A new type of lipid organization is observed in mixtures of phosphatidyl-choline with cardiolipin (in the presence of Ca2+), monoglycosyldiglyceride and phosphatidylethanolamine (in the presence of cholesterol). This phase is characterised by an isotropic 31P NMR signal and is visualised by freeze-fracturing as particles and pits on the fracture faces of the lipid bilayer. As the most favourable model for this phase we propose the inverted micelle sandwiched in between the two monolayers of the lipid bilayer.

Abstract. The structural interaction of the epidermal growth factor (EGF) receptor and the cytoskeleton of A431 cells has been studied using a monoclonal anti-EGF receptor antibody. This has been done with immunogold labeling using a variety of electron microscopical preparation procedures and EGF binding studies. By providing an image of the membraneassociated cytoskeleton, the dry cleavage method reveals a preferential localization of EGF receptors superimposed upon cytoskeletal filaments. The colocalization of gold particles with cytoskeletal filaments is not affected when pre-labeled cells are extracted with the non-ionic detergent Triton X-100, as visualized by dry cleavage. Using surface replication, this treatment results in visualization of the cytoskeleton. In these latter preparations, it is also observed that EGF receptor-coupled gold particles remain associated with cytoskeletal elements. Moreover, Triton extraction performed before immunogold labeling of EGF receptors demonstrates that isolated cytoskeletons contained binding sites for anti-EGF receptor antibodies. Using stereo micrographs of replica's obtained from these isolated cytoskeletons, it is shown that gold-labeled EGF receptors are exclusively present on the cortical membrane-associated region of the cytoskeleton and not on more intracellular-located filaments.Scatchard analysis of EGF binding to cells fixed with glutaraldehyde and treated with Triton X-100 before and after EGF binding indicates that a high affinity EGF binding site is associated with the Triton X-100 insoluble cytoskeleton.PIDERMAL growth factor (EGF) 1 is one of the most intensively studied polypeptide growth factors, and detailed knowledge has been obtained of the various effects of EGF in its target cells as well as of the molecular characteristics of EGF and EGF receptor (for reviews see references 7, 10, 27). Among the effects of EGF in its target cells are morphological changes, such as rounding up of cells (9) and induction of membrane ruffling and extension of filopodia (8). Furthermore, it has been shown that EGF causes alterations in the distribution of actin and tt-actinin (26). Since morphology and dynamics of the cell are largely maintained by an integrated action of cytoskeletal systems (2, 12), these observations suggest that EGF causes changes in the organization of the cytoskeleton via a direct or indirect coupling. Interestingly, an association of the EGF receptor kinase with the Triton X-100-insoluble cytoskeleton of A431 cells has been described recently (17). Furthermore, an enhanced phosphorylation of high molecular weight cytoskeletal proteins was determined as a consequence of EGFinduced kinase activity (17). In addition to the A431 cells, binding of EGF to purified cytoskeleton of the pheochromocytoma cell line PC12 has also been demonstrated (29).1. Abbreviation used in this paper: EGF, epidermal growth factor. So far, no morphological evidence at the ultrastructural level has been presented for an interaction of EGF receptors with the cytoskele...

Single bilayer lipid vesicles were formed by removal of Triton X‐100 with Bio Beads SM‐2 from a mixture of egg lecithin and a Triton X‐100 extract of human erythrocyte ghosts. Upon freeze‐fracturing, these vesicles showed intramembrane particles, similar to those seen in the erythrocyte membrane. Similar particles were also observed when a partially purified band 3 preparation was used instead of the crude Triton X‐100 extract. In the reconstituted vesicles an equal distribution of the intramembrane particles between the two fracture faces was observed. This is in contrast to the unequal distribution of the particles in the erythrocyte membrane, which did not seem to be altered by removal of the extrinsic proteins. From digestion studies with trypsin and chymotrypsin of vesicles, reconstituted from the crude X‐100 extract, it is concluded that band 3 protein in the vesicle bilayer has a similar orientation as in the native membrane.

A B S T R A C T Platelets are a rich source for the study of inositol lipids in man. The substitution of an EDTAKCl solution for the water component of the Bligh and Dyer procedure permitted quantitative extraction of polyphosphoinositides. The latter, with monophosphoinositide, were found to comprise, on a molar basis, 6.7% of total platelet phospholipids. Study of the incorporation of orthophosphate-'P into platelet phospholipids was further simplified by separating eight 'P-labeled lipids, including the inositides, with a single chromatographic development on formaldehyde-treated paper. Particular attention was paid to the influence of ionic environment on the pattern and degree of labeling.In 300 mOsm media major phospholipids other than the inositides were not labeled. Small amounts of label appeared in certain trace phospholipids, notably phosphatidic acid. In 150 mOsm media, labeling of inositides was moderately increased, that of trace phospholipids enormously so. The increased labeling was not solely due to thrombocytolysis since (a) platelet disruption by sonication or freeze-thawing abolished 'P incorporation into phospholipids and (b) in timed studies, restoration of osmolarity to 300 mOsm by addition of hypertonic sorbitol blunted the enhancement effect of previous 150 mOsm exposure. Lowering K and compensatorily increasing Na concentration of 300 mOsm media also stimulated aP labeling of inositides and, to a lesser extent, the trace phospholipids. However, the pattern and degree of stimulation were not as strikingly altered as in the osmolarity studies.

The synthesis and characterization of amphiphile dimethyl-«-hexadecyl[l l-((l-isocyanoethyl)carbonyloxy)unde-