bluuebunny/crossref_metadata_2025_split huggingface dataset:
Insulin binding and glucose transport in the R3230AC mammary adenocarcinoma
Authors: Harmon Joan Thilly , Hilf Russell 
Published: May 2005

AbstractCells dissociated from the R3230AC mammary adenocarcinoma from intact and diabetic rats were examined for insulin binding and glucose transport. The Kd for insulin binding, ∼ 10−10 M, was similar in all tumors studied. However, the apparent number of receptor sites per cell increased in cells from diabetic rats. Kinetic analysis of 3‐0‐methyl glucose (3‐OMG) entry showed both diffusional and passive carrier characteristics. Insulin (4 × 10−9 M) in vitro did not affect diffusional entry, whereas the hormone altered the passive carrier system, as reflected by an increase in Km and Vmax. Insulin decreased initial velocity of glucose transport at 4–6 mM glucose levels but increased initial velocity of glucose transport at 20 mM glucose. An explanation of the role of insulin on tumor growth in vivo from effects on glucose transport in vitro is proposed.
Cyclic nucleotides and cell growth
Authors: Seifert W. 
Published: May 2005

AbstractGrowth induction in resting fibroblast cultures by serum or growth factors induces a fast, transient cGMP peak which may constitute the intracellular signal for growth. A similar cGMP peak occurs when 3T3 cells arrested at the restriction point or in G0 by starvation for certain amino acids are induced for growth by readdition of the lacking nutrients. Both 3T3 and SV3T3 cells which are arrested randomly all around the cell cycle do not exhibit major changes in cyclic nucleotides after growth induction.Determination of intracellular cAMP and cGMP levels in normal and transformed fibroblasts under different growth conditions shows that the transition between growing and resting state (G0 arrest) is accompanied and probably induced by characteristic changes in cAMP to cGMP ratios. cGMP is decreased 2‐5‐fold in resting as compared to growing cultures, and increased 10‐20‐fold in activated cultures 20 min after serum induction. No major cGMP change was observed in growing, confluent, or serum‐activated cultures of transformed cells.Measurement of guanylcyclase under unphysiological conditions (2 mM Mn++) in crude and purified membranes from 3T3 and SV3T3 cultures did not show increased enzyme activity in the transformed cells. Significant differences may only show up when synchronized cells pass through the restriction point in G1 phase. As a hypothesis it is proposed that transformed cells have an activated guanylcyclase system or a relaxed cGMP‐pleiotypic response mechanism at the restriction point of their cell cycle.
Vasopressin‐sensitive kidney adenylate cyclase: Modulation of the hormonal response
Authors: Roy Christian 
Published: May 2005

AbstractVasopressin‐sensitive pig kidney adenylate cyclase is sensitive to several effectors, such as Mg2+, other divalent cations, and guanyl nucleotides. The purpose of the present study was to compare the main characteristics of adenylate cyclase activation by vasopressin, Mg2+, and GMPPNP, respectively. Mg2+·ions were shown to exert at least three different effects on adenylate cyclase. The substrate of the adenylate cyclase reaction is the Mg‐ATP complex. Mg2+ interacts with an enzyme regulatory site. Finally, Mg2+ can modulate the hormonal response, with Mg2+ions affecting the coupling function–that is, the quantitative relationship between receptor occupancy and adenylate cyclase activation. At all the magnesium concentrations tested, from 0.25 mM to 16 mM, adenylate cyclase activation was not a direct function of receptor occupancy. At low Mg2+ concentrations, adenylate cyclase activation dose‐response curve to the hormone tended to be superimposable to the hormone dose‐binding curve. These results suggest a role of magnesium at the coupling step between the hormone‐receptor complex and adenylate cyclase response. Cobalt, but not calcium, ions could exert the same effects as Mg2+ ions on this coupling step.GMPPNP induced considerable adenylate cyclase activation (15 to 35 times the basal value). Activation by GMPPNP was highly time and temperature dependent. At 30° C, a 20 to 60 min preincubation period in the presence of GMPPNP was needed to obtain maximal activation. The higher the dose of GMPPNP in the medium, the longer it took to reach equilibrium. At 15° C, activation was still increasing with time after 3 hr preincubation in the presence of the nucleotide. GMPPNP was active in a 10−8 M to 10−5 M concentration range. Unlike the results obtained with lysine vasopressin, the kinetic characteristics of dose‐dependent adenylate cyclase activation curves by GMPPNP were unaffected by varying Mg2+ concentrations except for the increase in velocity when raising Mg2+ concentration. It was not clear whether or not the activation processes by the hormone and by GMPPNP had common mechanisms.
Effect of cholinergic ligands on the lipids of acetylcholine receptor‐rich membrane preparations from torpedo californica
Authors: Martinez‐Carrion M. , Raftery M. A. , Thomas J. K. , Šator V. 
Published: May 2005

