COMPARISON OF ATTACHED DIATOM COMMUNITIES ON NATURAL AND ARTIFICIAL SUBSTRATES1

A study of attached diatom communities on artificial and natural substrates was conducted in Wheelwright Pond, New Hampshire, during 1975. There were differences in the species composition growing on artificial glass slides and natural substrates. The slides favored the accumulation of Achnanthes minutissima Kütz. and decreased the relative abundance of Eunotia incisa W. Sm. ex. Greg. and Cocconeis placentula v. euglypta (Ehr.) Cleve. Large growths of planktonic species were also noted on the slides. Compositional differences between slides positioned at 15–30 cm (upper) and 1 m (lower) from the surface of the water were minimal. A similar community composition of periphytic diatoms was found on five aquatic macrophytes.     

Radioreceptor assay for epidermal growth factor.

An established cell line of human lung fibroblasts with a high number of surface receptorsfor mouse epidermal growth factor (mEGF) was used to develop a simple and highly sensitive radioreceptor assay for EGF. ¹²⁵I-Labeled mEGF competed mole for mole with unlabeled mEGF for specific receptors. Optimal range for discriminating EGF concentrations in body fluids and tissue extracts by a competitive binding assay was between 5 and 100 ng/ml. Interassay correlation of variation was 8.47% and the recovery of highly purified mEGF added to serum and urine samples was greater than 95%. Human serum and amniotic fluids contained about 24 and 4 ng/ml, respectively, of mEGF equivalents. Concentrations of mEGF in mouse urine and serum were highly variable and were 2- to 10-fold greater than that previously detected by radioimmune assay. Hypophysectomy nearly abolished submaxillary mEGF content in both male and female mice, but testosterone treatment of hypophysectomized animals restored normal concentrations of mEGF to the glands. mEGF added to culture medium disappeared with time as a function of the number of cellular EGF receptors indicating cellular degradation of the growth factor. The radioreceptor assay for EGF is based on the close biologic relationship between the cell receptor site and the native hormone and should prove to be a useful complementary tool to characterize the physiological role of EGF.     

Modification of oligosaccharide-lipid synthesis and protein glycosylation in glucose-deprived cells

Incubation of vesicular stomatitis virus-infected glucose-starved baby hamster kidney cells with [³⁵S]methionine results in the synthesis of all viral proteins. However, as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and tryptic peptide mapping, the G protein is abnormally glycosylated. Metabolic labeling of the oligosaccharide-lipid precursors with [³H]mannose for 15 min, followed by Chromatographic and enzymatic analysis, indicates that the radiolabeled lipid-linked oligosaccharides are devoid of glucose in contrast to the glucosylated oligosaccharide-lipids synthesized by cells grown in the presence of glucose. Also, in contrast to control cells, examination of the glycopeptide fraction reveals the presence of [³H]mannose-labeled glycopeptides which are resistant to erado-β-N-acetylglucos-aminidase H and are smaller in size than glycopeptides from mature vesicular stomatitis virus. In order to observe these effects, a minimum time of 5 h of glucose deprivation is necessary and the addition of 55 μm glucose or mannose to the medium reverses these effects. These results indicate that vesicular stomatitis virus-infected BHK cells deprived of glucose are unable to glucosylate the oligosaccharide-lipid intermediates and, consequently, are unable to glycosylate the G protein normally.     

