bluuebunny/crossref_metadata_2025_split huggingface dataset:
Direct linkage of thrombin to its cell surface receptors in different cell types
Authors: Simmer Robert L. , Baker Joffre B. , Cunningham Dennis D. 
Published: May 2005

AbstractWhen 125I‐thrombin was incubated with foreskin fibroblasts, cervical carcinoma cells or fibrosarcoma cells of human origin, or with secondary chick embryo cells or Chinese hamster lung cells, it became directly linked to its cell surface receptors. The thrombin‐receptor complex (TH‐R) was derived exclusively from a pool of 125I‐thrombin that had become specifically bound to the cell surface. The linkage was probably covalent, since the complex was resistant to boiling in sodium dodecyl sulfate and 2‐mercaptoethanol. Raising the pH to 12 disrupted TH‐R, but did not affect a similar complex between epidermal growth factor and its receptor, suggesting that the linkage of these mitogens to their receptors was different. Mild trypsin treatment removed the ability of cells to form TH‐R; however, after a 24‐h incubation in serum‐free medium, trypsin‐treated cells recovered the capacity to form TH‐R, suggesting that TH‐R resulted from interaction of 125I‐thrombin with a cellular rather than a serum component. The mitogenic response of cells to thrombin was inversely related to the fraction of specifically bound 125I‐thrombin represented by TH‐R. The role of TH‐R in mitogenesis may be clarified in future studies by obtaining clones of Chinese hamster lung cells that vary in their capacities to form TH‐R and to respond to the mitogenic action of thrombin.
Role of the microfibrillar system in knob action of transformed cells
Authors: Meek William D. , Puck Theodore T. 
Published: May 2005

AbstractTransformed cells often display knobs (or blebs) distributed over their surface throughout most of interphase. Scanning electron microscopy (SEM) and time‐lapse cinematography on CHO‐K1 cells reveal roughly spherical knobs of 0.5–4 μm in diameter distributed densely around the cell periphery but sparsely over the central, nuclear hillock and oscillating in and out of the membrane with a period of 15–60 sec. Cyclic AMP derivatives cause the phenomenon of reverse transformation, in which the cell is converted to a fibroblastic morphology with disappearance of the knobs. A model was proposed attributing knob formation to the disorganization of the jointly operating microtubular and microfilamentous structure of the normal fibroblast. Evidence for this model includes the following: (1) Either colcemid or cytochalasin B (CB) prevents the knob disappearance normally produced by cAMP, and can elicit similar knobs from smooth‐surfaced cells; (2) knob removal by cAMP is specific, with little effect on microvilli and lamellipodia; (3) immunofluorescence with antiactin sera reveals condensed, amorphous masses directly beneath the membrane of CB‐treated cells instead of smooth, parallel fibrous patterns of reversetransformed cells or normal fibroblasts; (4) transmission electron microscopy (TEM) of sections show dense, elongated microfilament bundles and microtubules parallel to the long axis of the reverse‐transformed CHO cell, but sparse, random microtubules throughout the transformed cell and an apparent disordered network of 6‐nm microfilaments beneath the knobs; (5) cell membranes at the end of telophase, when the spindle disappears and cleavage is complete, display typical knob activity as expected by this picture.
Selective protein transport: Characterization and solubilization of the phosvitin receptor from chicken oocytes
Authors: Woods John W. , Roth Thomas F. 
Published: May 2005

AbstractPhosvitin (PV), a subunit of a female‐specific protein, vitellogenin, binds to oocyte membranes with a KD of 10−6 M. Binding reaches equilibrium within 30 min after incubation at 25°C. Bound 125I‐PV dissociates from the membrane with a t1/2 of 13 h when incubated in buffer. However, when 125I‐PV‐labeled membranes are incubated in buffer containing 10−5 M unlabeled PV, 50% of the initially bound 125I‐PV dissociates from the membrane within 10 min. These results support the conclusion that PV binds to a membrane‐associated receptor.Solubilization studies show that Triton X‐100 solubilizes up to 45% of the total membrane‐bound 125I‐PV. Gel‐exculsion chromatography of the solubilized material yields a 500,000 dalton 125I‐PV‐containing complex separated from free 125I‐PV. The 500,000 dalton complex completely dissociates to yield free 125I‐PV when incubated with excess unlabeled PV. However, when incubated with (1) no addition, (2) IgG, or (3) serum albumin, the extent of dissociation is significantly reduced and is consistent with that which would be predicted on the basis of the observed dissociation rate in the absence of unlabeled PV.These results suggest that bound 125I‐PV can only be displaced by unlabeled PV. These results also indicate that the 500,000 dalton species is a solubilized PV‐receptor complex and that it is possible to solubilize the PV‐receptor in an active form.
Uptake and binding of riboflavin by membrane vesicles of bacillus subtilis
Authors: Cecchini Gary , Kearney Edna B. 
Published: May 2005

