse u-PA-binding protein. In the monocyte macrophage cell line P388D.1, trypsin-treatment of intact cells could remove only the large mouse u-PAR variant (M(r) 60,000) indicating that only this type was a cell-surface-exposed molecule. The smaller mouse u-PAR variant (M(r) 45,000), was deglycosylated by the enzyme endo-beta-N-acetylglucosaminidase H and is probably an intracellular precursor form carrying only high-mannose carbohydrate. Deglycosylation of this variant yielded a polypeptide with an apparent M(r) of about 30,000, which corresponds to the Mr calculated from the cDNA derived protein sequence of mouse u-PAR. Receptor-bound mouse u-PA could be released by phosphatidylinositol-specific phospholipase C treatment, indicating that mouse u-PAR is attached to the cell surface by glycosylphosphatidylinositol. Purification of the two mouse u-PAR variant proteins by diisopropylfluorophosphate-inactivated mouse u-PA-Sepharose affinity chromatography yielded two silver-stained bands when analysed by SDS/PAGE, corresponding in electrophoretic mobility to those seen by ligand-blotting analysis.(ABSTRACT TRUNCATED AT 400 WORDS)     

Interactions among coexisting larval Odonata: an in situ experiment using small enclosures

Field experiments using small replicated enclosures focused on interactions between larval populations of Epitheca cynosura and Ladona deplanata (Odonata: Anisoptera) — two species that emerge in early spring. The presence of Epitheca reduced the total biomass of Ladona, but Ladona had no significant effect on Epitheca. These early-emerging species reduced the biomass of small instars of late-emerging Anisoptera which colonized enclosures during the experiments; and the late-emerging Anisoptera seem to have inhibited colonization by Zygoptera larvae. Results are consistent with the importance of predatory (cannibalism or mutual predation) interactions in this community.     

Extraction and immunochemical assays of a tubulin-like factor in cotton seedlings

Antibodies to tubulin were prepared in rabbits by immunization with reduced-carboxymethylated calf-brain tubulin. In immunodiffusion tests the antibodies showed full cross reactivity with the immunogen as well as with native calf-brain tubulin. The same antibodies showed cross reactivity with a factor in extract of cotton (Gossypium hirsutum L.) cotyledons but there was no full immunological identity between calf-brain tubulin and this factor. A solid-phase radioimmunoassay for quantitative estimation of this plant tubulin-like factor was developed. It measured the binding of antibodies to immobilized calf-native tubulin. Competition between the unknown soluble tubulin-like factor, and immobilized tubulin was assayed at serum dilution of 1:50. Extraction conditions which preserved the antigenic properties of the tubulin-like factor from cotton cotyledons were defined. The radioimmunoassay measured quantities of the tubulin-like factor in the range of 0.1–10 g-equivalents of calf-brain tubulin. Immediately after homogenization of the tissue only 25% of the total amount of tubulin-like activity was present in soluble form, while most of it remained in the insoluble fraction. Apparent maximal solubilization was achieved spontaneously 10 h after homogenization or by treatment with guanidine hydrochloride. These results indicate that in this material, tubulin is not released immediately by homogenization but remains assembled in microtubules and-or in a bound or sequestered form.Abbreviations NRS normal rabbit serum - RCM reduced-carboxymethylated     

Site-specific integration of an F'' lac pro factor in the region of the replication origin (oriC) of E. coli

Summary An episome, F 128, which carries approximately 8x10⁴ base pairs of chromosomal DNA homologous to the lac pro region of the E. coli chromosome, has been found to integrate into the oriC region of the chromosome in a site specific reaction. While the event appears to be recA-dependent, no homology between the episome and this region of the chromosome was detected. The Hfr strains formed result from the integration of intact F 128 molecules. The structure of the Hfr strains generated has been determined and their transfer properties analyzed.     

Quantitative structure-activity relationships (QSAR) in sweet aspartyl dipeptide methyl esters

In drug design the pharmacochemists frequently use physicochemicalconstants to correlate the structure with the observed potency.Curiously this approach has hardly been followed by psychophysiciststo indicate the increase of taste potency in a series of structurallyrelated compounds of the same stimulus. In the present experiments we correlated the relative sweetness(S) of 40 aspartyl dipeptide methyl esters [general formulaCH₂(COO^–). CH(NH₃⁺ ).CO.NH.CH(R).COOCH₃] with 8 physicochemicalparameters. Among the compounds we had 7 non-sweet stimuli whilethe potency of the remaining 33 peptide esters varied from 1to 27,000 (1 = sucrose). We calculated for the side chain Rthe values of the parachor parameter P, the hydrophobic fragmentalconstant f and 5 STERIMOL parameters (L, B₁ up to B₅). A multipleregression analysis programme selected by stages the most relevantparameters and tested their significance. We observed that the criterion whether a dipeptide ester issweet or not, is among others defined by the L and B₅ parametersof the side chain R. Compounds are sweet provided L is confinedto certain limits (0.50 nm<L<0.62 nm), or otherwise whenL exceeds these limits, the B₅ parameter has to be greater than0.45 nm (when L<0.50 nm) or smaller than 0.72 nm (when L>0.62nm). The sweet potency defined as log S correlated very significantlywith the parameters P and B₄ (n=33, r=0.812, s=0.60, F=29.06).When two compounds, which were shown to be situated at the borderlineof the length and volume parameters, were omitted in the analysis,the correlation improved (n=31, r=0.909, s=0.40, F=42.60). Inthe latter situation we found the following equation when theintercept was set at zero: log S=0.194f + 1.472.10^–2P—3.357B₅ A previously proposed conformation of aspartame (R=CH₂-Øat the receptor site was computed in detail. We calculated thedistances of the AH-B moieties to the third binding site (thecentre of Ø) and indicated the width of the receptoraccess for this series of sweet, structurally related, dipeptidemethyl esters.     

Using all Gene Families Vastly Expands Data Available for Phylogenomic Inference

Traditionally, single-copy orthologs have been the gold standard in phylogenomics. Most phylogenomic studies identify putative single-copy orthologs using clustering approaches and retain families with a single sequence per species. This limits the amount of data available by excluding larger families. Recent advances have suggested several ways to include data from larger families. For instance, tree-based decomposition methods facilitate the extraction of orthologs from large families. Additionally, several methods for species tree inference are robust to the inclusion of paralogs and could use all of the data from larger families. Here, we explore the effects of using all families for phylogenetic inference by examining relationships among 26 primate species in detail and by analyzing five additional data sets. We compare single-copy families, orthologs extracted using tree-based decomposition approaches, and all families with all data. We explore several species tree inference methods, finding that identical trees are returned across nearly all subsets of the data and methods for primates. The relationships among Platyrrhini remain contentious; however, the species tree inference method matters more than the subset of data used. Using data from larger gene families drastically increases the number of genes available and leads to consistent estimates of branch lengths, nodal certainty and concordance, and inferences of introgression in primates. For the other data sets, topological inferences are consistent whether single-copy families or orthologs extracted using decomposition approaches are analyzed. Using larger gene families is a promising approach to include more data in phylogenomics without sacrificing accuracy, at least when high-quality genomes are available.