Subcellular localization of RNAs in transfected cells: role of sequences at the 5' terminus

We have investigated the intracellular location of RNAs transcribed from transfected DNA. COS cells transfected with a clone containing the human adult beta globin gene contain three classes of globin RNAs. Their 3' termini and splice sites are indistinguishable from those of mature reticulocyte beta globin mRNA, and they are polyadenylated. However, as determined by S1 mapping, their 5' sequences are different. The 5' terminus of one is the same as that of mature beta globin mRNA (+1, cap site). The presumed 5' terminus of the second is located 30 nucleotides downstream from the cap site (+30). The third class contains additional nucleotides transcribed from sequences located 5' to the cap site (5' upstream RNA). The 5' upstream RNA molecules are restricted to the nucleus and are more stable than heterogeneous nuclear RNA. The +30 and +1 RNAs are located primarily in the cytoplasm. The data support the notion that nucleotide sequences and/or secondary modifications in the 5' region determine if an RNA is to be transported.     

Ornithogenic soils of the Cape Bird adelie penguin rookeries,Antarctica

Summary Ammonia evolution and urease, phosphatase and protease activities were measured after field incubations of soil from 5 sites, differently influenced by penguin guano, from Cape Bird, Antarctica. Rates of NH₃ production were low considering the large amounts of free NH₃ in these soils. Enzyme activities were higher than expected and, under optimum conditions of temperature, moisture and substrate availability, were sufficient to degrade rapidly the accumulated organic matter in these soils. Laboratory incubations confirmed these high levels of activity; they also showed high dehydrogenase, but not sulphatase, activity. The surface horizon from the site that received continuous additions of fresh penguin excreta had considerably more enzyme activity than the other samples. The optimum activity of phosphatase in this sample was at pH 9.5-10. These enzyme activities are compared with those of New Zealand topsoils, and considered in relation to activities likely to occur under field conditions. Recommendations are made for further biochemical studies at Cape Bird.     

Litter production and forest floor nutrient dynamics in pine and hardwood stands of New Brunswick, Canada

Biomass and nutrient transfer (N, P, K, Ca, Mg) of overstory (branches and leaves) and understory litter fall were examined over a two year period in four jack pine stands aged 16, 29, 49 and 57 years and four mixed hardwood stands aged 7, 17, 20 and 29 years. Relative amounts of the five nutrients in litter fall for both series of stands were N > K ≷ Ca > P = Mg. Return of mineral elements to the forest floor was generally twice as high on the hardwood stands as for similarly aged pine stands. Overall return of nutrients plotted versus stand age generally exhibited a plateau relationship, with relatively little difference among stands; however, some exceptions occurred. Understory contribution to litter fall was very important on these stands, since in most cases the nutrient mass in understory litter was usually similar to or higher than that from the tree layer. Data on forest floor biomass, nutrient distribution and turnover rates of these stands were also presented; mobility of nutrients in the forest floor was in the order K > Mg ≥ P ≥ Ca ≥ N.     

The cost of cardiac surgery.

A study in Wessex has shown that at 1977 prices, and excluding the cost of equipment already installed in the unit, the cost of replacing a man''s aortic valve in this unit is about 1800 pounds. Nevertheless, this seems a small price to pay for return to health and full working capacity, particularly since such patients no longer need to draw social security benefits and their tax contributions will return to normal, thus probably paying for the operation within two years.     

Membrane particle changes attending the acrosome reaction in guinea pig spermatozoa

To examine the freeze-fracture appearance of membrane alterations accompanying the preparation of sperm membranes for fusions-the first preparatory stage occurring before physiological release of the acrosomal content, the second afterward-we induced the acrosome reaction in capacitated guinea pig spermatozoa by adding calcium to the mixture. The most common features observed before fusion of the acrosomal and plasma membranes were the deletion of fibrillar intramembranous particles from the E-fracture faces of both membranes, and the clearance of globular particles from the P face of the plasma membrane-events taking place near the terminus of the equatorial segment. Large particles, >12nm, remained not far from the cleared E-face patches. The P face of the outer acrosomal membrane is virtually clear from the outset. In addition, when fusion was completed, occasional double lines of large particles transiently embossed the P face of the plasma membrane (postacrosomal) side of the fusion zone. Behind the line of fusion, another series of particle-cleared foci emerged. We interpreted these postfusion membrane clearances as a second adaptation for sperm-egg interaction. Induction of the acrosome reaction in media containing phosphatidylcholine liposomes resulted in their apparent attachment, incorporation, or exchange in both the originally and secondarily cleared regions. Our observations support the concepts that membranes become receptive to union at particle- deficient interfaces, and that the physiologically created barren areas in freeze-fracture replicas may herald incipient membrane fusion.     