AbstractIon permeation, triggered by ligand‐receptor interaction, is associated with the primary events of membrane depolarization at the neuromuscular junction and synaptic connections. To explore the possible sites of ion permeation, the long‐lived fluorescent probe pyrene (fluorescence lifetime ∼400 nsec) has been inserted into the lipid phase of acetylcholine receptor‐rich membrane (AcChR‐M) preparations from Torpedo californica. The pyrene probe is susceptible to both fluidity and permeability changes in the lipid bilayer. These changes are detected by variations in the rate of decay of the excited singlet state of pyrene after pulsation with a 10‐nsec ruby laser flash. Variations of these lifetimes in the membrane preparations alone or in the presence of quenchers show that binding of cholinergic agonists and antagonists, neurotoxins, and local anesthetics to AcChR‐M produces varying effects on the properties of the pyrene probe in the lipid phase.It is concluded that binding of cholinergic ligands to the receptor does not significantly alter the fluidity or permeability of the lipids in the bilayer in contact with pyrene. On the other hand, local anesthetics do affect these properties.
Protein transport: A selective membrane mechanism
Authors: Roth Thomas F. , Cutting John A. , Atlas Susan B. 
Published: May 2005

AbstractProteins are selectively sequestered by a number of cell types. However, only in oocytes is the process sufficiently aggravated and specific to be readily studied. In these cells certain serum proteins are taken up in proportions different from those found in the serum.In vitro incubations of hormonally stimulated and synchronous mosquito oocytes show that the only protein capable of initiating the transport process is the female specific yolk protein. Heterologous proteins such as IgG, bovine serum albumin, cytochrome C, and ferritin are inactive. The female specific protein is a phosphoglycolipoprotein. It is synthesized in the fat body, a liver analog in the insect, and passed into the serum before being transported into the oocytes. Preliminary kinetic analysis shows the uptake process to be specific with an apparent Km of about 10−7 M. Glycolytic inhibitors stop protein uptake.The receptor‐mediated binding steps in the transport process are most easily studied in the chicken because of the enormous amount of oocyte membrane available from a given oocyte and because up to 1 gm of protein is normally transported per day per oocyte. IgG and the hen specific phosvitin lipovitellin are two of the physiologically important proteins that are transported intact into the chicken oocytes. The uptake appears selective as shown by studies with iodinated proteins. Ferritin conjugated to IgG is shown by electron microscopy to bind to isolated plasma membranes only where coated pits have formed, whereas ferritin alone is not seen localized on any membrane surface. These very specialized regions of the membrane are similar to micropinocytotic pits but, in addition, possess on their cytoplasmic side dense ridges that form the coat. Transport involves binding to the coated pits, the pinching off of the pits, and the subsequent movement of the coated vesicles in the cytoplasm.
Purification of plasma membranes from rat mammary gland by a density perturbation procedure
Authors: Huggins John W. , Carraway Kermit L. 
Published: May 2005

AbstractA highly purified plasma membrane fraction was obtained from a microsomal subfraction of rat mammary gland after treatment with digitonin to increase its density. The purified membranes were enriched 70‐fold overall in 5′‐nucleotidase with essentially no contamination from glactosyltransferase, succinate‐INT reductase, or NADPH‐cytochrome c reductase. The mermbranes were also highly enriched in sialoglycoproteins.
A study of adenosine 3′–5′ cyclic monophosphate binding sites of human erythrocyte membranes using 8‐azidoadenosine 3′–5′ cyclic monophosphate, a photoaffinity probe
Authors: Owens James R. , Haley Boyd E. 
Published: May 2005

