Patterns of female dominance in Propithecus diadema edwardsi of Ranomafana National Park, Madagascar

Many lemur species are characterized by some form of female dominance, ranging from female feeding priority to complete female dominance, although this is a rare trait in primates and other mammals. The status of the Milne-Edwards' sifaka (Propithecus diadema edwardsi), a diurnal lemur, is ambiguous. Some short-term studies have found little or no aggression. The aim of the current, long-term study was to quantify the intersexual-dominance patterns of this sifaka. The distribution, outcome, and context of aggressive interactions were studied in four groups of wild sifakas. The majority of intersexual aggressive interactions were decided, with the loser expressing submissive behavior. Intersexual aggressive interactions occurred in all social contexts, and within all social contexts the females won the vast majority (92.7-96.0%) of aggressive interactions. While aggression rates were low (0.22/hr), this evidence suggests female dominance. We propose that female dominance exists because it provides a fitness advantage to both males and females.     

Influence of Heparin Mimetics on Assembly of the FGF·FGFR4 Signaling Complex

Fibroblast growth factor (FGF) signaling regulates mammalian development and metabolism, and its dysregulation is implicated in many inherited and acquired diseases, including cancer. Heparan sulfate glycosaminoglycans (HSGAGs) are essential for FGF signaling as they promote FGF·FGF receptor (FGFR) binding and dimerization. Using novel organic synthesis protocols to prepare homogeneously sulfated heparin mimetics (HM), including hexasaccharide (HM₆), octasaccharide (HM₈), and decasaccharide (HM₁₀), we tested the ability of these HM to support FGF1 and FGF2 signaling through FGFR4. Biological assays show that both HM₈ and HM₁₀ are significantly more potent than HM₆ in promoting FGF2-mediated FGFR4 signaling. In contrast, all three HM have comparable activity in promoting FGF1·FGFR4 signaling. To understand the molecular basis for these differential activities in FGF1/2·FGFR4 signaling, we used NMR spectroscopy, isothermal titration calorimetry, and size-exclusion chromatography to characterize binding interactions of FGF1/2 with the isolated Ig-domain 2 (D2) of FGFR4 in the presence of HM, and binary interactions of FGFs and D2 with HM. Our data confirm the existence of both a secondary FGF1·FGFR4 interaction site and a direct FGFR4·FGFR4 interaction site thus supporting the formation of the symmetric mode of FGF·FGFR dimerization in solution. Moreover, our results show that the observed higher activity of HM₈ relative to HM₆ in stimulating FGF2·FGFR4 signaling correlates with the higher affinity of HM₈ to bind and dimerize FGF2. Notably FGF2·HM₈ exhibits pronounced positive binding cooperativity. Based on our findings we propose a refined symmetric FGF·FGFR dimerization model, which incorporates the differential ability of HM to dimerize FGFs.     

Mutation of the cyclin-dependent kinase phosphorylation site in simian virus 40 (SV40) large T antigen specifically blocks SV40 origin DNA unwinding.

A mutant simian virus 40 (SV40) large tumor (T) antigen bearing alanine instead of threonine at residue 124 (T124A) failed to replicate SV40 DNA in infected monkey cells (J. Schneider and E. Fanning, J. Virol. 62:1598-1605, 1988). We investigated the biochemical properties of T124A T antigen in greater detail by using purified protein from a baculovirus expression system. Purified T124A is defective in SV40 DNA replication in vitro, but does bind specifically to the viral origin under the conditions normally used for DNA replication. The mutant protein forms double-hexamer complexes at the origin in an ATP-dependent fashion, although the binding reaction requires somewhat higher protein concentrations than the wild-type protein. Binding of T124A protein results in local distortion of the origin DNA similar to that observed with the wild-type protein. These findings indicate that the replication defect of T124A protein is not due to failure to recognize and occupy the origin. Under some conditions T124A is capable of unwinding short origin DNA fragments. However, the mutant protein is almost completely defective in unwinding of circular plasmid DNA molecules containing the SV40 origin. Since the helicase activity of T124A is essentially identical to that of the wild-type protein, we conclude that the mutant is defective in the initial opening of the duplex at the origin, possibly as a result of altered hexamer-hexamer interactions. The phenotype of T124A suggests a possible role for phosphorylation of threonine 124 by cyclin-dependent kinases in controlling the origin unwinding activity of T antigen in infected cells.     

