Structure of human phenylethanolamine n-methyltransferase gene: existence of two types of mRNA with different transcription initiation sites

The chromosomal gene for human phenylethanolamine N-methyltransferase (PNMT; EC 2.1.1.28) was isolated from a human genomic library using a cloned human PNMT cDNA as a probe, and the nucleotide sequence was determined. PNMT is encoded in a single gene which consists of three exons. We observed newly the presence of minor PNMT mRNA (type B) besides the major mRNA (type A) as reported previously (Kaneda et al., J. Biol. Chem. 263, 7672–7677, 1988) by Northern hybridization. Type B mRNA carries an approximately 700 nucleotide-long untranslated region in the 5′ terminus. This suggests that two types of mRNA are produced from a single gene through the use of two alternative promoters. A TATA-like sequence locates 30 base pair upstream from the cap site of type A mRNA. Upstream of the cap site, there are several sequences resembling Spl binding sites and glucocorticoid responsive elements, with the latter also found in the first intron.     

Nucleotide sequence of Suncus murinus immunoglobulin mu gene and comparison with mouse and human mu genes

RNA polymerase from the archaebacterium Sulfolobus acidocaldarius was chemically modified with AMP o-formylphenyl ester followed by reduction with borohydride. The modified protein catalyzes the labeling of its own largest subunit when incubated with [-³³P]UTP in the presence of poly[d(A-T)]. On cleaving of the labeled protein using cyanogen bromide, hydroxylamine or amino acid-specific endoproteinases for a very brief period, the pattern and size of the radioactive fragments formed are best explained by attachment of the label between Gly⁸⁴³ and Met⁸⁹⁵ of the largest subunit. In this region there exists a highly conserved sequence which is also found in other archaebacterial, eukaryotic and prokaryotic RNA polymerases. This suggests that the binding site for the initiating substrate of RNA polymerases has been conserved during evolution.     

Role of cell-mediated immunity in the resistance to experimental sporotrichosis in mice

The in vitro activity of several new imidazoles, cloconazole, sulconazole, butoconazole, isoconazole and fenticonazole, were compared with those of amphothericin B, flucytosine, and three azoles: econazole, miconazole and ketoconazole against isolates of pathogenic Candida. A total of 186 clinical isolates of 10 species of the genus Candida and two culture collection strains were tested by an agar-dilution technique. Isoconazole was the most active azole, followed by butoconazole and sulconazole. Differences between some of the species in their susceptibility to the antifungal agents were noted. Sulconazole and cloconazole had the highest activity in vitro against 106 isolates of C. albicans. Butoconazole and isoconazole were also very active against isolates of C. albicans, and were the most active azole compounds against 80 isolates of Candida spp.     

Induction of Cytoplasmic Streaming and Movement of Chloroplast Induced by L-Histidine and its Derivatives in Leaves of Egeria densa

Rotational cytoplasmic streaming in leaves of Egeria densa wasinduced by light as well as by L-histidine (L-His). During bothtreatement with light and with L-His chloroplasts on the periclinalface were dislodged and moved to the anticlinal face where rotationalcytoplasmic streaming occurred. The effective concentrationof L-His was about 0.01 mM and the effect was almost saturatedat 0.1 mM. A derivative of L-His, 3-methyl-L-histidine, wasslightly less effective than L-His. By contrast, 1-methyl-L-histidinewas almost ineffective for induction of streaming, not onlyin Egeria but also in Vallisneria. Our resutlts are in markedcontrast to Fitting's result (1936) that 1-M-L-His is more effectivethan L-His. In Egeria, 1-methyl-L-His counteracted the stimulativeeffect of L-His. 1-Methyl-L-His penetrated into leaf cells ofEgeria to the same extent as 3-methyl-L-His and to a greaterextent than L-His. This observation excludes the possibilitythat the impermeability of leaves to 1-M-L-His might be responsiblefor its ineffectiveness. 1-M-L-His did not interfere with photodinesis.Differences in the mechanism of induction of rotational streamingby L-His and by light are discussed. ⁴ Present address: Fukui Institute of Technology, Gakuen, Fukui,910 Japan (Received July 16, 1990; Accepted December 20, 1990)     

Expression of four types of human tyrosine hydroxylase in COS cells

Alternative splicing from a single gene produces four kinds of human tyrosine hydroxylase (types 1-4), which have structural diversity only in the N-terminal region. We attempted expression of the type 1-4 enzymes in COS cells and performed kinetic analyses. All had enzymatic activities. The Km values of the four types for L-tyrosine and 6-methyl-5,6,7,8-tetrahydropteridine were similar, although their relative homospecific activities were clearly different. The type 1 enzyme displayed the highest activity.     

Quantitative measurement of endotoxin in rainbow trout (Salmo gairdneri) serum by the chromogenic substrate method

The amount of endotoxin in serum collected from normal rainbow trout ( Salmo gairdneri) and trout inoculated with viable Vibrio anguillarum or lipopolysaccharide (LPS) extracted from bacteria was determined by the chromogenic substrate method. The mean values of endotoxin in four different groups of normal rainbow trout sera ranged from 31.9 to 65.3 pg/ml. When fish were inoculated with viable bacteria (1 × 10⁸), they became septicaemic and a large amount of endotoxin (> 14 ng/ml) was detected in the sera. In fish inoculated with a smaller number of bacteria the amount of endotoxin was several times higher than that of normal fish in spite of failure of bacterial isolation. Although the endotoxin level in serum increased rapidly (> 100 ng/ml) after intraperitoneal inoculation with purified V. anguillarum LPS (540 μg), no fish died during the experiment. The high level of endotoxin in normal rainbow trout and the resistance of trout to endotoxin are in striking contrast to those of mammalian and avian species.     

DETERMINATION OF GUSTATORY SUCROSE THRESHOLD IN WATER BY USE OF A LINEAR SUCROSE GRADIENT

A novel experimental method was developed which allows the determination of the threshold concentration of sucrose by use of a linear sucrose gradient in water. With this method a continuous tasting of the test-liquid is possible. A panel of 15 persons experienced in taste-testing was used. Three gradients of different steepness were applied: 0 to 1.5% (w/w) sucrose in 2 min (I), 3 min (II) and 4 min (III). The results of the new method were compared with those of the standard method (DIN). With gradients I and II we found values which were significantly higher than those of the standard method (I: 0.49% (w/w); II: 0.46% (w/w); DIN: 0.31% (w/w)), whereas with gradient III the same threshold value was found as with the DIN-Method (III: 0.32% (w/w)).