AbstractRiboflavin uptake and membrane‐associated riboflavin‐binding activity have been investigated in Bacillus subtilis. The uptake and binding activity of the vitamin were found to be repressed coordinately by riboflavin present in the growth medium. The uptake of riboflavin has been shown to have properties of a carrier‐mediated process, and membrane vesicles have been shown to demonstrate riboflavin counterflow and exchange. The membrane‐associated binding activity for riboflavin has been solubilized with detergents, and a procedure for the partial purification of this component is described. The partially purified riboflavin‐binding component has properties expected for a carrier involved in riboflavin uptake, as it shows saturation kinetics and is inhibited by riboflavin analogues. Evidence is also presented showing that reduced riboflavin binds to a greater extent than oxidized riboflavin, and the possible role of the reduced riboflavin in riboflavin uptake is discussed.
A proton nuclear magnetic resonance investigation of histine‐binding protein J of salmonella typhimurium: A model for transport of L‐histidine across cytoplasmic membrane
Authors: Ho Chien , Giza Yueh‐hua , Takahashi Seizo , Ugen Kenneth E. , Cottam Patricia F. , Dowd Susan R. 
Published: May 2005

AbstractGenetic evidence suggests that the high‐affinity L‐histidine transport in Salmonella typhimurium requires the participation of a periplasmic binding protein (histidine‐binding protein J) and two other proteins (P and Q proteins). The histidine‐binding protein J binds L‐histidine as the first step in the high‐affinity active transport of this amino acid across the cytoplasmic membrane. High‐resolution proton nuclear magnetic resonance spectroscopy at 600 MHz is used to investigate the conformations of this protein in the absence and presence of substrate. Previous nuclear magnetic resonance results reported by this laboratory have shown that there are extensive spectral changes in this protein upon the addition of L‐histidine. When resonances from individual amino acid residues of a protein can be resolved in the proton nuclear magnetic resonance spectrum, a great deal of detailed information about substrate‐induced structural changes can be obtained. In order to gain a deeper insight into the nature of these structural changes, deuterated phenylalanine or tyrosine has been incorporated into the bacteria. Proton nuclear magnetic resonance spectra of selectively deuterated histidine‐binding protein J were obtained and compared to the normal protein. Several of the proton resonances have been assigned to the various aromatic amino acid residues of this protein. A model for the high‐affinity transport of L‐histidine across the cytoplasmic membrane of S typhimurium is proposed. This model, which is a version of the pore model, assumes that both P and Q proteins are membrane‐bound and that the interface between these two proteins forms the channel for the passage of substrate. The histidine‐binding protein J serves as the “key” for the opening of the channel for the passage of L‐histidine. In the absence of substrate, this channel or gate is closed owing to a lack of appropriate interactions among these three proteins. The channel can be opened upon receiving a specific signal from the “key”; namely, the substrate‐induced conformational changes in the histidine‐binding protein J molecule. This model is consistent with available experimental evidence for the high‐affinity transport of L‐histidine across the cytoplasmic membrane of S typhimurium.
5′‐p‐Fluorosulfonylbenzoyladenosine as an ATP site affinity probe for Na+,K+‐ATPase
Authors: Cooper James B. , Winter Charles G. 
Published: May 2005

