bcd: A mutation affecting the width of organelle domains in the cortex of Tetrahymena thermophila

Summary A single-gene recessive mutation, bcd (broadened cortical domains), of Tetrahymena thermophila is characterized by a variable broadening of the spatial domains within which cortical organelles, including both the contractile vacuole pores (CVP) and oral apparatus (OA), are formed. The phenotype is not temperature-sensitive. During the development of the organelles of the mutant prior to cell division, extra CVPs and extra oral primordia (OP) appear near ciliary rows adjacent to the rows at which these structures normally form. In the later stages of development, some, but not all, of these extra structures are resorbed, or in the case of the oral domain, multiple adjacent OPs may be completely or partially integrated into a single enlarged OA. When multiple OAs persist, one or more of these may display a reversed orientation reminiscent of those encountered in janus mutants. However, unlike janus, bcd cells do not express any sign of a mirror-image global organization.Our results can best be accounted for by postulating that the bcd mutation affects some common determinant of the widths of both CVP and OA domains. Studies are in progress which explore the relationship between this width-determining mechanism(s) and the mechanism(s) determining the location of cortical organelles around the cell circumference.     

Regulation of beta-glucosidase in Bacteroides ruminicola by a different mechanism: growth rate-dependent derepression.

Bacteroides ruminicola B(1)4, a predominant ruminal and cecal bacterium, was grown in batch and continuous cultures, and beta-glucosidase activity was measured by following the hydrolysis of p-nitrophenyl-beta-glucopyranoside. Specific activity was high when the bacterium was grown in batch cultures containing cellobiose, mannose, or lactose (greater than 286 U/g of protein). Activity was reduced approximately 90% when the organism was grown on glucose, sucrose, fructose, maltose, or arabinose. The specific activity of cells fermenting glucose was initially low but increased as glucose was depleted. When glucose was added to cultures growing on cellobiose, beta-glucosidase synthesis ceased immediately. Catabolite repression by glucose was not accompanied by diauxic growth and was not relieved by cyclic AMP. Since glucose-grown cultures eventually exhibited high beta-glucosidase activity, cellobiose was not needed as an inducer. Catabolite repression explained beta-glucosidase activity of batch cultures and high-dilution-rate chemostats where glucose accumulated, but it could not account for activity at slow dilution rates. Maximal beta-glucosidase activity was observed at a dilution rate of approximately 0.35 h-1, and cellobiose-limited chemostats showed a 15-fold decrease in activity as the dilution rate declined. An eightfold decline was observed in glucose-limited chemostats. Since inducer availability was not a confounding factor in glucose-limited chemostats, the growth rate-dependent derepression could not be explained by other mechanisms.     

Effect of extracellular pH on growth and proton motive force of Bacteroides succinogenes, a cellulolytic ruminal bacterium.

The utilization of cellulose or cellobiose by Bacteroides succinogenes S85 was severely inhibited at pH values of less than 5.7. Since low pH inhibited the utilization of both cellobiose and cellulose, changes in cellulase activity could not explain the effect. At an extracellular pH of 6.9, the pH gradient (delta pH) across the cell membrane was only 0.07 U. As extracellular pH declined from 6.9 to 5.7, intracellular pH decreased to a smaller extent than extracellular pH and delta pH increased. Below pH 5.7, there was a linear and nearly proportional decrease in intracellular pH. B. succinogenes took up the lipophilic cation tetraphenylphosphonium ion (TPP+) in the presence of cellobiose, and uptake was sensitive to the ionophore valinomycin. As pH was decreased with phosphoric acid, the cells lost TPP+ and electrical potential, delta psi, decreased. From extracellular pH 6.9 to 5.7, the decrease in delta psi was compensated for by an increase in delta pH, and the proton motive force ranged from 152 to 158 mV. At a pH of less than 5.7, there was a large decrease in proton motive force, and this decrease corresponded to the inhibition of cellobiose utilization.     

