RAM, a gene of yeast required for a functional modification of RAS proteins and for production of mating pheromone a-factor

We have identified a gene (SUPH) of S. cerevisiae that is required for both RAS function and mating by cells of a mating type. supH is allelic to ste16, a gene required for the production of the mating pheromone a-factor. Both RAS and a-factor coding sequences terminate with the potential acyltransferase recognition sequence Cys-A-A-X, where A is an aliphatic amino acid. Mutations in SUPH-STE16 prevent the membrane localization and maturation of RAS protein, as well as the fatty acid acylation of it and other membrane proteins. We propose the designation RAM (RAS protein and a-factor maturation function) for SUPH and STE16. RAM may encode an enzyme responsible for the modification and membrane localization of proteins with this C-terminal sequence.     

Methylammonium uptake by Rhodobacter capsulatus

The ammonium uptake system of Rhodobacter capsulatus B100 was examined using the ammonium analog methylammonium. This analog was not transported when cells were grown aerobically on ammonium. When cultured on glutamate as a nitrogen source, or when nitrogen-starved, cells would take up methylammonium. Therefore, in cells grown under nitrogen-limiting conditions, a second system of ammonium uptake (or a modified form of the first) is present which is distinguished by its capacity for transporting the analog in addition to ammonium. The methylammonium uptake system exhibited saturation kinetics with a K _m of 22 M and a V _max of about 3 nmol per min · mg protein. Ammonium completely inhibited analog transport with a K _i in the range of 1 M. Once inside the cell methylammonium was rapidly converted to -N-methylglutamine; however, a small concentration gradient of methylammonium could still be observed. Kinetic parameters reflect the effects of assimilation.The methylammonium uptake system was temperature and pH dependent, and inhibition studies indicated that energy was required for the system to be operative. A glutamine auxotroph (G29) lacking the structural gene for glutanime synthetase did not accumulate the analog, even when nitrogen starved. The Nif^- mutant J61, which is unable to express nitrogenase structural genes, also did not transport methylammonium, regardless of the nitrogen source for growth. However, the mutant exhibited wild-type ammonium uptake and glutamine synthetase activity. These data suggest that transport of ammonium is required for growth on limited nitrogen and is under the control of the Ntr system in R. capsulatus.Abbreviations CCCP carbonyl cyanide-m-chlorophenyl hydrazone - CHES cyclohexylaminoethanesulfonic acid - DMSO dimethyl sulfoxide - GMAD -N-methylglutamine - GS glutamine synthetase - MES 2-(N-morpholino) ethanesulfonic acid - MSX methionine-Dl-sulfoximine - pCMB p-chloromercuribenzoate - Tricine N-tris(hydroxymethyl)methylglycine     

The complete coding sequence of the human raf oncogene and the corresponding structure of the c-raf-1 gene.

The complete 648 amino acid sequence of the human raf oncogene was deduced from the 2977 nucleotide sequence of a fetal liver cDNA. The cDNA has been used to obtain clones which extend the human c-raf-1 locus by an additional 18.9 kb at the 5' end and contain all the remaining coding exons.     

Temporal Distribution of [3H]-Imipramine Binding in Rat Brain Regions is Not Changed by Chronic Methamphetamine

Specific binding of [³H]-imipramine in the rat suprachiasmatic nuclei, occipital cortex and caudate putamen underwent significant and replicable changes throughout 24 hr under a light-dark cycle or under constant conditions. Daily variations were also found in the medial and dorsal raphe nuclei and the lateral hypothalamus. Methamphetamine, a psychoactive drug with marked effect on circadian rhythms in physiological and hormonal parameters and adrenergic receptors, did not have any significant effect on imipramine binding rhythms in eight discrete brain regions. Thus a drug known to reduce serotoninergic neurotransmission did not change characteristics of the modulatory binding site related to serotonin uptake.     

Autophosphorylation of type 2 casein kinase TS at both its α- and β-subunits: Influence of different effectors

Rat liver casein kinase TS (Ck-TS) having quarternary structure α₂β₂, autophosphorylates at its 25 kDa, β-subunits, incorporating up to 1.2 mol P/mol enzyme. According to their effects on the autophosphorylation pattern the effectors of Ck-TS activity can be grouped into 3 classes: (i) inhibitors, like heparin, which also prevent the autophosphorylation of the β-subunit; (ii) stimulators possessing several amino groups (like spermine) which increase the autophosphorylation at the β-subunit; (iii) stimulators possessing several guanido groups, like protamines and related peptides, which prevent the phosphorylation of the β-subunit, while promoting the autophosphorylation of the 38 kDa α-subunit. In the presence of such polyarginyl effectors the 130 kDa Ck-TS is converted into forms with higher sedimentation coefficient.     

Morphological variation of Keratella cochlearis (Gosse) in Lake Biwa,Japan

The length of the lorica (LL) of Keratella cochlearis cochlearis and of K. cochlearis tecta and the length of posterior spine (PSL) of the latter morphotype were measured in the strongly eutrophic basin and also in the mesotrophic basin of Lake Biwa, Japan, from September to December, 1980. In the population from the mesotrophic basin, the individuals with longer PSL prevail and the tecta forms are extremely rare. The LL values of both morphotypes from one sample do not differ. In December the LL increased to 95 µm in both morphotypes from 80 µm observed in September, while the PSL values decreased abrubtly in both basins in the middle of this period. It is suggested that the observed increase of LL could be related to the thermic factor, i.e. a steady decrease of water temperature, and the changes of PSL are correlated with the increase of nannoplankton and detritus aggregates noted in November. In this month an increase in fecundity and in the total numbers of rotifers took place as well (Hillbricht-Ilkowska, in press).     

