Lipoxygenase, Hydroperoxide Lyase and Volatile C6-Aldehyde Formation from C18-Fatty Acids during Development of Phaseolus vulgaris L.

Kidney bean plants (Phaseolus vulgaris) were found to have thecapability to produce C₆-aldehydes (hexanal and hexenals) fromlinoleic and linolenic acids. The various organs tested hadlipoxygenase and hydroperoxide lyase activities responsiblefor the C₆-aldehyde formation. Young leaves showed relativelyhigh activities for C₆-aldehyde formation. However, the activitiesof the leaves decreased gradually with leaf expansion. Seedlingsand seeds containing cotyledons showed low activities for C₆-aldehydeformation, because of the occurrence of an inhibitory factorin the cotyledons. The substrate specificity of the enzymeswas essentially the same among the various developmental stagesof leaves tested. (Received February 5, 1982; Accepted March 19, 1982)     

Increase of superoxide dismutase activity in various human leukemia cells.

(1) Superoxide dismutase activity in polymorphonuclear cells from human blood is considerably lower than that in lymphocytes. Macrophages from ascites show the middle level between the other two cells. (2) In myelocytic, monocytic, and lymphocytic leukemia cells, the enzyme activities are increased compared to those in the corresponding normal cells. (3) Gel electrophoresis patterns of all normal cells reveal bands corresponding to the cytosol and mitochondrial bands reported in previous studies. However, the mitochondrial Mn-containing superoxide dismutase activities are diminished or absent in leukemia cells. CN-insensitive superoxide dismutase activity in leukemia cells is not detected under the conditions.     

Cells capable of uptake of horseradish peroxidase in some circumventricular organs of the cat and rat

Summary The structure of mesenchymal cells distributed in some of the hypendymal organs of the circumventricular system in the cat and rat was demonstrated after intravenous injection of high doses of horseradish peroxidase. These cellular elements were observed in the vicinity of blood vessels of the organon vasculosum laminae terminalis, subfornical organ and area postrema. Electron-microscopically, these cells located between the basal laminae of the brain parenchyma and the blood capillaries show long cellular processes encircling fenestrated capillaries. Light and electron-microscopic examination revealed that this cell type is identical with the horseradish peroxidase-uptake cells, previously reported in the vicinity of the hypophysial portal system. Such phagocytic cells may be considered as a cellular component intervening between the brain parenchyma and the blood stream, playing a role in selective barrier functions in the above-mentioned circumventricular organs where a blood-brain barrier in the classical sense of the definition is lacking.This work was supported by grant No. 437002 from the Ministry of Education, Science and Culture, Japan     

Effect of leaf age on light and dark 14CO2 fixation in a CAM plant, Bryophyllum calycinum

Leaves of different ages from B. calycinum were exposed to ¹⁴CO₂in light during day and night. The labelling pattern on thechromatogram differed with leaf age. Young leaves had similarpatterns to those of C3 plants during both day and night. Matureleaves showed high incorporation of ¹⁴C into C4 acids, especiallyat night. In contrast, no significant difference with leaf agewas observed in the pattern of dark ¹⁴CO₂ fixation products.Study of the enzyme activity and the content of titratable acidat each leaf age suggested that high incorporation of ¹⁴C inC4 acids during the night was due to the simultaneous absorptionof CO₂ by both enzymes RuDPcarboxylase and PEPcarboxylase. (Received November 24, 1977; )     

Monoclonal antibodies recognizing amino-terminal and carboxy-terminal regions of human aldolase A: probes to detect conformational changes of the enzyme.

Three monoclonal antibodies (MAbs1A2, 3C5, and 4C2) for human aldolase A [EC 4.1.2.13] were established. MAbs1A2, 3C5, and 4C2 were shown to belong to subclasses IgM, IgG1, and IgG2a, respectively. None of the MAbs inhibits aldolase A activity. Their epitopes were mapped in detail on the molecule by examining the reactivities of the MAbs to chimeric proteins between aldolases A and B [Kitajima et al. (1990) J. Biol. Chem. 265, 17493-17498] in ELISA and to the CNBr-cleaved fragments of aldolase A in immuno-blotting. MAbs1A2 and 3C5 reacted with sites located within amino acid residues 306-363 at the C-terminal region of the enzyme. MAb4C2 recognized an epitope of the enzyme present within amino acid residues 34-108 at the N-terminal region. In a competitive binding assay, MAbs1A2 and 3C5 competed with each other for binding to the antigen and also interfered with the binding of MAb4C2, whereas MAb4C2 failed to inhibit the binding of MAbs1A2 and 3C5 to the antigen. MAb3C5 showed a species-specificity in the reaction with the antigen; it reacted with human and rabbit aldolase A with similar reactivity but not at all with the rat and mouse enzymes, which differ from the human and rabbit enzymes in two amino acid residues at positions 328 and 348. Reactivities of MAbs to aldolase A were further examined with engineered enzymes containing an amino acid substitution.(ABSTRACT TRUNCATED AT 250 WORDS)     

Dynamic sweating response of man to infrared irradiation in various spectral regions

In an attempt to detect differences in the thermal effect of infrared irradiation of different wavelengths, transient sweating response to infrared irradiation in various spectral regions was examined. In Series 1, the ventral or dorsal surface of the nude subject was irradiated repetitively for a period of 4 min (2 min on, 2 min off) by each of three kinds of infrared heaters with main emissivity in near-infrared (NIR; 0.7–2.8 m), intermediate-infrared (MIR; 1.5–5.8 m), and far-infrared (FIR; 2.8–25 m) regions. The sweating response on a non-irradiated area tended to be the greatest with MIR, while the magnitude of the sweating response on the irradiated area showed no consistent differences among various wavelengths. The results infer that MIR stimulated cutaneous thomoreceptors most effectively, while its direct effect on local sweat gland activity was minimal. In Series 2, the effects of 9–12 min irradiations in more restricted ranges of wavelength were compared by the combination of the three kinds of heaters with filters (translucent to wavelength ranges of 1.3–2.7, 2.7–3.5, 3.6–8.0 m, respectively). The sweating response on a remote area was predominantly greater with the range of 2.7–3.5 m than with the other wavelength ranges, while the local effect on sweating was minimal with this range. The results of Series 2 reinforce those of Series 1, indicating that the degree of stimulation of cutaneous thermoreceptors and of direct thermal effect on sweat gland activity differ with spectral regions incident on the skin, thus affecting local and remote effects on the sweating response.     

