The Escherichia coli cytochrome b556 gene, cybA, is assignable as sdhC in the succinate dehydrogenase gene cluster

Abstract The cytochrome b556 -deficient mutant Escherichia coli K12 strain TK3D11 [7] could not grow with succinate as the sole carbon source, but could grow well on dl -lactate. This finding suggested that cytochrome b556 is primarily responsible for oxidative metabolism and utilization of succinate. 24 Amino acid residues at the amino-terminal of purified cytochrome b556 were determined. This sequence coincided completely with amino acid residues 4 to 27, predicted from the DNA sequence of the sdhC gene, one of the unassigned open reading frames of the sdh gene cluster recently reported by Wood et al. [16]. Based on these and other results, we concluded that cybA , the gene for cytochrome b556 , is assignable as sdhC .     

Characterization of a growth-inhibiting protein present in rat serum that exerts a differential effect onin vitro growth of nonmalignant rat liver cells when compared with Rous sarcoma virus-transformed rat liver cells

Summary We have previously reported the transformation by Rous sarcoma virus of a cloned epithelial cell line (BRL) established from Buffalo rat liver by H. Coon. The nontransformed (BRL) and transformed (RSV-BRL) cells grew at comparable rates in culture, whereas only the transformed cells were tumorigenic in vivo. We report here on the existence in rat and mouse sera of a growth inhibitor for the nontransformed BRL cells. The transformed BRL cells (RSV-BRL) were insensitive to this inhibitor. The inhibitory activity was not prominent in sera from other species of animals tested except for rabbit; this serum inhibited the growth of RSV-BRL cells more strongly than that of BRL cells. The growth inhibitor was partially purified from rat serum. It is a protein free of lipid and has a molecular weight of about 220 000. The inhibitor could be separated into three components of pI 4.6, 5.2 (major) and 5.6 by isoelectric electrophoresis. EDITOR'S STATEMENT Although compelling theoretical arguments sometimes can be made for the likely existence of growth-inhibitory substances of physical relevance in the control of cell proliferation, experiments aimed at identifying and studying such factors often are difficult to design and interpret, and little strong data exists to suggest that growth-inhibitory substances are important regulatorsin vivo. The information presented in this paper represents a start toward developing a useful system for studying growth-inhibitory factor. David W. Barnes     

The morphology and anatomy ofHydrastis (Ranunculales): Systematic reevaluation of the genus

Evidence from morphology and anatomy (including embryology), as well as from palynology, chemistry and cytology, indicates thatHydrastis is quite divergent from Ranunculaceae (in which the genus has been most often included) as well as from both Glaucidiaceae and Berberidaceae. Distinctive features ofHydrastis, which demarcate it from Ranunculaceae but which are sometimes shared by Berberidaceae, are: the unique mode of origin of the vascular supply to stamens and carpels; the micropyle being formed by both integuments; the xylem not V-shaped in cross section; scalariform vessel perforations present; haploid chromosome number 13; pollen tectum consisting of a compound layer of striae; leaf mesophyll not differentiated; the unique course of stem medullary bundles; D-galactose present. Its distinctive higher haploid chromosome number, as well as its many less-specialized character states (in floral structure, leaf anatomy, and xylem and vessel morphology), suggest thatHydrastis is a relictual primitive group which diverged early from a common ancestral stock of Ranunculaceae, Berberidaceae and probably of Circaeasteraceae; at least some of the features shared byHydrastis and one or another of the families concerned seem to be a heritage from their common ancestor. We propose a reestablishment of a monotypic family, Hydrastidaceae.     

In vitro characteristics of rat mesangial cells in comparison with aortic smooth muscle cells and dermal fibroblasts

Rat glomerular mesangial cells were cultured and their antigens were compared with those of aortic vascular smooth muscle cells and dermal fibroblasts. Glomeruli, aortic, and dermal explants were cultured for 3 weeks and subcultured in the same conditions. These cultured cells were evaluated by indirect immunofluorescence studies using antibodies against Thy-1 antigen, desmin, and chicken gizzard actin. Most of mesangial cells were positive for Thy-1, desmin, and actin. On the other hand, fibroblasts were negative for desmin, and smooth muscle cells stained Thy-1 scarcely, and were negative for desmin. In the latter two cells, actin-positive fibrils were thinner and fainter than mesangial cells. These results indicated that mesangial cells could be distinguished in vitro from vascular smooth cells and fibroblasts by immunofluorescence microscopy.     

Spontaneous Mutations Modifying the Activity of Alcohol Dehydrogenase (Adh) in DROSOPHILA MELANOGASTER

In a marked-inversion balanced lethal system of the second chromosome of Drosophila melanogaster, mutations were accumulated under minimum pressure of natural selection in 1000 individual lines that originated essentially from two individuals. After about 300 generations, the specific activities of alcohol dehydrogenase of 69 randomly selected individual lines were measured with replications using four replicated vials (on 2 days—two replications per day) by observing the reduction of NAD⁺ to NADH at 340 nm. Total soluble protein as the basis of standardization of enzyme activity was measured by the Lowry method for each vial. A control experiment was made immediately after the establishment of 20 individual lines from a single genotype. A significant increase in genetic variance was observed among the mutation-accumulating lines but was not detected in the control experiment. The statistical analysis of the data on the basis of the one-band/one-gene hypothesis suggests that many mutations controlling the activity of alcohol dehydrogenase occurred in regions different from the alcohol dehydrogenase locus itself, mainly in the noncoding DNA. Furthermore, it is suggested that transposon-like elements are related to the induction of these changes in alcohol dehydrogenase specific activities. Additional experimental evidence supporting this conclusion is also given.     

