New cellulose synthesizing complexes (terminal complexes) involved in animal cellulose biosynthesis in the tunicateMetandrocarpa uedai

Summary Subcellular compartments comprising the endomembrane system in filamentous fungi are poorly characterized with most showing significant morphological differences from eukaryotic cells. For example, many filamentous fungi lack stacked Golgi-body cisternae, but contain Golgi equivalents — single cisternae or tubules which appear to serve the same functions. To help identify fungal endomembrane compartments and interrelationships between them we used a pharmacological agent, brefeldin A, known to affect specific endomembrane organelles in other organisms, most prominently the Golgi apparatus. At 10 g/ml brefeldin A, radial hyphal growth of the rice blast pathogenMagnaporthe grisea on solid agar medium was reduced by 96% over an initial 48 h, but recovered and was reduced by only 20% over a subsequent 72 h exposure. Light microscopic examination of individual living hyphae showed that apical elongation generally halted within 1 min after exposure to brefeldin A. Acute effects of 14 g/ml brefeldin A were characterized ultrasiructurally in cells prepared by freeze substitution. These included the appearance of two types of cisternae with unusual morphology, associated with ca. 45 nm diameter vesicles, as well as the unexpected persistence and increase in complexity of the Golgi equivalents. Also observed were (1) reduced numbers of apicale vesicles and disruption of Spitzenkörper organization, (2) apical clusters of 30–35 nm diameter microvesicles and associated tubular arrays, (3) dilation of rough endoplasmic reticulum, (4) packets of membrane-bounded electron-opaque cell wall inclusions, and (5) altered morphology of some vacuolar compartments. The distribution of concanavalin A binding sites, previously mapped to particular endomembrane compartments, was documented to aid the interpretation of these results. We conclude that brefeldin A effects on cells ofM. grisea differ from those reported with plant and animal cells, perhaps reflecting underlying differences in the endomembrane systems among these eukaryotes.Abbreviations BFA brefeldin A - ConA concanavalin A - ER endoplasmic reticulum - PDA potato dextrose agar - RER rough endoplasmic reticulum     

Bilateral giant myelolipoma in the adrenal of a cotton-top tamarin (Saguinus oedipus)

Abstract: A rare case of bilateral adrenal myelolipomas in a female cotton-top tamarin is reported. Large bilateral masses in the adrenal glands were composed of mature adipose cells containing varying amounts of hematopoietic cells of the myeloid, erythroid, and megakaryocyte series. The gross and histologic features of this case closely resemble human “giant” adrenal myelolipomas.     

Purification and characterization of microbial gellan lyase.

Gellan lyase was purified from the culture fluid of soil samples incubated in a medium containing gellan as a sole carbon source. The enzyme was a monomer with a molecular mass of 140 kDa and was most active at pH 7.5 and 45 degrees C. The enzyme was highly specific to gellan and lowered the viscosity of the polymer.     

The carboxy-terminal region of mammalian HSP90 is required for its dimerization and function in vivo.

The majority of mouse HSP90 exists as alpha-alpha and beta-beta homodimers. Truncation of the 15-kDa carboxy-terminal region of mouse HSP90 by digestion with the Ca(2+)-dependent protease m-calpain caused dissociation of the dimer. When expressed in a reticulocyte lysate, the full-length human HSP90 alpha formed a dimeric form. A plasmid harboring human HSP90 alpha cDNA was constructed so that the carboxy-terminal 49 amino acid residues were removed when translated in vitro. This carboxy-terminally truncated human HSP90 alpha was found to exist as a monomer. In contrast, loss of the 118 amino acid residues from the amino terminus of human HSP90 alpha did not affect its in vitro dimerization. Introduction of an expression plasmid harboring the full-length human HSP90 alpha complements the lethality caused by the double mutations of two HSP90-related genes, hsp82 and hsc82, in a haploid strain of Saccharomyces cerevisiae. The carboxy-terminally truncated human HSP90 alpha neither formed dimers in yeast cells nor rescued the lethal double mutant.     

