It has been shown that the entry of the malaria parasite into the red blood cell requires the presence of ATP in the host cell cytoplasm. In red blood cell ghosts that contain no ATP the receptor on the extracellular surface remains in place and parasites will bind to the membrane, but will not enter. ATP is thus necessary for one of the steps in the invasion sequence that follows recognition and attachment. The process of entry appears to involve the active participation of the host cell membrane cytoskeleton. We have suggested that the function of the intracellular ATP may be to regulate phosphorylation of the cytoskeleton. We now present evidence that the activity of the membrane-associated cyclic AMP-independent kinase of the red blood cell is inseparable from invasion; the active substrate may be spectrin.

Animals↗

PubMedDate not supplied

Membrane proteins at the interface of erythrocytes fused by treatment with polyethylene glycol.

Fusion of human red cells through the action of polyethylene glycol gives rise to pairs or higher clusters with a common membrane envelope, in which a barrier at the position of the original interface can be seen in phase contrast. At early times this septum contains lipids, as judged by labelling with a fluorescent lipophile, and transmembrane protein; this was shown by the presence of the preponderant component, band 3, detected by a fluorescent label, covalently attached before fusion at an extracellular site, or by immunofluorescence with anti-band 3 antibody. Ankyrin, which is bound to band 3, is also observed in the septum. The lipid thereafter disappears from the interface, carrying most of the band 3 with it. A continuous membrane skeletal network, defined by the presence of spectrin (detected by immunofluorescent staining in epifluorescence and confocal microscopy) appears to persist for long periods, but in many cells interruptions develop in the septum. In other fused pairs, particularly at longer times, the interface is seen to have vanished completely. Protease inhibitors have no discernible effect on any of these observations. The results suggest a model for the events that follow fusion. Covalent cross-linking of membrane proteins beyond a critical level causes inhibition of fusion, suggesting that proteins, probably the membrane skeletal network, regulate the fusion process. The efficiency of fusion is strikingly dependent on the composition of the isotonic medium, being relatively high at an orthophosphate concentration of 5 mM and minimal at 20 mM.

Anion Exchange Protein 1, Erythrocyte↗

PubMedDate not supplied

Structural determinants of the rigidity of the red cell membrane.

Although the stability and viscoelasticity of the red cell membrane are undoubtedly governed by the membrane's underlying protein skeleton, the mechanism by which this network controls elasticity is uncertain. The structural constraints, that impose end-to-end spacing on the spectrin molecules well below that in free solution, may impart rubbery (entropic) elasticity to the system. However, other enthalpic and entropic contributions due to interactions between spectrin chains or between spectrin and other proteins, the lipid bilayer or the solvent must also prevail. To relate structural features to elasticity, explicit measurements of membrane rigidity are required. The most widely used measurement is that of the membrane shear elastic modulus by micropipette aspiration. Analysis of genetic variants of membrane structure have shown that the density of spectrin is directly correlated with membrane rigidity. Although cross-linking of the skeleton increases rigidity, interruption of the continuity of the network by dissociating spectrin tetramers into dimers does not reduce rigidity as might be expected. On the other hand, external ligands that cause new interactions between integral proteins and the skeletal network do increase rigidity. Moreover, hereditary ovalocytes, which have a deletion of 9 amino acids from band 3 at the first point of entry into the membrane, are extremely rigid. This mutation is associated with decreased translational and rotational mobility of the band 3, and may impair flexural freedom of its cytoplasmic domain. It thus appears that elasticity may be regulated not only by the structure of the spectrin network, but also by its interactions with and freedom of motion relative to the lipid bilayer.

Chemical Phenomena↗