Integration of foreign genome fragments into cells transformed or cotransformed with fragmented adenoviral DNA

The integration of DNA of highly oncogenic simian adenovirus type 7 (SA7) and non-oncogenic human adenovirus type 6 (Ad6) into the genome of newborn rat kidney cells transformed by fragmented DNA preparations was studied using reassociation kinetics and spot hybridization. Transforming DNA was fragmented with the specific endonuclease SalI (SA7) and BglII (Ad6). In contrast to the cell transformation by intact viral DNA, transformation by fragmented DNA resulted in integration into the cellular genome of not only the lefthand fragment with the oncogene but also of other regions of the viral genome. Additionally integrated fragments were stable and preserved during numerous passages of cells lines, although they were no expressed, at least in the case of the Ad6-transformed cell line. The integration of the fragments of SA7 DNA was accompanied by loss of 25-50% of the mass of each fragment. Adding the linear form of the pBR322 plasmid to the preparation of transforming Ad6 DNA also contributed to its cointegration into the genome of the transformed cell. This technique of cell cotransformation with any foreign DNAs together with the viral oncogens may be used as an equivalent of an integration vector for eukaryotic cells.     

The transition of the thick ascending limb of Henle's loop into the distal convoluted tubule in the nephron of the rat kidney

Summary Examination of serial semithin sections of rat kidney cortex and a subsequent electron microscopic study of selected areas revealed that the characteristic epithelium of the cortical part of the thick ascending limb of Henle extends for a varying distance beyond the macula densa. The transition from the relatively thin epithelium of the thick ascending limb at this site to the three -or even four-fold thicker epithelium of the convoluted part of the distal tubule is sharply defined and occurs without the interposition of an intermediate cell type.The position of the macula densa at the end but still clearly within the ascending limb of Henle's loop is functionally interpreted to guarantee the separation of the sensor point macula densa from disturbing influences which might arise from the secretory activity of the subsequent tubular portion.Investigations supported by the Deutsche Forschungsgemeinschaft. The skillful technical assistance of Mrs. Saliha Sabanovic is gratefully acknowledged     

Species-specific aggregation factor in sponges. XIII. Entire and core structure of the large circular proteid particle from Geodia cydonium

High-molecular weight particles have been isolated from the sponge Geodica cydonium. In the "native" from these particles consist of a spherical center and have 25-30 filaments attached to it. The core structure of the particles is assembled of a central circle and 25 radially-arranged filaments. The core structure is obtained from the entire structure by incubation in a medium, containing a non-ionic detergent and EDTA. The molecular weight of the enitre structure was in the range of 1.4 X 10(9) daltons or more and of the core structure 6.1 x 10(8) daltons. Two functional proteins are released from the "native" particles: the aggregation factor and the sialytransferase.     

Molecular cloning of bovine leukemia virus DNA integrated into the bovine tumor cell genome

The bovine leukemia virus (BLV) DNA harbored in the bovine tumor cell genome was cloned in lambda Charon 4A phage. Using either representative or 3' half-enriched BLV cDNA as a blot hybridization probe, clone lambda BLV-1 was shown to carry 9 kb of the BLV genome, flanked by cellular sequences at both ends. Restriction mapping with twelve endonucleases and hybridization of the DNA fragments to BLV cDNA representing a 3'-end portion of the viral genome revealed the presence and precise location of two long terminal repeats (LTRs) and virus-cell junctions. Thus, lambda BLV-1 appears to contain the complete BLV genome and flanking tumor cellular sequences. The restriction map of the cloned BLV proviral DNA closely resembles that previously reported for unintegrated linear proviral DNA, but differs significantly from that of the integrated provirus of another BLV isolate, the difference occurring preferentially in the putative gag and pol genes.     

Ouabain-insensitive Na+-stimulated ATPase activity of basolateral plasma membranes from guinea-pig kidney cortex cells: II. Effect of Ca2+

