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S Mayor

Publications and source records attributed to S Mayor.

At least 73 records · Page 4Linked to original sources
PubMed1998

GPI-anchored proteins are organized in submicron domains at the cell surface.

Lateral heterogeneities in the classical fluid-mosaic model of cell membranes are now envisaged as domains or 'rafts' that are enriched in (glyco)sphingolipids, cholesterol, specific membrane proteins and glycosylphosphatidylinositol (GPI)-anchored proteins. These rafts dictate the sorting of associated proteins and/or provide sites for assembling cytoplasmic signalling molecules. However, there is no direct evidence that rafts exist in living cells. We have now measured the extent of energy transfer between isoforms of the folate receptor bound to a fluorescent analogue of folic acid, in terms of the dependence of fluorescence polarization on fluorophore densities in membranes. We find that the extent of energy transfer for the GPI-anchored folate-receptor isoform is density-independent, which is characteristic of organization in sub-pixel-sized domains at the surface of living cells; however, the extent of energy transfer for the transmembrane-anchored folate-receptor isoform was density-dependent, which is consistent with a random distribution. These domains are likely to be less than 70 nm in diameter and are disrupted by removal of cellular cholesterol. These results indicate that lipid-linked proteins are organized in cholesterol-dependent submicron-sized domains. Our methodology offers a new way of monitoring nanometre-scale association between molecules in living cells.

Animals↗
PubMed1998

Cholesterol-dependent retention of GPI-anchored proteins in endosomes.

Several cell surface eukaryotic proteins have a glycosylphosphatidylinositol (GPI) modification at the Cterminal end that serves as their sole means of membrane anchoring. Using fluorescently labeled ligands and digital fluorescence microscopy, we show that contrary to the potocytosis model, GPI-anchored proteins are internalized into endosomes that contain markers for both receptor-mediated uptake (e.g. transferrin) and fluid phase endocytosis (e.g. dextrans). This was confirmed by immunogold electron microscopy and the observation that a fluorescent folate derivative bound to the GPI-anchored folate receptor is internalized into the same compartment as co-internalized horseradish peroxidase-transferrin; the folate fluorescence was quenched when cells subsequently were incubated with diaminobenzidine and H2O2. Most of the GPI-anchored proteins are recycled back to the plasma membrane but at a rate that is at least 3-fold slower than C6-NBD-sphingomyelin or recycling receptors. This endocytic retention is regulated by the level of cholesterol in cell membranes; GPI-anchored proteins are recycled back to the cell surface at the same rate as recycling transferrin receptors and C6-NBD-sphingomyelin in cholesterol-depleted cells. Cholesterol-dependent endocytic sorting of GPI-anchored proteins is consistent with the involvement of specialized lipid domains or 'rafts' in endocytic sorting. These results provide an alternative explanation for GPI-requiring functions of some GPI-anchored proteins.

Animals↗
PubMed1998

A rationale for staged teaching of basic life support.

Basic life support is a crucial part of the Chain of Survival. Unfortunately, however the skill is complex and cannot readily be acquired--let alone retained--in the course of a single training session. Although the problem has long been recognised, no new strategies have been widely implemented to counter the problem. We believe that staged teaching of CPR might provide a solution, and we have devised a program to test this new method. It involves three stages of instruction that we have called bronze, silver, and gold standards. The bronze standard involves opening the airway and providing chest compression without active ventilation: this alone may widen the window of opportunity for successful defibrillation in adult victims in out-of-hospital cardiac arrest. Ventilation is introduced at silver stage using a ratio of 50:5, with emphasis on its value in the resuscitation of children being used as motivation to bring people back for a second period of instruction. The gold stage teaches conventional CPR. A pilot study has been encouraging and a randomized trial on skill acquisition and skill retention is planned.

Adult↗
PubMed1997

Bafilomycin A1 treatment retards transferrin receptor recycling more than bulk membrane recycling.

Treatment of Chinese hamster ovary cells with the vacuolar proton pump inhibitor bafilomycin A1 causes a 2-fold retardation in the rate of recycling of transfected human transferrin receptors back to the cell surface as measured using biochemical assays (Johnson, L. S. , Dunn, K. W., Pytowski, B., and McGraw, T. E. (1993) Mol. Biol. Cell 4, 1251-1266). We have used quantitative fluorescence microscopy to determine which step(s) in the endocytic recycling pathway are affected. We show that removal of transferrin from sorting endosomes and accumulation in the peri-centriolar endocytic recycling compartment takes place normally in bafilomycin A1-treated cells. However, the rate constant for exit of transferrin receptors from recycling endosomes (ke) is reduced from 0.063 min-1 in untreated cells to 0.034 min-1 in the presence of bafilomycin A1. This retardation appears to be dependent on the presence of internalization motifs in the cytoplasmic domain since modified receptors lacking these oligopeptide motifs do not show as large a decrease in recycling rate in the presence of bafilomycin A1. Bulk membrane recycling (measured by efflux of an internalized fluorescent lipid analog, 6-[N-[7-nitrobenzo-2-oxa-1, 3-diazol-4-yl--amino-hexoyl- sphingosylphosphorylcholine) is slowed from an exit rate constant of 0.060 min-1 without drug to 0.046 min-1 in the presence of bafilomycin A1. We conclude that bafilomycin A1 slows bulk membrane flow, but it causes additional inhibition of receptor recycling in a manner that is dependent on a peptide motif on the cytoplasmic domain.

4-Chloro-7-nitrobenzofurazan↗