AbstractWe have investigated the suitability of 5′‐p‐fluorosulfonylbenzoyladenosine (FSBA) as an ATP site affinity probe for the canine kidney Na+,K+‐ATPase. The purified enzyme is slowly inactivated by this compound in suitable buffers, losing about half of its activity over a two‐hour period. The rate of inactivation is more rapid in 0.1 M KCl than in 0.1 M NaCl. Low concentrations of ATP protect the enzyme against inactivation, with half‐maximal effects at 4 μM ATP in 0.1 M NaCl and 350 μM ATP in 0.1 M KCl. ADP also protects against FSBA inhibition, but AMP is ineffective when present at 100 μM levels. This pattern is consistent with the previously described nucleotide specificity of the Na+,K+‐ATPase. Addition of protective amounts of ATP after inactivation has occurred does not restore enzyme activity, indicating that inhibition is irreversible.Measurement of the concentration‐dependence of FSBA inactivation suggests an apparent Kd for binding of this compound well above 1 mM, the solubility limit of the analog. This finding is reinforced by the failure of 1 mM FSBA to compete effectively with ATP for the high‐affinity ATP site of the enzyme. Nevertheless, attachment of the analog to this site is indicated by its ability to prevent [3H]‐ADP binding in proportion to the number of sites it has inactivated. Studies with [3H]‐FSBA show that about 1 mole of the analog attaches specifically to the α subunit per mole of enzyme inactivated. A similar amount of nonspecific labeling also occurs with negligible effect on enzyme activity. These findings suggest that FSBA may be useful in probing the topography of the high‐affinity ATP binding site of the Na+,K+‐ATPase and related enzymes.
Release of adenosine by C1300 neuroblastoma cells in tissue culture
Authors: Green Richard D. 
Published: May 2005

AbstractPrevious work in our laboratory led us to postulate that N2a cells release adenosine into growth medium, where it acts at the extracellular adenosine receptors to modulate the sensitivity of the cells to the cyclic AMP‐elevating effect of adenosine [Green, RD, J Pharmacol Exp Ther 201:610, 1977]. We have now devised a high‐performance liquid chromatographic (HPLC) procedure capable of quantitating the concentrations of adenosine in cells and tissue culture media. Growth media of N2a cells and a variant of N2a cells deficient in hypoxanthine‐guanine phosphoribosyltransferase (HGPRT−) contain 10–20 nM adenosine, while that of a variant deficient in adenosine kinase (AK−) is elevated severalfold. It appears that the concentration of adenosine in growth media is determined by both the rate at which it is released by cells into the medium and the rate at which it is metabolized by adenosine deaminase present in the serum in the growth medium. Both N2a and AK− cells release considerable amounts of adenosine into serum‐free medium (SFM) over a short period. Adenosine release is greater from AK− cells and is accelerated by erythro‐9‐(2‐hydroxy‐3‐nonyl)‐adenine (EHNA), a potent adenosine deaminase inhibitor. This accelerated release is retarded by dipyridamole and homocysteine. Surprisingly, dipyridamole and 4‐(3‐butoxy‐4‐methoxybenzyl)‐2‐imidazolidinone (Ro 20 1724), a potent phosphodiesterase inhibitor, stimulate basal adenosine release from N2a but not from AK− cells. It remains to be determined if this is due to an effect of these compounds on adenosine kinase. These results give further support for the hypothesis that adenosine in growth medium modulates the sensitivity of the cells to the cyclic AMP‐elevating affect of adenosine, and furthermore they suggest that adenosine in growth media may tonically stimulate adenylate cyclase and affect processes controlled by the cyclic AMP:cyclic AMP‐dependent protein kinase system.
Cell‐cell contact and growth regulation of pinocytosis in 3T3 cells
Authors: Davies Peter F. 
Published: May 2005

AbstractIn sub‐confluent cultures of Balb/c‐3T3 cells, pinocytosis rates were increased after exposure to specific growth factors (serum; platelet‐derived growth factor, PDGF; epidermal growth factor, EGF). Conversely, as cells became growth‐inhibited with increasing culture density, there was a corresponding decline in pinocytosis rate per cell. In order to test whether density‐inhibition of pinocytosis was influenced either by the growth cycle or by cell contact independently of growth, cells were induced into a quiescent state at a range of subconfluent and confluent densities. Under such conditions, cell density did not significantly inhibit pinocytosis rate. When confluent quiescent cultures in 2.5% serum were exposed to 10% serum, the resulting round of DNA synthesis was accompanied by enhanced pinocytosis per cell, even though the cells were incontact with one another. Furthermore, in a SV40‐viral transformed 3T3 cell line, both the growth fraction and the pinocytosis rate per cell remained unchanged over a wide range of culture densities. These studies indicate that density‐dependent inhibition of pinocytosis in 3T3 cells appears to be secondary to growth‐inhibition rather than to any direct physical effects of cell–cell contact.
Expression of red cell membrane proteins in erythroid precursor cells
Authors: Yurchenco Peter D. , Furthmayr Heinz 
Published: May 2005