Coagulation factor X activating enzyme from Russell's viper venom (RVV-X). A novel metalloproteinase with disintegrin (platelet aggregation inhibitor)-like and C-type lectin-like domains.

We determined the complete amino acid sequence of RVV-X, the blood coagulation factor X activating enzyme, isolated from Russell's viper venom and studied structure-function relationships. RVV-X (M(r) 79,000) consists of a disulfide-bonded two-chain glycoprotein with a heavy chain of M(r) 59,000 and a light chain of heterogeneous M(r) 18,000 (LC1) and 21,000 (LC2). These chains were separated after reduction and S-pyridylethylation, and the isolated major component LC1 was used for sequence analysis. The heavy chain consists of 427 residues containing four asparagine-linked oligosaccharides, and its entire sequence was similar to that of the high molecular mass hemorrhagic protein, HR1B, isolated from the venom of Trimere-surus flavoviridis. The heavy chain contains three distinct domains, metalloproteinase, disintegrin (platelet aggregation inhibitor)-like and unknown cysteine-rich domains. On the other hand, light chain LC1 consists of 123 amino acid residues containing one asparagine-linked oligosaccharide and shows sequence homology similar to that found in the so-called C-type (Ca(2+)-dependent) lectins. Therefore, RVV-X is a novel metalloproteinase containing a mosaic structure with distintegrin-like, cysteine-rich, and C-type lectin-like domains. RVV-X potently inhibits collagen- and ADP-stimulated platelet aggregations, probably via its distintegrin-like domain, although this domain does not contain the Arg-Gly-Asp sequence which is conserved in various venom distintegrins and which is thought to be one of the interaction sites for platelet integrins. Our findings also indicate that snake venom factor IX/factor X-binding protein with a C-type lectin structure (Atoda, H., Hyuga, M., and Morita, T. (1991) J. Biol. Chem. 266, 14903-14911) inhibits RVV-X-catalyzed factor X activation; hence, the light chain of RVV-X probably participates in recognizing some portion of the zymogen factor X.     

A mitogen-activated protein (MAP) kinase activating factor in mammalian mitogen-stimulated cells is homologous to Xenopus M phase MAP kinase activator.

The mitogen-activated protein (MAP) kinases, a family of 40-45-kDa kinases whose activation requires both tyrosine and threonine/serine phosphorylations, are suggested to play key roles in various phosphorylation cascades. A previous study of Krebs and co-workers (Ahn, N. G., Seger, R., Bratlien, R. L., Diltz, C. D., Tonks, N. K., and Krebs, E. G. (1991) J. Biol. Chem. 266, 4220-4227) detected an activity in epidermal growth factor (EGF)-stimulated 3T3 cells that can stimulate inactive MAP kinases. We observed this activity in rat 3Y1 cells treated with various mitogenic factors and in PC12 cells treated with nerve growth factor (NGF). Its kinetics of activation and deactivation following EGF or NGF stimulation roughly paralleled that of MAP kinase. The MAP kinase activator required the presence of ATP and a divalent cation such as Mn2+ and Mg2+ and was inactivated by phosphatase 2A treatment in vitro. This activator has been isolated from EGF-stimulated 3Y1 cells by sequential chromatography and identified as a 45-kDa monomeric protein. It was able to activate mammalian and Xenopus MAP kinases in vitro and was very similar to Xenopus M phase MAP kinase activating factor, which was purified previously from mature oocytes (Matsuda, S., Kosako, H., Takenaka, K., Moriyama, K., Sakai, H., Akiyama, T., Gotoh, Y., and Nishida, E. (1992) EMBO J. 11, 973-982), in terms of its functional, immunological, and physicochemical properties. Thus, the same or a similar upstream activating factor may function in mitogen-induced and M phase-promoting factor-induced MAP kinase activation pathways.     

Activation of mitogen-activated protein kinase and its activator by ras in intact cells and in a cell-free system.

Mitogen-activated protein (MAP) kinase is a serine/threonine kinase whose function is thought to be essential for the transduction of mitogenic signals. MAP kinase is activated by phosphorylation induced by a variety of extracellular stimuli, and its direct upstream activator has been identified. Using amphibian and mammalian systems, we show here that ras can activate MAP kinase and its activator. Injection of v-Ha-ras p21 into Xenopus immature oocytes activated both MAP kinase and maturation-promoting factor (MPF) activities. The activation of MAP kinase preceded that of MPF, demonstrating that ras activates MAP kinase in an MPF-independent pathway. Moreover, we found that the MAP kinase activator is also activated in ras-injected oocytes. Activation of MAP kinase and its activator occurred also when the v-Ki-ras gene was conditionally induced in rat fibroblastic 3Y1 cells. Furthermore, we observed that ras activated MAP kinase and its activator in a cell-free system prepared from Xenopus oocytes. Using an antibody against the Xenopus 45-kDa MAP kinase activator, we demonstrated that the 45-kDa activator molecule was activated by ras. These findings suggest that the MAP kinase activator/MAP kinase system may be the downstream components of ras signal transduction pathways.