Thermodynamic and kinetic characterization of ligand binding to the purine riboswitch aptamer domain

Riboswitches are cis-acting genetic regulatory elements found commonly in bacterial mRNAs that consist of a metabolite-responsive aptamer domain coupled to a regulatory switch. Purine riboswitches respond to intracellular concentrations of either adenine or guanine/hypoxanthine to control gene expression. The aptamer domain of the purine riboswitch contains a pyrimidine residue (Y74) that forms a Watson-Crick base-pairing interaction with the bound purine nucleobase ligand that discriminates between adenine and guanine. We sought to understand the structural basis of this specificity and the mechanism of ligand recognition by the purine riboswitch. Here, we present the 2,6-diaminopurine-bound structure of a C74U mutant of the xpt-pbuX guanine riboswitch, along with a detailed thermodynamic and kinetic analysis of nucleobase recognition by both the native and mutant riboswitches. These studies demonstrate clearly that the pyrimidine at position 74 is the sole determinant of purine riboswitch specificity. In addition, the mutant riboswitch binds adenine and adenine derivatives well compared with the guanine-responsive riboswitch. Under our experimental conditions, 2,6-diaminopurine binds the RNA with DeltaH=-40.3 kcal mol(-1), DeltaS=-97.6 cal mol(-1)K(-1), and DeltaG=-10.73 kcal mol(-1). A kinetic determination of the slow rate (0.15 x 10(5)M(-1)s(-1) and 2.1 x 10(5)mM(-1)s(-1) for 2-aminopurine binding the adenine-responsive mutant riboswitch and 7-deazaguanine-binding guanine riboswitch, respectively) of association under varying experimental conditions allowed us to propose a mechanism for ligand recognition by the purine riboswitch. A conformationally dynamic unliganded state for the binding pocket is stabilized first by the Watson-Crick base pairing between the ligand and Y74, and by the subsequent ordering of the J2/3 loop, enclosing the ligand within the three-way junction.     

Effects of serum type on growth and permeability properties of cultured endothelial cells

Serum is frequently added to defined basal media as a source of certain nutrients and macromolecular growth factors essential for cell growth. The many different sera commercially available may not be equally suitable for all cell types. The effects of four sera, fetal bovine serum (FBS), calf bovine serum (CS), equine serum (ES-1), and plasma-derived equine serum (ES-2), on growth and permeability properties of cultured porcine endothelial cells were determined. The rate of DNA synthesis, measured as [3H]thymidine incorporation, reached a peak at around 24 h, regardless of serum type, and was most marked with ES-1- or ES-2-treated cells. However, when estimated by total DNA, FBS, CS, or ES-1 treatment resulted in greater cell proliferation than ES-2. Based on protein synthetic rate and total cell protein, both FBS and CS appeared to be most growth supporting. At 72 h after cell plating, albumin passage across cultured endothelial monolayers was elevated in ES-1- and ES-2-treated cells compared with FBS- or CS-treated cells. "Leaky" cell monolayers were most marked with ES-1-treated cells. Cells grown in ES-2- and particularly in ES-1-enriched media were larger and more spindle-shaped compared with the typical cobblestone appearance of cells cultured in media enriched with either FBS or CS. These data suggest that CS, but not ES-1 or ES-2, is an excellent substitute for FBS to support desirable growth properties of macrovascular endothelial cells in culture.     

PAURODENDRON,A RHIZOMORPHIC LYCOPOD

From middle Pennsylvanian coal balls collected in Kansas and Illinois the ligules and rhizomorph of Paurodendron have been illustrated. Assignment of Paurodendron to the Lycopsida is corroborated. Discovery of the root-bearing portion of this Carboniferous lycopod establishes a diminutive, phyletic line in the Paleozoic with rhizomorphs smaller than many species of Isoeles. Paurodendron resembles the selaginelloid line more closely than other rhizomorphic types. Anatomical comparisons of Paurodendron with roots and root-bearing structures of lycopods are made; the root-bearing and a representative leaf-bearing portion of Paurodendron fraiponti have been reconstructed. The specific diagnosis has been emended, and the three species described by Fry have been reduced to one.     