AbstractSpecific antibodies to human glycophorin A and spectrin were used to study the expression of these membrane proteins in normal and pathologic human bone marrow. In immunofluorescence experiments spectrin and glycophorin A are found in 50–60% of the nucleated cells in normal bone marrow. These two proteins are expressed at all stages of red cell differentiation and can be traced at least to the earliest morphologically recognizable nucleated red cell precursor, the proerythroblast; the two proteins are specific for cells of the red cell series and are not found to be expressed in lymphocytic, granulocytic cells or platelets. These conclusions were drawn from studies on bone marrow in patients with a temporary block in erythropoiesis at the level of stem cells or of the pronormoblast. Bone marrow from these individuals either lacked all nucleated cells stainable for glycophorin A and spectrin or contained only pronormoblasts. Similar findings were obtained on spleen cells from mice which were made severely anemic by multiple injections with N‐acetyl‐phenylhydrazine. Antibodies to a sialoglycoprotein isolated from mouse red cell membranes stain 70–80% of all cells in the spleen of anemic animals, while only 1–2% of such cells are seen in the spleen of normal animals. Spectrin and glycophorin A could be labeled metabolically and isolated using specific antibodies. The human tumor cell line K562 expresses both membrane proteins, but induction experiments with various agents thus far have failed to change their expression.
Activation of T lymphocytes by the Fc portion of immunoglobulin
Authors: Thoman Marilyn L. , Morgan Edward L. , Weigle William O. 
Published: May 2005

AbstractT lymphocytes are stimulated to release T‐cell‐replacing factors in response to Fc fragments of human IgG. Lyt 1+23− T cells are directly triggered to factor production by Fc subfragments, derived from intact Fc fragments by macrophage‐dependent enzymatic cleavage. These factor(s) replace T cell function in two Fc‐mediated immune responses; induction of polyclonal antibody synthesis, and potentiation of anti‐SRBC responses.
Long‐term maintenance of “cloned” human PLT cells in TCGF with LCL cells as a feeder layer
Authors: Hank Jacquelyn A. , Inouye Hiroo , Guy Lynn A. , Alter Barbara J. , Bach Fritz H. 
Published: May 2005

AbstractThe long‐term maintenance of T cells “cloned” by limiting dilution in TCGF was enhanced by the use of irradiated autologous lymphoblastoid cell line (LCL) cells as well as irradiated LCL cells of the individual to which the T cells were originally primed. It was possible to obtain more than 1 × 1012 cells from a “clone” seeded at one cell per well. Some of the clones tested express primed LD‐typing activity.
Divalent cation dependent ATPase activities of red blood cell membranes: Influence of the oxidation of membrane thiol groups close to each other
Authors: Scutari G. , Ballestrin G. , Covaz A. L. 
Published: May 2005

AbstractAn Mg2+‐dependent low ATPase activity can be detected in erythrocyte “white membranes,” in addition to that of the well known (Ca2+ + Mg2+)‐ATPase. The thiol oxidizing agent diamide affects both activities. The oxidation of neighboring thiols seems to leave the mechanism of the (Ca2+ + Mg2+)‐ATPase amplification system evoked by Ca2+ largely unaffected. The perturbation caused by diamide in the membranes seems to affect primarily a step of the ATP hydrolysis mechanism that is common to both ATPase activities. The effectiveness of diamide seems to be the same when either Ca2+ and Mg2+, or Mg2+ alone are present during the reagent action. Reduction of disulfide bonds by DTE after diamide treatment restores the (Ca2+ + Mg2+)‐ATPase activity but is unable to take the Mg2+‐ATPase activity back to the original level.The hypothesis is discussed that the redox state of one (or more than one) couple of SH close to each other and possibly connected to the active site, may be an important factor in optimizing the efficiency of Ca action on the (Ca2+ + Mg2+)‐ATPase.
Modulation of EGF binding and action by succinylated concanavalin A in fibroblast cell cultures
Authors: Ballmer Kurt , Burger Max M. 
Published: May 2005