Ascorbic Acid Inhibits 125I-SCH 23982 Binding but Increases the Affinity of Dopamine for D1 Dopamine Receptors

Abstract: Effects of ascorbic acid (AA) on ¹²⁵I-SCH 23982 binding to D₁ dopaminergic receptors in membrane preparations from rat striatum were investigated. AA in the range of 0.03 µ M –0.33 m M inhibited 75% of specific binding of ¹²⁵I-SCH 23982 in a dose-dependent manner. At higher concentrations, this inhibition of binding activity by AA was less potent, and 3.3 m M AA inhibited only 30% of specific binding. Reduced glutathione did not alter the inhibition of binding by 0.33 m M AA, but reduced the inhibition by 3.3 m M AA to 8% of specific binding. The loss of specific binding by AA was rescued by 1 m M EDTA, an inhibitor of lipid peroxidation. In the absence of AA, competition experiments with the agonist, dopamine, revealed the presence of high-affinity ( K _h = 224.9 ± 48.9 n M ) and low-affinity ( K _l = 21,100 ± 2,400 n M ) binding sites. Although the maximum binding of ¹²⁵I-SCH 23982 decreased to 40% without affecting the K _D value in the presence of 1.67 m M AA, the value of the high-affinity site for dopamine was increased ( K _h = 23.3 ± 9.4 n M ) and that of the low-affinity site was decreased ( K _l = 136,800 ± 40,900 n M ). These results suggest that AA may affect D₁ dopamine receptor function by lipid peroxidation, competition with dopamine for low-affinity sites, and reduced oxidation of dopamine.     

Temperature Sensitivity of Agonist High-Affinity Binding Sites of Solubilized and Reconstituted D1 Dopamine Receptors

Abstract: Solubilization of rat striatal membranes with sodium cholate, followed by reconstitution into phospholipid vesicles, leads to a 6.5-fold increase in the agonist high-affinity binding sites of the D₁ dopamine receptor. These high-affinity binding sites display differential sensitivity toward temperature. When reconstituted receptors were preincubated for 1 h at 0–4°C (on ice) or at 22°C (room temperature) followed by radioligand binding assays with dopamine, neither the high-affinity values of the receptor for dopamine nor the percent receptors in the high-affinity state (31–39%) were changed from control reconstituted receptors, which were not subject to any preincubations. At 30°C, there was a partial loss in the number of high-affinity D₁ receptors with only 25% of the total receptor population in the high-affinity state; there was no change in the affinity values of the high-affinity binding sites. At 37°C, there was a 40% loss in total number of D₁ receptor binding sites. All the high-affinity binding sites were lost and the remaining 60% of binding activity represented the low-affinity binding state of the receptor. These results indicate that the high-affinity binding sites of the reconstituted D₁ dopamine receptors are uniquely sensitive to higher temperatures.     

Analysis of N-terminal processing of hepatitis C virus nonstructural protein 2.

We determined the partial amino (N)-terminal amino acid sequence of hepatitis C virus p21 (nonstructural protein 2 [NS2]). Cleavage at the p21 (NS2) N terminus depended on the presence of microsomal membranes. The amino-terminal position of p21 (NS2) was assigned to amino acid 810 of the hepatitis C virus strain IIJ precursor polyprotein. Mutation of the alanine residue at position P1 of the putative cleavage site inhibited membrane-dependent processing. This alteration in processing together with the fact that hydrophobic amino acid residues are clustered upstream of the putative cleavage site suggested the involvement of a signal peptidase(s) in the cleavage. Furthermore, mutation analysis of this possible cleavage site revealed the presence of another microsome membrane-dependent cleavage site upstream of the N terminus of p21 (NS2).     

Inactivation of bleomycin by an N-acetyltransferase in the bleomycin-producing strain Streptomyces verticillus

Abstract Bleomycin-producing Streptomyces verticillus ATCC15003 possesses a bleomycin acetyltransferase which inactivates the drug in the presence of acetyl coenzyme A. The site of acylation in enzymically prepared acetylbleomycin A2 was determined by nuclear magnetic resonance analysis; the primary amino group of the β-aminoalanine moiety of bleomycin was acetylated. Acetylbleomycin A2 had no detectable antibacterial activity and did not induce in vitro DNA degradation.     

Characterization of a Chlamydia pneumoniae Strain Isolated from a 57-Year-Old Man

The isolation of Chlamydia pneumoniae, especially from elderly persons, is generally not easy. Recently, we succeeded in isolating a chlamydial strain, which was designated KKpn-15, from a 57-year-old man suffering from acute bronchitis. It was compared with well established strains of C. pneumoniae, C. trachomatis and C. psittaci, and its biological properties, such as the morphology of elementary bodies (EBs) and inclusions, and the immunochemistry of EB proteins, were investigated. Based on the results obtained in the present study, it was confirmed that the new chlamydial strain, KKpn-15, is a member of the C. pneumoniae strain and that the organisms of KKpn-15 are useful as an antigen for the serodiagnosis and epidemiology of C. pneumoniae infection.