Electrophoretic and immunologic characterization of proteins of merozoites of Eimeria acervulina, E. maxima, E. necatrix, and E. tenella.

Merozoites of Eimeria acervulina, Eimeria maxima, Eimeria necatrix, and Eimeria tenella were compared by gel electrophoresis, western-blotting with chicken antiserum, indirect fluorescent antibody reactions, and antiserum neutralization. Merozoites from the 4 species had dissimilar patterns of proteins and antigens in soluble and membrane fractions. Coomassie blue staining of SDS-PAGE gels revealed 16-22 protein bands depending on the species of merozoite but only 3 bands per species in the membrane fractions. Homologous and heterologous antisera recognized 5-12 soluble fraction bands and 3-7 membrane fraction bands on immunoperoxidase-stained western blots, depending on the species. When antisera from infected chickens were used in an indirect fluorescent antibody reaction, the merozoites of E. tenella and E. necatrix had a strong reaction with homologous and heterologous antisera. Merozoites of E. acervulina and E. maxima reacted with homologous antisera but had a weak or no reaction with heterologous antisera. Chicken antiserum against E. tenella had no effect on the viability of E. tenella merozoites when they were inoculated into chicken embryos.     

New generation pharmacogenomic tools: a SNP linkage disequilibrium Map,validated SNP assay resource,and high-throughput instrumentation system for large-scale genetic studies

Since public and private efforts announced the first draft of the human genome last year, researchers have reported great numbers of single nucleotide polymorphisms (SNPs). We believe that the availability of well-mapped, quality SNP markers constitutes the gateway to a revolution in genetics and personalized medicine that will lead to better diagnosis and treatment of common complex disorders. A new generation of tools and public SNP resources for pharmacogenomic and genetic studies--specifically for candidate-gene, candidate-region, and whole-genome association studies--will form part of the new scientific landscape. This will only be possible through the greater accessibility of SNP resources and superior high-throughput instrumentation-assay systems that enable affordable, highly productive large-scale genetic studies. We are contributing to this effort by developing a high-quality linkage disequilibrium SNP marker map and an accompanying set of ready-to-use, validated SNP assays across every gene in the human genome. This effort incorporates both the public sequence and SNP data sources, and Celera Genomics' human genome assembly and enormous resource ofphysically mapped SNPs (approximately 4,000,000 unique records). This article discusses our approach and methodology for designing the map, choosing quality SNPs, designing and validating these assays, and obtaining population frequency ofthe polymorphisms. We also discuss an advanced, high-performance SNP assay chemisty--a new generation of the TaqMan probe-based, 5' nuclease assay-and high-throughput instrumentation-software system for large-scale genotyping. We provide the new SNP map and validation information, validated SNP assays and reagents, and instrumentation systems as a novel resource for genetic discoveries.     

A novel cytokine-inducible gene CIS encodes an SH2-containing protein that binds to tyrosine-phosphorylated interleukin 3 and erythropoietin receptors.

Cytokines manifest their function through alteration of gene expression. However, target genes for signals from cytokine receptors are largely unknown. We therefore searched for immediate-early cytokine-responsive genes and isolated a novel gene, CIS (cytokine inducible SH2-containing protein) which is induced in hematopoietic cells by a subset of cytokines including interleukin 2 (IL2), IL3, granulocyte-macrophage colony-stimulating factor (GM-CSF) and erythropoietin (EPO), but not by stem cell factor, granulocyte colony-stimulating factor and IL6. The CIS message encodes a polypeptide of 257 amino acids that contains an SH2 domain of 96 amino acids in the middle. To clarify the function of CIS in cytokine signal transduction, we expressed CIS in IL3-dependent hematopoietic cell lines under the control of a steroid-inducible promoter. The CIS product stably associated with the tyrosine-phosphorylated beta chain of the IL3 receptor as well as the tyrosine-phosphorylated EPO receptor. Forced expression of CIS by steroid reduced the growth rate of these transformants, suggesting a negative role of CIS in signal transduction. CIS induction requires the membrane-proximal region of the cytoplasmic domain of the EPO receptor as well as that of the common beta chain of the IL3, IL5 and GM-CSF receptor, whereas CIS binds to the receptor that is tyrosine phosphorylated by cytokine stimulation. Thus CIS appears to be a unique regulatory molecule for cytokine signal transduction.