scholarly journals External labeling of galactose in surface membrane glycoproteins of the intact myelin sheath.

1976 ◽  
Vol 251 (1) ◽  
pp. 153-158
Author(s):  
J F Poduslo ◽  
R H Quarles ◽  
R O Brady
Keyword(s):  
Myelin Sheath ◽  
Surface Membrane ◽  
Membrane Glycoproteins

scholarly journals Synthesis of membrane glycoproteins in rat small-intestinal villus cells. Redistribution of l-[1,5,6-3H]-fucose-labelled membrane glycoproteins among Golgi, lateral basal and microvillus membranes in vivo

1979 ◽  
Vol 182 (1) ◽  
pp. 203-212 ◽  
Author(s):  
Andrea Quaroni ◽  
Katharina Kirsch ◽  
Milton M. Weiser
Keyword(s):  
Golgi Complex ◽  
Membrane Fraction ◽  
Surface Membrane ◽  
Sodium Dodecyl ◽  
Membrane Glycoproteins ◽  
Intestinal Villus ◽  
Microvillus Membrane ◽  
Small Intestinal ◽  
Small Intestinal Villus

The biogenesis of plasmalemma glycoproteins of rat small-intestinal villus cells was studied by following the incorporation of l-[1,5,6-3H]fucose, given intraperitoneally with and without chase, into Golgi, lateral basal and microvillus membranes. Each membrane fraction showed distinct kinetics of incorporation of labelled fucose and was differently affected by the chase, which produced a much greater decrease in incorporation of label into Golgi and microvillus than into lateral basal membranes. The kinetic data suggest a redistribution of newly synthesized glycoproteins from the site of fucosylation, the Golgi complex, directly into both lateral basal and microvillus membranes. The observed biphasic pattern of label incorporation into the microvillus membrane fraction may be evidence for a second indirect route of incorporation. The selective effect of the chase suggests the presence of two different pools of radioactive fucose in the Golgi complex that differ in (1) their accessibility to dilution with non-radioactive fucose, and (2) their utilization for the biosynthesis of membrane glycoproteins subsequently destined for either the microvillus or the lateral basal parts of the plasmalemma. The radioactively labelled glycoproteins of the different membrane fractions were separated by sodium dodecyl sulphate/polyacrylamide-slab-gel electrophoresis and identified by fluorography. The patterns of labelled glycoproteins in Golgi and lateral basal membranes were identical at all times. At least 14 bands could be identified shortly after radioactive-fucose injection. Most seemed to disappear at later times, although one of them, which was never observed in microvillus membranes, increased in relative intensity. All but two of the labelled glycoproteins present in the microvillus membrane corresponded to those observed in Golgi and lateral basal membranes shortly after fucose injection. The patterns of labelled glycoproteins in all membrane fractions were little affected by the chase. These data support a flow concept for the insertion of most surface-membrane glycoproteins of the intestinal villus cells.

scholarly journals Distribution of the cell substratum attachment (CSAT) antigen on myogenic and fibroblastic cells in culture.

1985 ◽  
Vol 100 (5) ◽  
pp. 1528-1539 ◽  
Author(s):  
C H Damsky ◽  
K A Knudsen ◽  
D Bradley ◽  
C A Buck ◽  
A F Horwitz
Keyword(s):  
Cell Adhesion ◽  
Surface Membrane ◽  
Membrane Glycoproteins ◽  
Myogenic Cells ◽  
Microfilament Bundle ◽  
Cell Matrix ◽  
Cell Biol ◽  
The Stability ◽  
Adhesive Contacts ◽  
Fibroblastic Cells

Previous studies (Neff et al., 1982, J. Cell. Biol. 95:654-666; Decker et al., 1984. J. Cell. Biol. 99:1388-1404) have described a monoclonal antibody (CSAT Mab) directed against a complex of three integral membrane glycoproteins of 120,000-160,000 mol wt (CSAT antigen [ag]) involved in the cell matrix adhesion of myoblasts and fibroblasts. In localization studies on fibroblasts presented here, CSAT ag has a discrete, well-organized distribution pattern. It co-aligns with portions of stress fibers and is enriched at the periphery of, but not directly beneath vinculin-rich focal contacts. In this last location, it co-distributes with fibronectin, consistent with the suggestion that the CSAT ag participates in the mechanism by which fibroblasts attach to fibronectin. In prefusion myoblasts, which are rapidly detached by CSAT Mab, CSAT ag is distributed diffusely as are vinculin, laminin, and fibronectin. After fusion, myotubes become more difficult to detach with CSAT Mab. The CSAT ag and vinculin are organized in a much more discrete pattern on the myotube surface, becoming enriched at microfilament bundle termini and in lateral lamellae which appear to attach myotubes to the substratum. These results suggest that the organization of CSAT ag-adhesive complexes on the surface of myogenic cells can affect the stability of their adhesive contacts. We conclude from the sum of the studies presented that, in both myogenic and fibroblastic cells, the CSAT ag is localized in sites expected of a surface membrane mediator of cell adhesion to extracelluon of CSAT ag-adhesive complexes on the surface of myogenic cells can affect the stability of their adhesive contacts. We conclude from the sum of the studies presented that, in both myogenic and fibroblastic cells, the CSAT ag is localized in sites expected of a surface membrane mediator of cell adhesion to extracellular matrix. The results from studies that use fibroblasts in particular suggest the involvement of CSAT ag in the adhesion of these cells to fibronectin.