AbstractAn earlier report (1a) has shown the utility of 8‐N3cAMP (8‐azidoadenosine‐3′, 5′‐cyclic monophosphate) as a photoaffinity probe for cAMP binding sites in human erythrocyte membranes. The increased resolution obtained using a linear‐gradient SDS polyacrylamide gel system now shows that: (1) both cAMP and 8‐N3cAMP stimulate the phosphorylation by [γ‐32P]‐ATP of the same red cell membrane proteins; (2) the protein of approximately 48,000 molecular weight whose phosphorylation by [γ‐32P]‐ATP is stimulated by cAMP and 8‐N3cAMP migrates at a solwer rate than the protein in the same molecular weight range which is heavily photolabeled with [32P]‐8‐N3cAMP; (3) other cyclic nucleotide binding sites exist besides those initailly reported; (4) the variation in the ratio of incorporation of 32P‐8‐N3cAMP into the two highest affinity binding sites appears to be the result of a specific proteolysis of the larger protein.
Morphological and biochemical abnormalities in hearts of cardiac mutant salamanders (Ambystoma mexicanum)
Authors: Lemanski Larry F. 
Published: May 2005

AbstractThe effect of homozygosity for recessive gene c in Ambystoma mexicanum is the absence of a heartbeat even though initially heart development appears normal. Mutant embryos (c/c) are first distinguishable from their normal siblings (+/+; +/c) at stage 34 (7 days after fertilization) when the normals develop contracting hearts. The mutant hearts at this stage, upon gross examination, appear structurally normal but fail to beat. Nevertheless, the mutants survive through stage 41, which is about 20 days beyond the heartbeat stage, and they exhibit normal swimming movements, indicating that gene c does not affect skeletal muscle. Electron microscopic studies of normal hearts show some myofibrils to be present at stage 34; by stage 41, the normal myocardial cells have become highly differentiated muscle cells. Although some mutant heart cells contain a few thin 60 A and thick 150 A filaments, organized myofibrils are absent. Instead, amorphous proteinaceous collections are prominent. Heavy meromyosin (HMM) binding experiments were performed on mutant hearts to determine whether the myocardial cells contain actin. Mutant myocardial cells that are glycerinated but not treated with HMM contain intact amorphous bodies. After incubation in HMM, the amorphous collections are no longer present and large numbers of decorated actin filaments appear. The results suggest that the amorphous proteinaceous collections contain actin in a nonfilamentous form, and the addition of HMM induces this actin to polymerize into filaments. SDS‐polyacrylamide gel electrophoresis of mutant heart tissue supports this conclusion by showing a prominent 43,000 dalton band suggestive of actin. The electrophoresis experiments also demonstrate a significant reduction of myosin heavy chain (200,000 daltons) in mutant hearts when compared to normal, and this latter observation is confirmed by radioimmunoassay experiments. Muscle tropomyosin (34,000 daltons), prominent in normal hearts, is virtually nonexistent in mutants. Thus, it appears that this single gene mutation affects the accumulation and organization of several different muscle proteins, including actin, myosin, and tropomyosin.
Suppression of stimulating cell activity by microtubule‐disrupting alkaloids
Authors: Ranney D. F. , Pincus J. H. 
Published: May 2005

AbstractMicrotubule‐disrupting alkaloids and protein fixatives were used to investigate the nature of an active process that must occur within stimulator cells in order for them to intiate a unidirectional mixed lymphocyte response (MLR). Brief treatment of the stimulator cells (SC) with glutaraldehyde (0.15%), formalin (0.6%), or lanthanum chloride (10−3 M) abolished their capacity to activate responder cells (RC). Pre‐treatment of SC with the microtubule‐disrupting alkaloids, colchicine (c) (10−4 to 10−6) or clochicine + vincristine (c+v) (10−4 to 10−6 M) also abrogated their stimulating capacity. This capacity was not restored by the addition of supernates from untreated cultures, thereby excluding the possibility that the alkaloids acted by decreasing the release of soluble stimulatory factors from SC. The introduction of alkaloid‐inactivated, mitomycin‐treated RC as drug carriers did not affect the mitogenic response of untreated RC to concanavalin A. This excluded a significant leakage of alkaloids from the treated SC and uptake by RC during culture. Lumicolchicine produced no decrease in the stimulating capacity of SC. This suggested that the suppression induced by low concentrations of colchicine resulted from its specific disruption of microtubules. None of the above treatements quantitatively reduced the antigenicity of SC, as evaluated by humoral and cell‐mediated lysis of the treated cells. Also, these treatements produced no significant changes in the specific binding of concanavalin A by SC. These indicate there is a functional interaction of microtubular structures with cell surface antigens that appears to regulate either the capacity of SC to associate with RC, or the ability of SC to form and stabilize stimulatory antigenic configurations on the cell surface.
Nature and origin of patterns of changes in cell shape in embryos
Authors: Jacobson Antone G. , Gordon Richard 
Published: May 2005

