Identification of a major GTP-binding protein in bovine aortic smooth muscle membranes as smg p21, a GTP-binding protein having the same effector domain as ras p21s

At least two GTP-binding proteins (G proteins) with Mr values of about 20,000 were extracted from bovine aortic smooth muscle membranes by sodium cholate. The most abundant G protein (22K G) was purified to near homogeneity by successive column chromatographies of Ultrogel AcA-44, phenyl-Sepharose CL-4B, hydroxyapatite and Mono Q HR5/5. 22K G showed kinetic and physical properties very similar to those of smg p21, a G protein recently isolated from bovine brain and human platelet membranes, having the same effector domain as ras p21s. Moreover, 22K G was recognized specifically by the anti-smg p21 antibody. These results indicate that the major G protein in bovine aortic smooth muscle membranes is smg p21.     

Regeneration of bovine and octopus opsins in situ with natural and artificial retinals

We consider the problem of color regulation in visual pigments for both bovine rhodopsin (lambda max = 500 nm) and octopus rhodopsin (lambda max = 475 nm). Both pigments have 11-cis-retinal (lambda max = 379 nm, in ethanol) as their chromophore. These rhodopsins were bleached in their native membranes, and the opsins were regenerated with natural and artificial chromophores. Both bovine and octopus opsins were regenerated with the 9-cis- and 11-cis-retinal isomers, but the octopus opsin was additionally regenerated with the 13-cis and all-trans isomers. Titration of the octopus opsin with 11-cis-retinal gave an extinction coefficient for octopus rhodopsin of 27,000 +/- 3000 M-1 cm-1 at 475 nm. The absorption maxima of bovine artificial pigments formed by regenerating opsin with the 11-cis dihydro series of chromophores support a color regulation model for bovine rhodopsin in which the chromophore-binding site of the protein has two negative charges: one directly hydrogen bonded to the Schiff base nitrogen and another near carbon-13. Formation of octopus artificial pigments with both all-trans and 11-cis dihydro chromophores leads to a similar model for octopus rhodopsin and metarhodopsin: there are two negative charges in the chromophore-binding site, one directly hydrogen bonded to the Schiff base nitrogen and a second near carbon-13. The interaction of this second charge with the chromophore in octopus rhodopsin is weaker than in bovine, while in metarhodopsin it is as strong as in bovine.     

DNA ploidy pattern and amplification of ERBB and ERBB2 genes in human gastric carcinomas

The DNA ploidy pattern and amplification of ERBB and ERBB2 genes were examined in paraffin-embedded tissue from gastric carcinomas using flow cytometry and a slot-blot hybridization technique. The incidence of aneuploidy in well differentiated adenocarcinomas (56%) was significantly higher (p less than 0.05) than that in poorly differentiated adenocarcinomas (21%). The DNA ploidy pattern was not remarkably different between the primary tumors and metastatic deposits in lymph nodes. Of the nine specimens having an aneuploid stem cell line in the primary tumor and/or in metastases, three showed ERBB2 gene amplification and one showed ERBB gene amplification. The incidence of epidermal growth factor (EGF) immunoreactivity in tumor cells showed no difference between diploid and aneuploid tumors. These findings indicate that aneuploidy is frequently associated with amplification of ERBB and ERBB2 genes.     

Roles of the complement receptor type 1 (CR1) and type 3 (CR3) on phagocytosis and subsequent phagosome-lysosome fusion in Salmonella-infected murine macrophages

Abstract The receptors involved in the recognition of Salmonella typhimurium and S. typhi by murine macrophages were identified, and their relevance to phagosome-lysosome fusion was also investigated. Phagocytosis of S. typhimurium by murine macrophages was dependent on the opsonization with normal fresh serum, although the opsonin had no triggering activity in phagosome-lysosome fusion. In contrast, the opsonization of S. typhi with normal fresh serum efficiently triggered both phagocytosis and following phagosome-lysosome fusion. Anti-murine CR1 antibody suppressed phagocytosis of S. typhimurium by 36%, whereas anti-CR3 antibody, mannan, and advanced glycosylation endproducts (AGE)-BSA all failed to prevent phagocytosis of S. typhimurium , suggesting that CR1 may only contribute to the recognition of S. typhimurium and may possibly play a minor role. Other receptors involved may also influence the outcome phagocytosis in terms of phagosome-lysosome fusion. In the case of S. typhi , only anti-CR3 antibody significantly inhibited not only phagocytosis of S. typhi but also following phagosome-lysosome fusion. Treatment with K76COONa, an inhibitor of C3bINA (I factor), resulted in a marked inhibition of phagosomelysosome fusion in S. typhi -infected macrophages, although no significant inhibition was observed on phagocytosis of S. typhi . These results suggest that S. typhimurium and S. typhi may be recognized at least in part by CR1 and CR3, respectively, and that the recognition by CR3 but not CR1 is functionally associated with subsequent phagosomelysosome fusion in murine macrophages.     