Hard quasispherical model for the viscosity of hemoglobin solutions.

The viscosity of concentrated hemoglobin solutions of moderate ionic strength at pH values near the isoelectric point may be quantitatively described by the generalized form of a relation commonly applied to suspensions of hard spherical particles. This finding is consistent with the hard quasispherical model previously proposed to account for the thermodynamic properties of concentrated hemoglobin solutions under comparable conditions (1).     

Reconstitution of band 3, the erythrocyte anion exchange protein

Band 3, the erythrocyte membrane protein thought to be responsible for anion transport, was purified to near homogeneity using a Concanavalin A affinity column. Band 3 was then combined with egg lecithin, erythrocyte lipid, cholesterol, and glycophorin, the major erythrocyte sialoglycoprotein, to form vesicles capable of rapid sulfate transport. The transport activity was sensitive to prior treatment of the erythrocytes with pyridoxal phosphate-NaBH₄, a potent inhibitor of anion transport in these cells.     

Two cholesterol ester hydrolases. Distribution in rat tissues and in cultured human fibroblasts and monkey arterial smooth muscle cells.

Hydrolytic activity against acetone-dispersed [4-14C]cholesterol oleate has been assayed as a function of pH in seven parenchymal tissues, blood cells, and plasma of the rat, as well as in cultured human fibroblasts and monkey (Macaca nemestrina) arterial smooth muscle cells. Both acid and neutral hydrolytic activities were present in all of these except rat plasma. The pH optima were in all cases close to pH 4.5 and pH 6.8. Acid activity was quite constant from tissue to tissue, while neutral activity varied greatly, being greatest in adrenal, testis, and adipose tissue. Subcellular fractionation of human fibroblasts allowed demonstration that activities at pH 4.5 and pH 6.8 were concentrated in different fractions, apparently lysosomal and polysomal, respectively. It appears most cell types, including fibroblasts and smooth muscle cells, contain two separate enzymes capable of hydrolyzing cholesterol esters. The neutral pH polysomal enzyme, which is especially prominent in certain tissues, may have a function related to the specialized roles of these tissues.     

Modifications of anionic-lipid domains preceding membrane fusion in guinea pig sperm

The relationship between anionic-lipid concentration and the functional properties of plasma-membrane domains was explored using the guinea-pig sperm membrane as a model, with polymyxin B (PXB) as a probe. Areas of plasmalemma specialized for fusion during the acrosome reaction had a higher affinity for the probe than adjacent nonfusigenic regions. In addition, capacitation--a process preceding acrosome:plasma-membrane fusion--markedly enlarged the area susceptible to PXB binding over the acrosomal cap. Protease treatment mimicked capacitation by increasing the acrosome-reaction incidence as well as PXB binding, at enzyme concentrations not affecting the surface coat nor altering filipin/sterol localization. Both proteolytic digestion and capacitation failed to augment PXB- or filipin-affinity in nonfusigenic zones, such as the post-acrosomal segment, including its particle-free maculae. Incubation of sperm in capacitating medium supplemented with 32P-labeled phosphate, followed by lipid extraction, thin-layer chromatography, and autoradiography, revealed a radioactive band comigrating with cardiolipin and phosphatidic acid. Vermiform protrusions elicited by PXB in the outer lamellae of cardiolipin- phosphatidylcholine liposomes resembled those seen in fusional regions of sperm membrane. We conclude that (a) differing concentrations of anionic lipids are found in adjacent domains of the sperm plasma membrane; (b) these domains mirror the functional regions of the membrane, with higher anionic-lipid concentrations localized over fusional zones; (c) the surface coat does not participate in the maintenance of such domains; (d) anionic-lipid synthesis may contribute to their formation; and (e) anionic-lipid concentrations increase as the membrane becomes fusionally competent, indicating that cellular modulation of lipid domains accompanies regulation of membrane function.