AbstractSpatial patterns of the future elongation of cells exist in the early embroyo. In the newt, such a pattern of changers of cell shape contributes to the formation of the neural plate. Regardless of where neural plate. Regardless of where neural plte cells are transplanted, they change shape as prescribed by tge pattern. Embryonic induction has a role in establishing this pattern.
Interaction of charged lipid vesicles with planar bilayer lipid membranes: Detection by antibiotic membrane probes
Authors: Cohen Joel A. , Moronne Mario M. 
Published: May 2005

AbstractA technique has been developed for monitoring the interaction of charged phospholipid vesicles with planar bilayer lipid membranes (BLM) by use of the antibiotics Valinomycin, Nonactin, and Monazomycin as surface‐charge probes. Anionic phosphatidylserine vesicles, when added to one aqueous compartment of a BLM, are shown to impart negative surface charge to zwitterionic phosphatidylocholine and phosphatidylethanolamine bilayers. The surface charge is distributed asymmertically, mainly on the vesicular side of the BLM, and is not removed by exchange of the vesicular aqueous solution. Possible mechanisms for the vesicle‐BLM interactions are discussed.
Measurement of the lateral mobility of cell surface components in single living cells by fluorescence recovery after photobleaching
Authors: Jacobson K. , Derzko Z. , Wu E‐S. , Hou Y. , Poste G. 
Published: May 2005

AbstractThe use of fluorescence recovery after photobleaching (FRAP) techniques to monitor the lateral mobility of plant lectin‐receptor complexes on the surface of single, living mammalian cells is described in detail. FRAP measurements indicate that over 75% of the wheat germ agglutinin receptor (WGA‐receptor) complexes on the surface of human embryo fibroblasts are mobile. These WGA‐receptor complexes diffuse laterally (as opposed to flow) on the cell surface with a diffusion coefficient in the range of 2 × 10−11 to 2 × 10−10 cm2/sec. Both the percentage of mobile WGA‐receptor complexes and the mean diffusion coefficient of these complexes are higher than that obtained from earlier FRAP measurements of the mobility of concanavalin A‐receptor (Con A‐receptor) complexes in a variety of cell types. The possible reasons for the differing mobilities of WGA and Con A receptors are discussed.
The natural occurrence of insulin receptors in groups on adipocyte plasma membranes as demonstrated with monomeric ferritin‐insulin
Authors: Jarett Leonard , Smith Robert M. 
Published: May 2005

AbstractThis study was designed to document whether the reported distribution of insulin receptors in small groups of receptor sites randomly distributed in the glycocalyx of adipocytes and isolated adipocyte plasma membranes was a naturally occurring phenomena or due to artifacts. Possible artifacts include: (1) oligomeric forms of ferritin in the ferritin‐insulin preparation, (2) an uneven distribution of the glycocalyx on the plasma membrane, or (3) ligand‐induced aggregation of occupied receptor complexes. Biogel A 1.5m chromatography of the ferritin‐insulin conjugate revealed the ferritin in the ferritin‐insulin complex to consist of 55% monomers, 15% dimers, and 30% oligomers. The monomer peak was purified (> 95%) for use in these studies. Cationic ferritin, a glycocalyx marker, when incubated with paraformaldehyde‐fixed plasma membranes, was found to be uniformly distributed on the surface of the plasma membrane indicative of uniformly distributed glycocalyx. The ability to demonstrate and inhibit ligand‐induced aggregation on the isolated plasma membrane was established with a multivalent ligand, ferritin‐concanavalin A. More than 66% of the ferritin‐concanavalin A receptors were found in large clusters of 5 or more and 34% as singletons or clusters of up to 4 when incubated at 24°C with fresh membranes. Only 38% of the ferritin‐concanavalin A receptors were in large clusters; 62% were singletons or clusters up to 4 on membranes prefixed with paraformaldehyde before incubation. The distribution of the monomeric ferritin‐insulin was similar on both adipocytes and purified adipocyte plasma membranes and was consistent with earlier reports with ferritin‐insulin. The quantitative distribution of the monomeric ferritin‐insulin as singletons or in groups of 2–6 was comparable between the intact cells and isolated membranes incubated at 24°C. The binding of 500 μUnits monomeric ferritin‐insulin per ml to the isolated plasma membranes was studied under incubation conditions similar to those used with ferritin‐concanavalin A. Under all three conditions, fresh membranes at 24°C and 0–4°C and prefixed membranes at 24°C, the pattern of distribution of the monomeric ferritin‐insulin as singletons or groups of 2–6 was identical, indicating that the ligand was not causing aggregation into clusters as did the concanavalin A. Thus, the occurrence of insulin receptors in small groups appears to be a natural phenomenon in the plasma membrane structure of adipocytes.
Mutations affecting the binding, internalization, and lysosomal hydrolysis of low density lipoprotein in cultured human fibroblasts, lymphocytes, and aortic smooth muscle cells
Authors: Brown Michael S. , Anderson Richard G. W. , Goldstein Joseph L. 
Published: May 2005