Cloning and sequencing of a ligninase gene from a lignindegrading basidiomycete,Phanerochaete chrysosporium

Summary A ligninase gene has been cloned from a Phanerochaete chrysosporium genomic DNA library. Nucleotide sequencing of the gene has revealed that the ligninase structural gene contains 1116 bp of the protein-encoding sequence, of which 84 bp encode the signal peptide. The protein-encoding sequence is interrupted by eight introns which conform to the universal G-T/A-G splicing rule observed for the 3 and 5 intron boundaries. The putative eukaryotic regulatory sequences, i.e. CAAT and TATA box-like sequences, are present in the 5 flanking region.     

Suppressive effect of ipriflavone on bone depletion in the experimental diabetic rat: dose response of ipriflavone

The dose dependent effect of ipriflavone (7-isopropoxy-isoflavone) on the femoral bone in streptozotocin-induced diabetic rats was studied by microdensitometric analysis. Diabetic rats showed severe hyperglycemia, glucosuria, hypoinsulinemia, associated with increased urinary calcium and hydroxyproline. Microdensitometric analysis revealed decreases in femoral length, bone width, and bone density. The dietary administration of ipriflavone (about 270 mg/kg/day) to the diabetic rats for 6 weeks prevented reduction of the cortical thickness index in the diaphysis and depletion of bone density in the distal metaphysis, and also reduced the inner diameter of the diaphysis; diabetic state was not improved. A simple correlation and linear regression analysis revealed that ipriflavone also significantly reduced the inner diameter in the diaphysis at a dose of 90 mg/kg/day, but not at one of 25 mg/kg/day. These results suggest that ipriflavone suppresses the depletion of the femoral bone through inhibition of bone resorption in a dose dependent fashion; its minimum effective dose is 90 mg/kg/day in experimental diabetes.     

Selection of an atrazine-resistant tobacco cell line having a mutant psbA gene

Summary A mutant cell line that shows high resistance to the photosynthesis-inhibiting herbicide atrazine was selected from cultured photomixotrophic Nicotiana tabacum cv. Samsun NN cells by repeated exposure to toxic levels of the herbicide. This resistance was confirmed by measurements of Hill reaction activity in isolated thylakoid membranes. Nucleotide sequencing revealed that the resistant cell line had a point mutation in its chloroplast psbA gene. The 264th codon, AGT (serine) was changed to ACT (threonine) in this mutant. This new type of mutation also conferred moderate cross-resistance to diuron and subsequently was stable in the absence of continued selection pressure.     

Structural characterization of lipid A component of Erwinia carotovora lipopolysaccharide

The chemical structure of the lipid A component of lipopolysaccharide excreted into the liquid medium by the plant pathogenic enterobacterium Erwinia carotovora FERM P-7576 was characterized. It consists of a -1, 6-linked glucosamine disaccharide which carries ester-and amide-bound fatty acids and phosphate similar to the lipid A from other gram-negative bacteria. The lipid A preparation was not uniform in the number and composition of the fatty acids linked to the disaccharide. Four prominent lipids A were involved, they were composed of five to seven residues of fatty acid. Among them the major component was hexa-acyl lipid A, in which the hydroxyl group at position 3 and the amino group of the non-reducing glucosamine unit carry 3-dodecanoyl-oxytetradecanoyl residues. Positions 2 and 3 of the reducing glucosamine unit were substituted by 3-hydroxytetradecanoic acid. In the hepta-acyl lipid A, an additional hexadecanoic acid was linked to the hydroxyl group of the 3-hydroxytetradecanoyl residue at position 2 of the hexa-acyl lipid A. Two penta-acyl lipids A were the homologs of the hexa-acyl lipid A with decreasing acylation. Dodecanoic acid was missing from one, and 3-hydroxytetradecanoic acid from another. 3-Dodecanoyloxytetradecanoyl residue at position 3 differentiates E. carotovora lipid A from that of other gram-negative bacteria.Abbreviations LPS lipopolysaccharide - GlcN glucosamine - KDO 3-deoxy-d-manno-octulosonic acid - FAB-MS fast atom bombardment mass spectrometry - u atomic mass unit     

Changes in plant growth processes under microgravity conditions simulated by a three-dimensional clinostat

We developed a three-dimensional (3-D) clinostat to simulate a microgravity environment and studied the changes in plant growth processes under this condition. The rate of germination of cress (Lepidium sativum), maize (Zea mays), rice (Oryza sativa), pea (Pisum sativum), or azuki bean (Vigna angularis) was not affected on the clinostat. The clinostat rotation did not influence the growth rate of their roots or shoots, except for a slight promotion of growth in azuki roots and epicotyls. On the contrary, the direction of growth of plant organs clearly changed on the 3-D clinostat. On the surface of the earth, roots grow downward while shoots upward in parallel to the gravity vector. On the 3-D clinostat, roots of cress elongated along the direction of the tip of root primordia after having changed the direction continuously. Rice roots also grew parallel to the direction of the tip of root primordia. On the other hand, roots of maize, pea, and azuki bean grew in a random fashion. The direction of growth of shoots was more controlled even on the 3-D clinostat. In a front view of embryos, shoots grew mostly along the direction of the tip of primordia. In a side view, rice coleoptiles showed an adaxial (toward the caryopsis) while coleoptiles of maize and epicotyls of pea and azuki bean an abaxial curvature. The curvature of shoots became larger with their growth. Such an autotropism may have an important role in regulation of life cycle of higher plants under a microgravity environment.