scholarly journals Biochemical and functional consequences of dissociation of the platelet membrane glycoprotein IIb-IIIa complex

Blood ◽  
1985 ◽  
Vol 66 (1) ◽  
pp. 92-98 ◽  
Author(s):  
SJ Shattil ◽  
LF Brass ◽  
JS Bennett ◽  
P Pandhi
Keyword(s):  
Platelet Aggregation ◽  
Surface Membrane ◽  
Broad Band ◽  
Adenosine Diphosphate ◽  
Platelet Membrane ◽  
Membrane Glycoproteins ◽  
Fibrinogen Receptor ◽  
Activated Platelets ◽  
Discrete Band ◽  
Functional Consequences

Abstract The platelet membrane glycoproteins, IIb and IIIa, form a Ca2+- dependent heterodimer complex that functions as the fibrinogen receptor in activated platelets to mediate platelet aggregation. Little is known about factors that affect the IIb-IIIa complex within the platelet membrane. It has been observed that platelets incubated with ethylene glycol tetra-acetic acid (EGTA) at 37 degrees C are unable to aggregate or to bind monoclonal antibodies specific for the IIb-IIIa complex. To determine whether this is due to a dissociation of IIb from IIIa, we developed a method for quantitating the complex on nondenaturing, polyacrylamide gradient gels. Platelets were surface-labeled with 125I and then solubilized and electrophoresed in 0.2% Triton and 10 mmol/L CHAPS. Under these conditions and in the presence of 1 mmol/L Ca2+, glycoproteins IIb and IIIa migrated on the gels as a discrete band at Rf = 0.33. Protein that was eluted from this band bound to an immunoaffinity column specific for the IIb-IIIa complex. In contrast, when the IIb-IIIa complex was solubilized and then dissociated with EGTA, the discrete band at Rf = 0.33 was no longer present, and IIb and IIIa were now found in a broad band at Rf = 0.45 to 0.50. To study IIb and IIIa within the surface membrane, the 125I-labeled platelets were first incubated with 0.5 mmol/L EGTA (1 nmol/L free Ca2+) at 22 degrees C and then solubilized in the absence of EGTA. The IIb and IIIa from these platelets migrated at Rf = 0.33, indicating the presence of the intact IIb-IIIa complex. In contrast, when the platelets were incubated at 37 degrees C for one hour with the EGTA, the discrete band at Rf = 0.33 representing the IIb-IIIa complex gradually disappeared. This phenomenon could not be reversed by adding Ca2+ back to the platelets before solubilization and electrophoresis. This loss of the IIb-IIIa complex from intact platelets was accompanied by (a) a progressive and irreversible decrease in adenosine diphosphate (ADP)-induced platelet aggregation and (b) decreased binding of a complex-dependent monoclonal antibody to the platelets. These studies demonstrate that when platelets are exposed to low Ca2+ at 37 degrees C, the IIb-IIIa heterodimer complexes in their surface membranes are irreversibly disrupted. Because intact IIb-IIIa complexes are required for platelet aggregation, the loss of these complexes may account for the failure of these platelets to aggregate in response to ADP.

Platelet Hyperreactivity to Submaximal Epinephrine: Biologic and Genetic Correlates.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 3562-3562
Author(s):  
Donald L. Yee ◽  
Carol W. Sun ◽  
Angela L. Bergeron ◽  
David S. Lopez ◽  
Jing-fei Dong ◽  
...  
Keyword(s):  
Healthy Subjects ◽  
Platelet Reactivity ◽  
Platelet Rich Plasma ◽  
Smoking Status ◽  
Surface Membrane ◽  
Thrombin Inhibitor ◽  
Study Cohort ◽  
Membrane Glycoproteins ◽  
Thrombotic Risk