AbstractStudies comparing the metabolism of low density lipoprotein (LDL) in normal cells and in cells cultured from patients with homozygous familial hypercholesterolemia have disclosed the existence of a receptor for plasma LDL. This receptor has been identified on the surface of human fibroblasts, lymphocytes, and aortic smooth muscle cells. An extension of these studies to cell strains derived from patients with other single gene defects in cholesterol metabolism has provided additional insight into the normal mechanisms by which cells regulate their cholesterol content and how alterations in these genetic control mechanisms may predispose to atherosclerosis in man.
Transport in halobacterium halobium: Light‐induced cation‐gradients, amino acid transport kinetics, and properties of transport carriers
Authors: Lanyi Janos K. 
Published: May 2005

AbstractCell envelope vesicles prepared from H. halobium contain bacteriorhodopsin and upon illumination protons are ejected. Coupled to the proton motive force is the efflux of Na+. Measurements of 22Na flux, exterior pH change, and membrane potential, ΔΨ (with the dye 3,3′‐dipentyloxadicarbocyanine) indicate that the means of Na+ transport is sodium/proton exchange. The kinetics of the pH changes and other evidence suggests that the antiport is electrogenic (H+/Na+ > 1). The resulting large chemical gradient for Na+ (outside > inside), as well as the membrane potential, will drive the transport of 18 amino acids. The 19th, glutamate, is unique in that its accumulation is indifferent to ΔΨ: this amino acid is transported only when a chemical gradient for Na+ is present. Thus, when more and more NaCl is included in the vesicles glutamate transport proceeds with longer and longer lags. After illumination the gradient of H+ collapses within 1 min, while the large Na+ gradient and glutamate transporting activity persists for 10–15 min, indicating that proton motive force is not necessary for transport. A chemical gradient of Na+, arranged by suspending vesicles loaded with KCl in NaCl, drives glutamate transport in the dark without other sources of energy, with Vmax and Km comparable to light‐induced transport. These and other lines of evidence suggest that the transport of glutamate is facilitated by symport with Na+, in an electrically neutral fashion, so that only the chemical component of the Na+ gradient is a driving force.The transport of all amino acids but glutamate is bidirectional. Actively driven efflux can be obtained with reversed Na+ gradients (inside > outside), and passive efflux is considerably enhanced by intravesicle Na+. These results suggest that the transport carriers are functionally symmetrical. On the other hand, noncompetitive inhibition of transport by cysteine (a specific inhibitor of several of the carriers) is only obtained from the vesicle exterior and only for influx: these results suggest that in some respects the carriers are asymmetrical.A protein fraction which binds glutamate has been found in cholate‐solubilized H. halobium membranes, with an apparent molecular weight of 50,000. When this fraction (but not the others eluted from an Agarose column) is reconstituted with soybean lipids to yield lipoprotein vesicles, facilitated transport activity is regained. Neither binding nor reconstituted transport depend on the presence of Na+. The kinetics of the transport and of the competitive inhibition by glutamate analogs suggest that the protein fraction responsible is derived from the intact transport system.