Transposon Mutants of Bradyrhizobium japonicum Altered in Attachment to Host Roots

Transposon mutants of Bradyrhizobium japonicum 110 ARS were produced and screened for changes in attachment ability. Mutant CFK4 produced twice as many piliated cells, attached in 2.5-fold-higher numbers to soybean root segments, and colonized roots in about 2-fold-higher numbers than did the parental strain, 110 ARS. Mutants CFK35 and CFK38 were reduced in their attachment about 2-fold and 3.5-fold, respectively. This corresponded to reductions in piliated cells in their populations, reduced reaction with anti-pilus antiserum, and reduced hydrophobic attachment. Mutants CFK4 and CFK38 nodulated soybeans at about the same level as the parent strain, but CFK35 induced only pseudonodules. Two-dimensional gel analyses of the proteins from the mutants showed relatively few changes in proteins.     

In vivo depletion of mannose receptor bearing cells from rat liver by ricin A chain: effects on clearance of beta-glucuronidase

Ricin A chain has previously been shown to intoxicate macrophages in vitro following binding and endocytosis by the macrophage mannose receptor. In this report it is demonstrated that the intravenous injection of ricin A chain in nephrectomized rats leads to a prolonged plasma half-life for [125I]beta-glucuronidase, a ligand for the mannose receptor. Clearance of [125I]asialofetuin, a ligand for the galactose receptor of hepatocytes, was unaffected by injection of A chain. Microscopic examination of the livers of A chain-treated animals revealed a loss of phagocytic cells from the liver sinusoids. These results suggest that ricin A chain may be useful as a toxin specific for mannose receptor bearing cells of the reticuloendothelial system.     

Interleukin 1 preferentially stimulates the production of tissue-type plasminogen activator by human articular chondrocytes

Interleukin 1, derived from human placenta, stimulates plasminogen activator activity in human articular chondrocytes. The stimulation of plasminogen activator activity can be abolished by preincubation of placental interleukin 1 with an antiserum to homogeneous 22K factor, a species of interleukin 1 beta, indicating that the stimulation of plasminogen activator activity is due to interleukin 1 and not contaminating factors. Chondrocytes produce three species of plasminogen activator, with apparent Mr approximately 50,000, 65,000 and 100,000 as determined after sodium dodecyl sulphate (SDS)-polyacrylamide gel electrophoresis with gels containing casein and plasminogen. Both placental interleukin 1 and 22K factor enhance the production of the species of Mr approximately 65,000 and 100,000. Comparison of the mobility of the plasminogen activator species on SDS-polyacrylamide gel electrophoresis with human urokinase (u-PA) and human melanoma tissue-type plasminogen activator (t-PA) and studies with antibodies to these enzymes indicate that the Mr approximately 50,000 species is a u-PA and the Mr approximately 65,000 a t-PA. The Mr approximately 100,000 species is possibly an enzyme-inhibitor complex. Interleukin 1 therefore appears to enhance the production of t-PA and a putative enzyme-inhibitor complex. Abolition of plasminogen activator activity in the fibrin plate assay with antibodies to t-PA and u-PA also confirms enhanced t-PA production on interleukin 1 stimulation, though there is also evidence for increased cell-associated production of u-PA.     

