Fatty acid composition of brown adipose tissue in genetically heat-tolerant FOK rats

The phospholipid fatty acid composition of brown adipose tissue (BAT) was examined in inbred heat-tolerant FOK rats and compared with that in conventional Wistar rats not previously exposed to heat. The FOK rats showed higher unsaturation states, as indicated by higher levels of polyunsaturated fatty acids and a higher unsaturation index and polyunsaturated fatty acids/saturated fatty acids ratio. This higher level of unsaturation was characterized by the higher amount of polyunsaturated fatty acids such as linoleic acid, arachidonic acid and docosahexaenoic acid. It may be concluded that the increased docosahexaenoic acid level in BAT phospholipids brings about the hyperplasia of BAT, causing an enhancement of its in vivo thermogernic activity as well as the systemic non-shivering thermogenesis observed in heat-tolerant FOK rats. Received: 17 May 2000 / Revised: 21 August 2000 / Accepted: 21 September 2000     

Leclercia adecarboxylata Gen. Nov., Comb. Nov., formerly known asEscherichia adecarboxylata.

The nameLeclercia adecarboxylata is proposed for a group of the family Enterobacteriacae previously known asEscherichia adecarboxylata. Leclercia adecarboxylata can be phenotypically differentiated from all other species of Enterobacteriaceae. The members of this species are positive for motility, indole production, methyl red, growth in the presence of KCN, malonate, beta-galactosidase, beta-xylosidase, esculin hydrolysis, gas production fromd-glucose, and acid production fromd-cellobiose,d-lactose, melibiose,l-rhamnose, adonitol,d-arabitol, dulcitol, and salicin; the strains were negative for Voges-Proskauer, citrate (Simmons), H₂S (Kligler), lysine and ornithine decarboxylases, arginine dihydrolase, phenylalanine deaminase, gelatinase, DNase, Tween-80 hydrolysis, and acid production from myoinositol and alpha-methyl-d-glucoside. Fermentation ofd-raffinose,d-sucrose, andd-sorbitol is variable with strains. DNA relatedness of 11 strains ofL. adecarboxylata to three strains including the type strain of this species averaged 80% in reactions at 65°C. DNA relatedness to other species in Enterobacteriaceae was 2%–32%, indicating that this species was placed in a new genusLeclercia gen. nov. The type strain ofL. adecarboxylata is ATCC 23216.     

Ca<Superscript>2+</Superscript>, calmodulin and phospholipids regulate nitricoxide synthase activity in the rabbit submandibular gland

Nitric oxide (NO) plays an important role as an intra- and intercellular signaling molecule in mammalian tissues. In the submandibular gland, NO has been suggested to be involved in the regulation of secretion and in blood flow. NO is produced by activation of NO synthase (NOS). Here, we have investigated the regulation of NOS activity in the rabbit submandibular gland. NOS activity was detected in both the cytosolic and membrane fractions. Characteristics of NOS in the cytosolic and partially purified membrane fractions, such as Km values for l-arginine and EC₅₀ values for calmodulin and Ca²⁺, were similar. A protein band that cross-reacted with anti-nNOS antibody was detected in both the cytosolic and membrane fractions. The membrane-fraction NOS activity increased 1.82-fold with treatment of Triton X-100, but the cytosolic-fraction NOS activity did not. The NOS activity was inhibited by phosphatidic acid (PA) and phosphatidylinositol 4,5-bisphosphate (PIP₂). The inhibitory effects of phospholipids on the NOS activity were relieved by an increase in Ca²⁺ concentrations. These results suggest that the Ca²⁺- and calmodulin-regulating enzyme nNOS occurs in cytosolic and membrane fractions, and PA and PIP₂ regulate the NOS activity in the membrane site by regulating the effect of Ca²⁺ in the rabbit submandibular gland.Communicated by I.D. Hume     

Hypoxia reduces the expression of heme oxygenase-2 in various types of human cell lines. A possible strategy for the maintenance of intracellular heme level

Heme oxygenase consists of two structurally related isozymes, heme oxygenase-1 and and heme oxygenase-2, each of which cleaves heme to form biliverdin, iron and carbon monoxide. Expression of heme oxygenase-1 is increased or decreased depending on cellular microenvironments, whereas little is known about the regulation of heme oxygenase-2 expression. Here we show that hypoxia (1% oxygen) reduces the expression levels of heme oxygenase-2 mRNA and protein after 48 h of incubation in human cell lines, including Jurkat T-lymphocytes, YN-1 and K562 erythroleukemia, HeLa cervical cancer, and HepG2 hepatoma, as judged by northern blot and western blot analyses. In contrast, the expression level of heme oxygenase-1 mRNA varies under hypoxia, depending on the cell line; it was increased in YN-1 cells, decreased in HeLa and HepG2 cells, and remained undetectable in Jurkat and K562 cells. Moreover, heme oxygenase-1 protein was decreased in YN-1 cells under the conditions used, despite the induction of heme oxygenase-1 mRNA under hypoxia. The heme oxygenase activity was significantly decreased in YN-1, K562 and HepG2 cells after 48 h of hypoxia. To explore the mechanism for the hypoxia-mediated reduction of heme oxygenase-2 expression, we showed that hypoxia shortened the half-life of heme oxygenase-2 mRNA (from 12 h to 6 h) in YN-1 cells, without affecting the half-life of heme oxygenase-1 mRNA (9.5 h). Importantly, the heme contents were increased in YN-1, HepG2 and HeLa cells after 48 h of incubation under hypoxia. Thus, the reduced expression of heme oxygenase-2 may represent an important adaptation to hypoxia in certain cell types, which may contribute to the maintenance of the intracellular heme level.     

