Interaction of microtubule proteins with phospholipid vesicles.

Abstract

We have examined the interaction of unilamellar dimyristoyl phosphatidylcholine liposomes with the high-speed supernate of brain homogenate and with tubulin purified through one or two cycles of microtubule assembly-disassembly. Tubulin and certain high molecular weight proteins are selectively adsorbed from these mixtures onto liposomes. The composition of adsorbed proteins is similar to that obtained during corresponding cycles of microtubule assembly, suggesting the equivalency of these processes. Adsorption induces stacking and/or fusion of liposomes into multilamellar structures indicating strong protein-lipid interaction. In addition, liposome-adsorbed tubulin forms extensive intermolecular disulfide bridges that are inert to reducing agents in the aqueous medium. The observations form a basis for further study of the distribution, function, and properties of membrane-bound tubulin. KEY WORDS tubulin membrane disulfides phospholipid microtubules liposomesPharmacological evidence has implicated microtubules in the control of a wide range of surface phenomena. For example, microtubule disassembly by colchicine enables the movement of surfacebound concanavalin A into caps in leukocytes (1, 35) and prevents the segregation of membrane proteins (26, 32) and probably lipids (4) that normally accompanies phagocytosis in neutrophils. In addition, studies ofneutrophils from mice with the Chedak-Higashi syndrome show that failure of microtubule assembly causes colchicine-like effects but in the absence of drugs (25). These results indicate that microtubule assembly is required to control and direct the distribution of membrane components.We have suggested that microtubule regulation of surface events may occur indirectly through control of the microfflament-contractile system (5). On the other hand, there is increasing evidence for a direct association of tubulin (the major subunit component of microtubules) with membranes. For example, there is specific colchicine binding to isolated membrane fractions from brain (6, 10); and tubulin has been identified in synaptic vesicles by polyacrylamide gel electrophoresis (7, 11,19), by reaction with specific antibodies (33), and recently by analysis on two-dimensional electropheretograms and by tryptic peptide mapping (17). Stephens' (30) analyses of membranes of the scallop have also supported the existence of membrane-bound tubulin: it was clearly shown that tubulin is associated with ciliary but not flageUar membranes. Finally, previous in vitro studies in our laboratory demonstrated fluorescence resonance energy transfer between fluorescein-labeled neutrophil membranes and rhodamine-labeled brain tubulin (2). At 37~ a high degree of transfer indicated close approximation of the membrane and tubulin. At 0~ transfer did not occur.While these studies indicate the presence of tubulin in isolated membrane fractions, they indicate neither the molecular nor functional basis of the tubulin-membrane interaction. In particular, a major difficulty in establishing the molecu...

Posttranslational modification of tubulin by palmitoylation: I. In vivo and cell-free studies.

“…This has led to the suggestion that the putative membrane tubulin may actually be contaminating cytoplasmic tubulin. Studies showing that cytoplasmic tubulin binds to artificial lipid membranes (Caron and Berlin, 1979;Klausner et al, 1981;Kumar et al, 1981) support this concern. Ideally, one would like to identify a membrane-associated form of tubulin that is biochemically distinct from cytoplasmic tubulin.…”

Appearance of cytoskeletal components on the surface of leukemia cells and of lymphocytes transformed by mitogens and Epstein-Barr virus.

“…Another possibility may be that the framework proteins are iptegral components of the cell membrane, as has been documented for tubulin at the synaptosomal cell junction in higher organisms (27) as well as in scallop gill cilia (28). A physicochemical basis for such associations is also provided by the recent finding that tubulin binds strongly with, and can be embedded within, simple phospholipid bilayers (29).…”

Folding of the Repeat Domain of Tau Upon Binding to Lipid Surfaces

“…54 High-resolution structural characterization of microtubule-bound proteins, however, remains largely impracticable because of the large size and heterogeneity of such systems. Interestingly, tau has also been reported to interact with lipid membranes in vitro [55][56][57] and in vivo, 17,21 and lipids, fatty acids and detergents can trigger tau aggregation. [58][59][60] We reasoned that such interactions might also involve a structural reorganization within tau.…”