The ouabain-insensitive, Mg²⁺-dependent, Na⁺-stimulated ATPase activity present in fresh basolateral plasma membranes from guinea-pig kidney cortex cells (prepared at pH 7.2) can be increased by the addition of micromolar concentrations of Ca²⁺ to the assay medium. The Ca²⁺ involved in this effect seems to be associated with the membranes in two different ways: as a labile component, which can be quickly and easily ‘deactivated’ by reducing the free Ca²⁺ concentration of the assay medium to values lower than 1 μM; and as a stable component, which can be ‘deactivated’ by preincubating the membranes for periods of 3–4 h with 2 mM EDTA or EGTA. Both components are easily activated by micromolar concentrations of Ca²⁺. The $\text{K}{}_{\mathrm{<mtext>a</mtext>}}$ of the system for Na⁺ is the same, 8 mM, whether only the stable component or both components, stable and labile, are working. In other words, the activating effect of Ca²⁺ on the Na⁺-stimulated ATPase is on the $\text{V}{}_{\mathrm{<mtext>max</mtext>}}$, and not on the $\text{K}{}_{\mathrm{<mtext>a</mtext>}}$ of the system for Na⁺. The activating effect of Ca²⁺ may be related to some conformational change produced by the interaction of this ion with the membranes, since it can also be obtained by resuspending the membranes at pH 7.8 or by ageing the preparations. Changes in the Ca²⁺ concentration may modulate the ouabain-insensitive, Na⁺-stimulated ATPase activity. This modulation could regulate the magnitude of the extrusion of Na⁺ accompanied by Cl^? and water that these cells show, and to which the Na⁺-ATPase has been associated as being responsible for the energy supply of this mode of Na⁺ extrusion.     

Na+-dependent transport of glycine in renal brush border membrane vesicles: Evidence for a single specific transport system

The uptake of glycine in rabbit renal brush border membrane vesicles was shown to consist of glycine transport into an intravesicular space. An Na⁺ electrochemical gradient (extravesicular>intravesicular) stimulated the initial rate of glycine uptake and effected a transient accumulation of intravesicular glycine above the steady-state value. This stimulation could not be induced by the imposition of a K⁺, Li⁺ or choline⁺ gradient and was enhanced as extravesicular Na⁺ was increased from 10 mM to 100 mM. Dissipation of the Na⁺ gradient by the ionophore gramicidin D resulted in diminished Na⁺-stimulated glycine uptake. Na⁺-stimulated uptake of glycine was electrogenic. Substrate-velocity analysis of Na⁺-dependent glycine uptake over the range of amino acid concentrations from 25 μM to 10 mM demonstrated a single saturable transport system with apparent K_m = 996 μM and V_max = 348 pmol glycine/mg protein per min. Inhibition observed when the Na⁺-dependent uptake of 25 μM glycine was inhibited by 5 mM extravesicular test amino acid segregated dibasic amino acids, which did not inhibit glycine uptake, from all other amino acid groups. The amino acids d-alanine, d-glutamic acid, and d-proline inhibited similarly to their l counterparts. Accelerative exchange of extravesicular [³H]glycine was demonstrated when brush border vesicles were preloaded with glycine, but not when they were preloaded with l-alanine, l-glutamic acid, or with l-proline. It is concluded that a single transport system exists at the level of the rabbit renal brush border membrane that functions to reabsorb glycine independently from other groups of amino acids.     

Renal sodium-d-glucose cotransport system. Involvement of tyrosine residues in sodium-transporter interaction

Using brush-border membrane vesicles isolated from calf kidney cortex the effect of tyrosine-reactive reagents on sodium-dependent d-glucose transport was investigated. Treatment of the membranes for 60 min with NBD-Cl (7-chloro-4-nitrobenzo-2-oxa-1,3-diazole), N-acetylimidazole or tetranitromethane decreased d-glucose uptake 50, 70 and 40%, respectively. Tracer exchange experiments revealed that the inhibition of transport is due to a direct modification of the sodium-d-glucose cotransport system. The modification by NBD-Cl decreases the apparent V_max of the transport system with respect to its interaction with sodium. In addition, the rate of inactivation of the transport system by NBD-Cl is reduced in the presence of high concentrations of sodium. The results indicate that tyrosine residues play an essential role in sodium-d-glucose cotransport and are probably involved in the binding and/or transport of sodium by the sodium-d-glucose cotransport system.