AbstractThe role of the binding of succinylated concanavalin A to tissue culture cells in influencing epidermal growth factor (EGF)‐mediated cell proliferation has been studied. Succinylated concanavalin A dramatically reduces the stimulation of 3T6 cells by EGF in Dulbecco's modified Eagle's medium (DME) containing insulin and vitamin B12 as additional growth factors, but no serum. Furthermore, binding studies using 125I‐labeled EGF have shown that the binding of EGF to the cell surface is reduced upon addition of succinylated concanavalin A.
Restricted infectivity of ecotropic type C retroviruses in mouse teratocarcinoma cells: Studies on viral DNA intermediates
Authors: Yang Wen K. , d'Auriol Luc , Yang Den‐Mei , Kiggans James O. , Ou Chin‐yih , Périès Jorge , Emanoil‐Ravicovitch Rodica 
Published: May 2005

AbstractReplication of Gross strain N‐tropic type C retrovirus was markedly restricted in a pluripotential undifferentiated embryonal cell line (PCC4) of murine teratocarcinoma, whereas the same virus could cause productive infection in a myoblast‐derived differentiated line (PCD1) of the same tumor origin. To investigate the restriction mechanism, we compared the initial viral DNA formation in these two cell lines. Analyses by means of a modified Hirt extraction procedure and a modified Southern gel transfer method indicated that PCC4 and PCD1 cells supported the synthesis of viral DNA intermediates after inoculation of the Gross virus. In both cells, a linear DNA duplex (form III viral DNA) appeared at 4 hr, reached a maximal level at 8–9 hr, and declined rapidly thereafter, while two closed‐circular supercoiled DNA duplexes (form I viral DNA) showed their appearance, increase and decline in the 8–24 hr period. During the period from 34 to 78 hr after virus inoculation, another burst of viral DNA synthesis occurred in PCD1 cells, presumably due to secondary virus infection, while at this period both form III and form I viral DNAs became undetectable in PCC4 cells. The Hirt supernatant DNAs prepared from PCD1 and PCC4 cells 10 hr after virus inoculation were equally infectious for NIH3T3 cells in a DNA transfection assay. Both PCD1 and PCC4 cells were very poor recipients for DNA transfection, although one positive result with PCD1 cells might suggest a difference between the two cell types in this aspect. These results indicate that restriction of type C retrovirus in undifferentated embryonl carcinoma cells occurs at a step subsequent to formation and maturation of viral DNA intermediates.
Inhibition of adipose conversion in 3T3‐L2 cells by retinoic acid
Authors: Murray Thomas , Russell Thomas R. 
Published: May 2005

AbstractThe role of retiinoic acid in modulating the differentiation of 3T3‐L2 fibroblasts into adipocytes has been examined. Results indicate that the retinoid is capable of effectively inhibiting the degree of adipose conversion which is brought about by treatment of preadipocytes with 1‐methyl‐3‐isobutylxanthine plus dexamethasone. Morphological and enzymatic (fatty acid synthetase activity) expression of the adipose phenotype are both inhibited more than 90% by 10−6 M retinoic acid. The inhibition is concentration dependent with retinoic acid levels as low as 10−11 M capable of reducing adipose conversion by 20%. Retinoic acid must be administered simultaneously with the triggering agents to be effective. Exposure of nongrowing preadipocytes to retinoic acid does not alter the ability of the cells to differentiate in response to a subsequent treatment with methylisobutylxanthine plus dexamethasone. Further, the inhibition is reversible. Cultures in which methylisobutylxanthine plus dexamethasone triggered differentiation has been blocked by addition of retinoic acid (10−6M) will readily undergo adipose conversion in response to a second treatment with methylisobutylxanthinthine plus dexamethasone in the absence of the retioid. Similar inhibition of differentiation was found when cultures were treated with drugs in medium supplemented with either newborn calf serum or fetal calf serum. However, the extent to which methylisobutylxanthine plus dexamethasone are able to promote differentiation in these cells is considerably greater in medium containing fetal calf serum.