Abstract Platelet hyperreactivity constitutes an important thrombotic risk factor; however, standardized methods for its measurement are lacking. We recently reported that aggregometry using a submaximal concentration of epinephrine (0.4 μM in citrated platelet-rich plasma) identifies individuals with in vitro platelet hyperreactivity; this hyperreactivity was reproducible on multiple occasions over long periods of time (up to 3 years). To better understand this aberrant reactivity, we studied in a larger group of subjects (n=404) the association between healthy individuals’ platelet reactivity to epinephrine and their platelet phenotype as measured by other functional and biochemical assays. Fourteen percent (n=56) of our study cohort showed a hyperreactive response (> 60% aggregation) to 0.4 μM epinephrine; the remainder showed a minimal response (see figure). Subjects with hyperreactivity to epinephrine were more likely to exhibit hyperaggregability to other agonists (ADP, arachidonic acid, collagen, collagen-related peptide and ristocetin; p<.04), increased spontaneous aggregation (p<.001), shorter PFA-100 closure times with both epinephrine (p<.001) and ADP (p<.02) cartridges and increased surface expression of P-selectin after exposure to ADP (p<.02) and to shear stress (p<.01). Subjects exhibiting platelet hyperreactivity in citrated specimens also did so in specimens anticoagulated with the thrombin inhibitor Phe-Pro-Arg-chloromethyl ketone, suggesting that the hyperreactive phenotype is independent of calcium concentration and residual plasma thrombin activity. Taken together, these data obtained using multiple assays and methods of platelet stimulation are most consistent with the existence of a hyperreactive phenotype that is not agonist specific, but is rather a "global" property of the platelet. Platelet hyperreactivity was not associated with age, race, body mass index, smoking status or presence of hypertension, but was independently associated with female gender (p=.02) and with higher fibrinogen levels (p=.002). To explore potential mechanisms responsible for platelet hyperreactivity to epinephrine, we measured expression of key surface membrane glycoproteins (GPs) and genotyped polymorphisms located on candidate genes relevant to epinephrine-mediated platelet activation. We found that hyperreactivity was strongly associated with increased expression of both quiescent and activated forms of the fibrinogen receptor GP IIb-IIIa (p<.005) and with the T allele of the C825T polymorphism on the gene (GNB3) encoding the beta-3 subunit of G proteins (p<.03), but not with the G1838A polymorphism on the gene encoding the α2A-adrenergic receptor. Aggregometry using submaximal concentrations of epinephrine thus identifies a global hyperreactive platelet phenotype and may be useful in settings where platelet hyperreactivity bears special relevance - for example, in populations of patients at risk for thrombosis. Our findings also suggest that the physiologic determinants of platelet hyperreactivity in response to different types of stimulation may share common signaling pathways, possibly involving G protein-coupled receptors and increased GP IIb-IIIa expression. Variation in Platelet Reactivity (404 healthy subjects) Variation in Platelet Reactivity (404 healthy subjects)

scholarly journals Membrane flow during nematode spermiogenesis.

1982 ◽  
Vol 92 (1) ◽  
pp. 113-120 ◽  
Author(s):  
T M Roberts ◽  
S Ward
Keyword(s):  
Caenorhabditis Elegans ◽  
Diffusion Coefficients ◽  
Surface Membrane ◽  
Lateral Diffusion ◽  
Bulk Flow ◽  
Membrane Glycoproteins ◽  
Membrane Flow ◽  
Latex Beads ◽  
Membrane Components

Two distinct types of surface membrane rearrangement occur during the differentiation of Caenorhabditis elegans spermatids into amoeboid spermatozoa. The first, detected by the behavior of latex beads attached to the surface, is a nondirected, intermittent movement of discrete portions of the membrane. This movement starts when spermatids are stimulated to differentiate and stops when a pseudopod is formed. The second type of movement is a directed, continual flow of membrane components from the tip of the pseudopod to its base. Both membrane glycoproteins and fluorescent phospholipids inserted in the membrane flow backward at the same rate, approximately 4 micrometers/min, although their lateral diffusion coefficients in the membrane differ by at least a factor of 5. These observations suggest that pseudopodial membrane movement is due to bulk flow of membrane components away from the tip of the pseudopod.

scholarly journals Hyposialylation of differentiation-inducer-resistant HL-60 cells

Blood ◽  
1986 ◽  
Vol 68 (6) ◽  
pp. 1402-1406
Author(s):  
RE Gallagher ◽  
DA Giangiulio ◽  
CS Chang ◽  
CJ Glover ◽  
RL Felsted
Keyword(s):  
Sialic Acid ◽  
Surface Membrane ◽  
Resistance Mechanism ◽  
Electrophoretic Pattern ◽  
Wild Type ◽  
Membrane Glycoproteins ◽  
Purine Base ◽  
Neuraminidase Treatment ◽  
Total Sialic Acid ◽  
Differentiation Inducer

The total sialic acid concent of retinoic acid (RA)-resistant or 6- thioguanine (6TG)-resistant HL-60 cells was more than tenfold lower and of dimethylsulfoxide (DMSO)-resistant HL-60 cells was approximately twofold lower than that of parental, wild-type (wt) HL-60 cells. Neuraminidase-inaccessible, ie residual cell-associated sialic acid after neuraminidase treatment, was four- to twelvefold lower in the three differentiation-inducer-resistant sublines than in the parent line. Neuraminidase treatment of 125I-labeled surface membrane glycoproteins (SMGs) from wt HL-60 cells converted the two-dimensional gel electrophoretic pattern to one having features in common with RA- and 6TG-resistant cells. However, neuraminidase treatment did not alter the sensitivity of wt HL-60 cells to differentiation induction by RA, hypoxanthine (purine base), or DMSO. These results indicate that differences in peripheral, neuraminidase-accessible sialic acids are important determinants of the gel electrophoretic mobility of the SMGs of the HL-60 line and sublines but are not likely related to the differentiation-resistance mechanism. Further studies are required to determine if hyposialylation of cryptic, neuraminidase-inaccessible sites has functional significance.

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