Comparison of various atmospheric conditions for isolation and subcultivation of Mycoplasma hyorhinis from cell cultures

The efficiency of aerobic incubation was compared with incubation under various oxygen and carbon dioxide conditions for the isolation and subcultivation of three strains of Mycoplasma hyorhinis from VERO-cell cultures and subcultivation of three laboratory strains. Under anaerobic conditions with a low oxidation-reduction potential (at or below -115 mV) as obtained in jars, with catalysts, containing mixtures of 5%-10% CO2 in H2, very poor or no growth of any of the six M. hyorhinis strains was observed. When traces of oxygen were present (that is, under conditions with higher oxidation-reduction potentials, e.g. when omitting the catalyst in the above gas mixtures or in 5% CO2 + 95% N2) isolation from cell cultures was successful in most tests, but subcultivation of these primary isolates was seldom possible under these semi-anaerobic conditions. However, in most cases these primary isolates could be subcultivated aerobically, although aerobic conditions were unsatisfactory for isolation in about half of the experiments. Isolation of M. hyorhinis was optimal in 5% O2 + 95% N2, under which condition the isolates could also always be subcultivated. Isolation failed occasionally when 5% O2 + 5% CO2 + 90% N2 was used, thus indicating that 5% CO2 was slightly inhibitory. 5% CO2 in air and 10% CO2 either in air, H2 or N2 were also inadequate for isolation from cell cultures. In contrast to the findings with these cell culture-adapted M. hyorhinis strains, the laboratory strains could be subcultivated easily under all conditions tested except those with an oxidation-reduction potential at or below -115 mV; 100% CO2 was inhibitory for all 6 strains. Our findings may partly explain why M. hyorhinis is often considered "non-cultivable" on artificial media once adapted to cell cultures. The findings emphasize the need to employ also a micro-aerophilic condition (5% O2 in 95% N2) in the examination of cell cultures for mycoplasma.     

Kinetics of expression of infectious ecotropic, xenotropic, and mink cell focus-forming murine leukemia virus after 5-iododeoxyuridine induction of cells from high- and low-leukemia mouse strains

Endogenous murine leukemia virus (MuLV) was induced with 5-iododeoxyuridine (IdUrd) from the high-leukemia mouse strain AKR and from two low-leukemia strains, C3H/He and BALB/c. A virus-free cell line from strain AKR readily gave rise to infectious, XC-positive MuLV upon treatment with IdUrd, whereas cells from strains C3H/He and BALB/c produced replication-deficient, XC-negative MuLV. IdUrd-induced cells also produced xenotropic and mink cell focus-forming MuLV. Xenotropic virus emerged at a higher titer from both AKR and BALB/c cells during two discrete time intervals, first at day 3 after induction and a second time during spread of the induced ecotropic MuLV. Xenotropic and mink cell focus-forming MuLVs were also produced by IdUrd-induced C3H/He cells but required another round of infection in Sc-1 cells for detection. The in vitro infectivity of endogenous ecotropic MuLV isolated by IdUrd induction from C3H/He cells correlated with pathogenicity in the Fv-1-compatible, leukemia-negative mouse strain NFS/N. Thus, the virulence of endogenous ecotropic MuLV may be an important factor in determining the leukemia incidence in these inbred strains of mice.     

Different structure of polytene chromosomes ofPhaseolus coccineus suspensors during early embryogenesis

Summary The pattern of DNA and RNA puffs in pair VII of polytene chromosomes has been investigated in the suspensor ofPhaseolus coccineus during early embryo development. The pattern of³H-TdR and³H-U incorporation has been also detected. Collected data indicate that: 1. both heterochromatic regions, p₁₁ and q_(111+112), of chromosome pair VII, organize large DNA puffs; 2. DNA puffs of both regions are specific of different embryo differentiation steps; 3. a seasonal influence on the DNA puffing seems also to be present, as demonstrated by the comparison of the results collected in two different crops; 4. the incorporation experiment by³H-TdR evidences that not all DNA puffs show clustered labeling; 5. the RNA puffing of the two regions seems also to be specific of determined embryo stages.     

Soybean leaf urease: Comparison with seed urease

Soybeans, Glycine max (L.) Merr., from ureides for transport of nitrogen from the root nodule to the shoot. The most direct routes for ureide utilization include the degradation of ureide-derived urea to NH₃ and CO₂. Ureolytic activity was found in leaf disks of soybean and exhbited optimal activity at pH 7 in the presence of a high concentration of urea (250 m M ). In vitro studies showed neither urea amidolyase nor urea dehydrogenase activity in soybean leaves and the ureolytic activity was characterized as urease. Several biochemical properties of soybean leaf urease were determined and compared to seed urease properties. Soybean leaf urease differed from that of seed in five ways: pH optima (5.25 and 8.75), apparent K_m (0.8 m M ), no inhibition by hydroxyurea, faster electrophoretic mobility and no cross-reactivity with soybean seed urease antibodies. The data suggest that urease is the primary urea metabolizing enzyme present in soybean leaves. The properties of soybean leaf urease support the conclusion that a unique isozyme of urease is present in leaf tissue.