Quantum Yields of Decomposition and Homo-Dimerization of Solid <Emphasis Type="SmallCaps">L</Emphasis>-Alanine Induced by 7.2 eV Vacuum Ultraviolet Light Irradiation: An Estimate of the Half-Life of <Emphasis Type="SmallCaps">L</Emphasis>-Alanine on the Surface of Space Objects

One of the leading hypotheses regarding the origin of prebiotic molecules on primitive Earth is that they formed from inorganic molecules in extraterrestrial environments and were delivered by meteorites, space dust and comets. To evaluate the availability of extraterrestrial amino acids, it is necessary to examine their decomposition and oligomerization rates as induced by extraterrestrial energy sources, such as vacuum ultraviolet (VUV) and X-ray photons and high energy particles. This paper reports the quantum yields of decomposition ((8.2 ± 0.7) × 10⁻² photon⁻¹) and homo-dimerization ((1.2 ± 0.3) × 10⁻³ photon⁻¹) and decomposition of the dimer (0.24 ± 0.06 photon⁻¹) of solid l-alanine (Ala) induced by VUV light with an energy of 7.2 eV. Using these quantum yields, the half-life of l-Ala on the surface of a space object in the present earth orbit was estimated to be about 52 days, even when only photons with an energy of 7.2 eV emitted from the present Sun were considered. The actual half-life of solid l-Ala on the surface of a space object orbit around the present day Earth would certainly be much shorter than our estimate, because of the added effect of photons and particles of other energies. Thus, we propose that l-Ala needs to be shielded from solar VUV in protected environments, such as the interior of a meteorite, within a time scale of days after synthesis to ensure its arrival on the primitive Earth.     

An engineered hyaluronan synthase: characterization for recombinant human hyaluronan synthase 2 Escherichia coli

The Class I hyaluronan synthase (HAS) is a unique glycosyltransferase synthesizing hyaluronan (HA), a polysaccharide composed of GlcUA and GlcNAc, by using one catalytic domain that elongates two different monosaccharides. As for the synthetic mechanism, there are two alternative manners for the sugar elongation process. Some bacterial HASs add new sugars to the non-reducing end of the acceptor to grow polymers. On the other hand, some vertebrate enzymes seem to transfer sugars to the reducing end. Expression of vertebrate HASs as active and soluble proteins will accelerate further precise insight into mechanisms of sugar elongation reactions by natural HASs. Since large scale production of HA polymers and oligomers would become powerful tools both for basic studies and new biotechnology to create functional carbohydrates in medicinal purposes, advent of an efficient method for the expression of HASs in Escherichia coli is strongly expected. Here we communicate the first success of the production of recombinant human HAS2 proteins composed of only the catalytic region in E. coli as the active form. It was demonstrated that an engineered HAS2 expressed in E. coli exhibited significant activity to synthesize a mixture of HAS oligomers from 8-mer (HA8) to 16-mer (HA16). Engineered HAS2 prepared herein elongated sugars from exogenous tetrasaccharide to form polymers with a direction to the non-reducing end. According to the present results, large scale production of engineered recombinant HASs is to be performed using E. coli that will provide practical and economic advantages in manufacturing enzymes for use in the synthesis of various oligomeric HA molecules and their industrial applications.     

Semaphorin 4C, a transmembrane semaphorin, [corrected] associates with a neurite-outgrowth-related protein, SFAP75

Semaphorin 4C (S4C, previously called M-SemaF) was recently identified as a brain rich transmembrane member of semaphorin family of the vertebrate. In the cytoplasmic domain of S4C there is a proline-rich region suggesting that the cytoplasmic domain may play an important role in Sema4C function. In this study, we have identified the cytoplasmic domain (cd) of M-SemaF(S4C)-associating protein with a Mr of 75 kDa, named SFAP75, from mouse brain. SFAP75 turned out to be the same as the recently reported neurite-outgrowth-related protein named Norbin. Deletion mutants analyses of S4C and SFAP75 revealed that the membrane-proximal region of S4Ccd binds to the intermediate region of SFAP75. Western blot and immunohistochemical analyses with anti-Sema4C and anti-SFAP75 antibodies indicated that S4C and SFAP75 were specially enriched in the brain with a similar distribution pattern to each other. These results suggest that S4C interacts with SFAP75 and plays a role in neural function in brain.