Spontaneous feline sporotrichosis: A fine structural study

Fine structural details of the parasitic yeastlike phase of Sporothrix schenckii contained in biopsy tissue from a naturally-occurring case of disseminated feline sporotrichosis are described and illustrated by electron microscopy. Both free and phagocytosed fungal cells were observed. The fungal cells were contained within an extracellular, electron transparent vacuolar area which was bounded by a limiting membrane of probable host origin. The yeastlike cells were characterized by a conspicuous layer of osmiophilic microfilaments which occurred along the outermost surface of the cell wall. In many yeastlike cells, scattered, membranebound vacuoles containing electron opaque material were observed in the cytoplasm. Asteroid bodies were not observed.     

8-Bromo-cyclic AMP induces phosphorylation of two sites in SRC-1 that facilitate ligand-independent activation of the chicken progesterone receptor and are critical for functional cooperation between SRC-1 and CREB binding protein

Elevation of intracellular 8-bromo-cyclic AMP (cAMP) can activate certain steroid receptors and enhance the ligand-dependent activation of most receptors. During ligand-independent activation of the chicken progesterone receptor (cPR(A)) with the protein kinase A (PKA) activator, 8-bromo-cAMP, we found no alteration in cPR(A) phosphorylation (W. Bai, B. G. Rowan, V. E. Allgood, B. W. O'Malley, and N. L. Weigel, J. Biol. Chem. 272:10457-10463, 1997). To determine if other receptor-associated cofactors were targets of cAMP-dependent signaling pathways, we examined the phosphorylation of steroid receptor coactivator 1 (SRC-1). We detected a 1.8-fold increase in SRC-1 phosphorylation in transfected COS-1 cells incubated with 8-bromo-cAMP. Phosphorylation was increased on two mitogen-activated protein kinase (MAPK) sites, threonine 1179 and serine 1185. PKA did not phosphorylate these sites in vitro. However, blockage of PKA activity in COS-1 cells with the PKA inhibitor (PKI) prevented the 8-bromo-cAMP-mediated phosphorylation of these sites. Incubation of COS-1 cells with 8-bromo-cAMP resulted in activation of the MAPK pathway, as determined by Western blotting with antibodies to the phosphorylated (active) form of Erk-1/2, suggesting an indirect pathway to SRC-1 phosphorylation. Mutation of threonine 1179 and serine 1185 to alanine in COS-1 cells coexpressing cPR(A) and the GRE(2)E1bCAT reporter resulted in up to a 50% decrease in coactivation during both ligand-independent activation and ligand-dependent activation. This was due, in part, to loss of functional cooperation between SRC-1 and CREB binding protein for coactivation of cPR(A). This is the first demonstration of cross talk between a signaling pathway and specific phosphorylation sites in a nuclear receptor coactivator that can regulate steroid receptor activation.     

Body Mass Index and the Prevalence of Hypertension and Dyslipidemia

Objective: To describe and evaluate relationships between body mass index (BMI) and blood pressure, cholesterol, high‐density lipoprotein‐cholesterol (HDL‐C), and hypertension and dyslipidemia. Research Methods and Procedures: A national survey of adults in the United States that included measurement of height, weight, blood pressure, and lipids (National Health and Nutrition Examination Survey III 1988–1994). Crude age‐adjusted, age‐specific means and proportions, and multivariate odds ratios that quantify the association between hypertension or dyslipidemia and BMI, controlling for race/ethnicity, education, and smoking habits are presented. Results: More than one‐half of the adult population is overweight (BMI of 25 to 29.9) or obese (BMI of ≥30). The prevalence of high blood pressure and mean levels of systolic and diastolic blood pressure increased as BMI increased at ages younger than 60 years. The prevalence of high blood cholesterol and mean levels of cholesterol were higher at BMI levels over 25 rather than below 25 but did not increase consistently with increasing BMI above 25. Rates of low HDL‐C increased and mean levels of HDL‐C decreased as levels of BMI increased. The associations of BMI with high blood pressure and abnormal lipids were statistically significant after controlling for age, race or ethnicity, education, and smoking; odds ratios were highest at ages 20 to 39 but most trends were apparent at older ages. Within BMI categories, hypertension was more prevalent and HDL‐C levels were higher in black than white or Mexican American men and women. Discussion: These data quantify the strong associations of BMI with hypertension and abnormal lipids. They are consistent with the national emphasis on prevention and control of overweight and obesity and indicate that blood pressure and cholesterol measurement and control are especially important for overweight and obese people.     

Development of an Assay for Phospholipase C Using Column-Reconstituted, Extruded Phospholipid Vesicles

The reconstitution of heterotrimeric G proteins into phospholipid vesicles has been widely used for the measurement of PLC-beta activity in vitro. We have developed an improved and sensitive method for the assay of PLC-beta activity. This approach involves reconstitution of purified betagamma dimers into extruded phospholipid vesicles containing phosphatidylinositol 4, 5-bisphosphate and using a gel-filtration technique to separate the reconstituted vesicles from monodispersed betagamma dimers and the detergent used to solubilize G proteins. The method provides physical information about the partitioning of betagamma dimers into phospholipid vesicles and was used to examine the effect of different prenyl groups on the gamma subunits in the activation of PLC-beta. The beta1gamma1 dimer (containing the farnesyl group) and the beta1gamma2 dimer (containing the geranylgeranyl group) were purified from baculovirus-infected Sf9 insect cells and were found to partition equally into phospholipid vesicles. The beta1gamma2 dimer is more potent and effective in stimulating PLC-beta activity than the beta1gamma1 dimer. The EC50 values of betagamma dimers for the activation of PLC-beta determined with this method were lower than those determined by previous methodology, showing that betagamma subunits have a subnanomolar affinity for PLC-beta.     

Drosophila acidic ribosomal protein rpA2: Sequence and characterization

A cDNA encoding the Drosophila melanogaster acidic ribosomal protein rpA2 was cloned and sequenced. rpA2 is homologous to the Artemia salina acidic ribosomal protein eL12′. In situ hybridization to salivary gland polytene chromosomes localizes the rpA2 gene to band 21C. It is a single copy gene, with an mRNA of 0.8 kb. Two-dimensional gel electrophoresis of Drosophila ribosomal proteins followed by immuno-blotting showed that the rpA2 protein has an apparent relative mobility in SDS of 17 kD and an isoelectric point less than pH 5.0. Although the Drosophila gene rp21C may be the same as rpA2, the reported sequences differ. Comparisons of the aligned nucleotide sequences coding for the acidic ribosomal proteins rpA1 and rpA2 of Drosophila with those of other eukaryotes support the view of two separate, though closely related, groups of acidic proteins. Comparison with the Artemia homologues suggests that nucleotide identity may have been conserved by some constraint that acts in addition to the requirement for substantial similarity of amino acid sequences. © 1993 Wiley-Liss, Inc.     

Two-dimensional protein electrophoretic analysis of postponed aging inDrosophila

Five populations ofDrosophila melanogaster that had been selected for postponed aging were compared with five control populations using two-dimensional protein gel electrophoresis. The goals of the study were to identify specific proteins associated with postponed aging and to survey the population genetics of the response to selection. A total of 321 proteins were resolvable per population; these proteins were scored according to their intensity. The resulting data were analyzed using resampling, combinatoric, and maximum parsimony methods. The analysis indicated that the populations with postponed aging were different from their controls with respect to specific proteins and with respect to the variation between populations. The populations selected for postponed aging were more heterogeneous between populations than were the control populations. Maximum parsimony trees separate the selected populations, as a group, from their controls, thereby exhibiting a homoplastic pattern.