Serum lipids and lipoproteins in the atherosclerosis prone LA/N corpulent rat

The LA/N rat is one of two congenic strains bred from the original obese, hyperphagic and hypertensive rats of Koletsky. With the exception of hypertension the LA/N strain, when homozygous for the corpulent gene, is phenotypically similar to the parent Koletsky strain and prone to the development of vascular and myocardial lesions. Here we report a detailed analysis of the serum lipids, lipoproteins and apolipoproteins B, E and A-I levels in young adult homozygous corpulent (cp/cp) rats of both sexes and in lean males of the same age which were demonstrable non-carriers (+/+) of the cp gene. Both male and female cp/cp rats were hypertriglyceridemic (282-512 mg/100 ml) and moderately hypercholesterolemic (74-84 mg/100 ml). Elevations in these lipids reflected the presence of large (622 A), triacylglycerol-rich and apoprotein-poor VLDL containing both apolipoproteins Bh and B1 and increased phospholipid-rich HDL. Similar, but less pronounced, elevations in serum apolipoproteins B and E in the cp/cp rats when compared to the +/+ animals were also noted. Apolipoproteins A-I levels were 2.7-3-fold higher in cp/cp rats. The levels of VLDL were significantly higher in female cp/cp rats; however, the levels of IDL (intermediate-density lipoproteins), LDL and HDL were significantly lower than in the more atherosclerosis prone male cp/cp rats. Similarly, apolipoprotein A-I was higher and apolipoprotein B lower in the male cp/cp than in the female cp/cp rats. The LDL (d = 1.030-1.063 g/ml) in cp/cp rats, like that in normal animals, was heterogeneous and contained apolipoproteins Bh, E, A-I and C. This fraction was significantly elevated in male cp/cp rats when compared to females but still represented less than 13% of the total serum cholesterol and less than 6% of the total serum lipids in 3-month-old cp/cp animals. The ratio of cholesterol to phospholipids was significantly lower for all lipoproteins in cp/cp rats when compared to +/+ males and these ratios for female cp/cp rats were in all cases lower than those of male cp/cp animals.     

Nucleoside triphosphate pyrophosphatase of rabbit matrix vesicles, a mechanism for the generation of inorganic pyrophosphate in epiphyseal cartilage

Inorganic pyrophosphate (PPi) may be important in the regulation of mineralisation but its origin in epiphyseal cartilage is ill-defined. Nucleoside triphosphate pyrophosphatase is one potential source, as this enzyme catalyses the formation of PPi from nucleoside triphosphates. This enzyme has been identified in matrix vesicles derived from rabbit epiphyseal cartilage and a method developed to measure the activity using ATP as substrate in intact matrix vesicles under relatively physiological conditions. The enzyme had a high affinity for ATP (Km less than 10 microM) and was also active towards GTP, CTP and UTP. Disruption of the matrix vesicle membrane by sonication failed to alter the activity. Treatment of sonicated matrix vesicles with Triton X-100 increased the activity which may indicate a direct effect of the detergent on the enzyme. Activity towards ATP was inhibited substantially by ADP and AMP and by another potential substrate beta,gamma-methyleneadenosine 5'-triphosphate. Dichloromethylene bisphosphonate, an analogue of the product PPi, inhibited the activity to a lesser extent. Two other potential substrates, NADP+ and thymidine 5'-monophosphate p-nitrophenyl ester were only weakly inhibitory as was 1-hydroxyethylidene 1,1-bisphosphonate. These results imply that nucleoside triphosphates are the substrates in vivo and the inhibitory effects of ADP and AMP suggest mechanisms whereby this activity could be regulated.     

The prickly-pears (Opuntia spp., Cactaceae): A source of human and animal food in semiarid regions

The Cactaceae contain many economically promising species primarily in the genus Opuntia. This genus appears to have its center of genetic diversity in Mexico where it is widely used as fodder, forage, fruit, and a green vegetable. In southwestern United States, the prickly-pears have been considered as both weeds and valuable forage plants. During the frequent, unpredictable droughts, propane torches known as “pear burners” are used to singe the spines off cactus pads so that they can be eaten by livestock. Although spineless varieties of Opuntia can be consumed directly by domestic livestock, they are extremely susceptible to herbivory by wildlife. The Cactaceae possess Crassulacean Acid Metabolism, which can be four- to five-fold more efficient in converting water to dry matter than the most efficient grasses. Some Opuntia strains grow rapidly with fresh-fruit yields of 8,000–12,000 kg/ha/ yr or more and dry-matter vegetative production of 20,000–50,000 kg/ha/yr.