Distribution of Leu 7 (HNK-1) antigen in human digestive organs: an immunohistochemical study with monoclonal antibody

Summary Leu 7 immunoreactivity was demonstrated with the indirect peroxidase-labelled antibody method on frozen and paraffin-embedded tissue sections of human digestive organs. Anti-Leu 7 monoclonal antibody, which allegedly detects mononuclear cells with natural killer or killer activity, recognized lymphoid cells among intestinal epithelial cells and in the germinal centres of solitary lymphoid follicles of small and large intestine, and a few in gallbladder, liver and the lamina propria of the intestine. In addition, peripheral nerve fibres, endocrine cells in the gut and pancreas and carcinoid and islet cell tumours were also positively stained. At the ultrastructural level, Leu 7 antigen was localized on the plasma membrane of granulated lymphoid cells in the gut mucosa and on the secretory granules of intestinal endocrine cells. In normal pancreas, Leu 7 immunoreactivity was demonstrated in most cells containing pancreatic polypeptide and in many cells containing somatostatin or glicentin. Insulin-containing cells, however, lacked Leu 7 immunoreactant. These findings were obtained in both frozen sections and paraffin-embedded sections. The possible cross-reactivities of monoclonal antibodies are discussed as they raise an important caveat in immunohistochemical studies using these antibodies.     

Structure and antitumor activity of an alkali-soluble polysaccharide from Cordyceps ophioglossoides

A water-insoluble extracellular glucan (CO-1) was isolated from the precipitate formed on incubation of the culture filtrate of Cordyceps ophioglossoides at 37 degrees for 19 h. CO-1 was homogeneous as judged by h.p.l.c., electrophoresis, and ultracentrifugation, and the average molecular weight was determined by h.p.l.c. to be 632,000. The 1H- and 13C-n.m.r. and the i.r. spectra indicated that the glucosidic linkages in CO-1 were beta. From the results of methylation analysis, Smith degradation, n.m.r. studies, and enzymic hydrolysis, it was concluded that CO-1 is composed of a backbone of (1----3)-linked beta-D-glucopyranosyl residues with a beta-D-glucopyranosyl group attached to O-6 of every second D-glucopyranosyl residue of the backbone. CO-1 strongly inhibited the growth of Sarcoma 180 solid-type tumor. CO-1 polyalcohol, which was prepared by Smith degradation of CO-1, exhibited more-potent antitumor activity than CO-1. The absorption maximum of Congo Red shifted significantly in the presence of CO-1.     

Differences in Ca2+ mobilization induced by alpha-adrenergic agonist and phosphatidic acid in cultured hepatocytes

In an attempt to elucidate the relationship between phosphatidylinositol breakdown and alpha-adrenergic responses, effects of phosphatidic acid and phosphatidylinositol related metabolites on Ca²⁺ mobilization and glucose output in cultured hepatocytes were examined. Norepinephrine induced the net ⁴⁵Ca²⁺ efflux from preloaded cells and stimulated glucose output via alpha-adrenergic receptor stimulation, whereas phosphatidic acid caused ⁴⁵Ca²⁺ uptake to cells and did not stimulate glucose output. Myo-inositol-monophosphate, diglyceride and arachidonic acid, which are released by phosphatidylinositol breakdown, had no effect on ⁴⁵Ca²⁺ efflux and glucose output in cells. These results suggest that phosphatidic acid and phosphatidylinositol related metabolites can not mimic the alpha-adrenergic actions in cultured hepatocytes.     

Chromosomal location of the Escherichia coli cytochrome b556 gene, cybA

Summary The amounts of cytochrome b556 in the cytoplasmic membranes of several Escherichia coli K12 strains having F-prime factors and a lambda transducing phage were determined. The amount was amplified about two-fold in strains having F100-12 and F152, but not in strains having F100-11, F8 and psu ⁺ 2glnS ⁺. The strain TK3D11, which lacks the kdp-gltA region (deletion D-01) of the E. coli chromosome, did not synthesize cytochrome b556 at all. From these results, the gene cybA encoding cytochrome b556 was located in the kdp-gltA region.In the cytochrome b556-deficient mutant, a novel b type cytochrome, cytochrome b561 which is a product of the gene cybB, was identified. It seems to function as a physiological electron transferring cytochrome in place of cytochrome b556 in this mutant.Abbeviations HPLC high performance liquid chromatography - EDTA ethylenediamine tetraacetic acid - SDS sodium dodecyl sulfate - NADH reduced form of nicotinamide adenine dinucleotide