Non-covalent binding of membrane lipids to membrane proteins

Polar lipids and membrane proteins are major components of biological membranes, both cell membranes and membranes of enveloped viruses. How these two classes of membrane components interact with each other to influence the function of biological membranes is a fundamental question that has attracted intense interest since the origins of the field of membrane studies. One of the most powerful ideas that driven the field is the likelihood that lipids bind to membrane proteins at specific sites, modulating protein structure and function. However only relatively recently has high resolution structure determination of membrane proteins progressed to the point of providing atomic level structure of lipid binding sites on membrane proteins. Analysis of X-ray diffraction, electron crystallography and NMR data over 100 specific lipid binding sites on membrane proteins. These data demonstrate tight lipid binding of both phospholipids and cholesterol to membrane proteins. Membrane lipids bind to membrane proteins by their headgroups, or by their acyl chains, or binding is mediated by the entire lipid molecule. When headgroups bind, binding is stabilized by polar interactions between lipid headgroups and the protein. When acyl chains bind, van der Waals effects dominate as the acyl chains adopt conformations that complement particular sites on the rough protein surface. No generally applicable motifs for binding have yet emerged. Previously published biochemical and biophysical data link this binding with function. This Article is Part of a Special Issue Entitled: Membrane Structure and Function: Relevance in the Cell's Physiology, Pathology and Therapy.     

ATP-dependent membrane assembly of F-actin facilitates membrane fusion

We recently established an in vitro assay that monitors the fusion between latex-bead phagosomes and endocytic organelles in the presence of J774 macrophage cytosol (). Here, we show that different reagents affecting the actin cytoskeleton can either inhibit or stimulate this fusion process. Because the membranes of purified phagosomes can assemble F-actin de novo from pure actin with ATP (), we focused here on the ability of membranes to nucleate actin in the presence of J774 cytosolic extracts. For this, we used F-actin sedimentation, pyrene actin assays, and torsional rheometry, a biophysical approach that could provide kinetic information on actin polymerization and gel formation. We make two major conclusions. First, under our standard in vitro conditions (4 mg/ml cytosol and 1 mM ATP), the presence of membranes actively catalyzed the assembly of cytosolic F-actin, which assembled into highly viscoelastic gels. A model is discussed that links these results to how the actin may facilitate fusion. Second, cytosolic actin paradoxically polymerized more under ATP depletion than under high-ATP conditions, even in the absence of membranes; we discuss these data in the context of the well described, large increases in F-actin seen in many cells during ischemia.     

Label-free imaging of membrane potential using membrane electromotility

Electrical activity may cause observable changes in a cell's structure in the absence of exogenous reporter molecules. In this work, we report a low-coherence interferometric microscopy technique that can detect an optical signal correlated with the membrane potential changes in individual mammalian cells without exogenous labels. By measuring milliradian-scale phase shifts in the transmitted light, we can detect changes in the cells' membrane potential. We find that the observed optical signals are due to membrane electromotility, which causes the cells to deform in response to the membrane potential changes. We demonstrate wide-field imaging of the propagation of electrical stimuli in gap-junction-coupled cell networks. Membrane electromotility-induced cell deformation may be useful as a reporter of electrical activity.     

Origin of magnetosome membrane: proteomic analysis of magnetosome membrane and comparison with cytoplasmic membrane

Prokaryotes are known to have evolved one or more unique organelles. Although several hypotheses have been proposed concerning the biogenesis of these intracellular components, the majority of these proposals remains unclear. Magnetotactic bacteria synthesize intracellular magnetosomes that are enclosed by lipid bilayer membranes. From the identification and characterization of several surface and transmembrane magnetosome proteins, we have postulated that magnetosomes are derived from the cytoplasmic membrane (CM). To confirm this hypothesis, a comparative proteomic analysis of the magnetosome membrane (MM) and CM of the magnetotactic bacterium, Magnetospirillum magneticum AMB-1, was undertaken. Based on the whole genome sequence of M. magneticum AMB-1, 78 identified MM proteins were also found to be prevalent in the CM, several of which are related to magnetosome biosynthesis, such as Mms13, which is tightly bound on the magnetite surface. Fatty acid analysis was also conducted, and showed a striking similarity between the CM and MM profiles. These results suggest that the MM is derived from the CM.     

Erythrocyte membrane proteins and membrane skeleton

Considerable advances in the research field of erythrocyte membrane were achieved in the recent two decades. New findings in the structure-function correlation and interactions of erythrocyte membrane proteins have attracted extensive attention. Interesting progress was also made in the molecular pathogenesis of erythrocyte membrane disorders. Advances in the composition, function and interaction of erythrocyte membrane proteins, erythrocyte membrane skeleton, and relevant diseases are briefly described and summarized here on the basis of domestic and world literatures. Translated from Life Science Research, 2005, 9(4): 283–291 [译自: 生命科学研究]     

Erythrocyte membrane proteins and membrane skeleton

Considerable advances in the research field of erythrocyte membrane were achieved in the recent two decades.New findings in the structure-function correlation and interactions of erythrocyte membrane proteins have attracted extensive attention.Interesting progress was also made in the molecular pathogenesis of erythrocyte membrane disorders.Advances in the composition,function and interaction of erythrocyte membrane proteins,erythrocyte membrane skeleton,and relevant diseases are briefly described and summarized here on the basis of domestic and world literatures.     

Interaction of membrane skeletal proteins with membrane lipid domain

The object of this paper is to review briefly the studies on the interaction of red blood cell membrane skeletal proteins and their non-erythroid analogues with lipids in model systems as well as in natural membranes. An important question to be addressed is the physiological significance and possible regulatory molecular mechanisms in which these interactions are engaged.     

Palaeontological evidence of membrane relationship in step-by-step membrane fusion

Studies on membrane fusion in living cells indicate that initiation of membrane fusion is a transient and hard to capture process. Despite previous research, membrane behaviour at this point is still poorly understood. Recent palaeobotanical research has revealed snapshots of membrane fusion in a 15-million-year-old fossil pinaceous cone. To reveal the membrane behaviour during the fusion, we conducted more observations on the same fossil material. Several discernible steps of membrane fusion have been fixed naturally and observed in the fossil material. This observation provides transmission electron microscope (TEM) images of the transient intermediate stage and clearly shows the relationship between membranes. Observing such a transient phenomenon in fossil material implies that the fixing was most likely accomplished quickly by a natural process. The mechanism behind this phenomenon is clearly worthy of further enquiry.     

Characteristics of a membrane reservoir buffering membrane tension.

When membrane-attached beads are pulled vertically by a laser tweezers, a membrane tube of constant diameter (tether) is formed. We found that the force on the bead (tether force) did not depend on tether length over a wide range of tether lengths, which indicates that a previously unidentified reservoir of membrane and not stretch of the plasma membrane provides the tether membrane. Plots of tether force vs. tether length have an initial phase, an elongation phase, and an exponential phase. During the major elongation phase, tether force is constant, buffered by the "membrane reservoir." Finally, there is an abrupt exponential rise in force that brings the tether out of the trap, indicating depletion of the membrane reservoir. In chick embryo fibroblasts and 3T3 fibroblasts, the maximum tether lengths that can be pulled at a velocity of 4 microm/s are 5.1 +/- 0. 3 and 5.0 +/- 0.2 microm, respectively. To examine the importance of the actin cytoskeleton, we treated cells with cytochalasin B or D and found that the tether lengths increased dramatically to 13.8 +/- 0.8 and 12.0 +/- 0.7 microm, respectively. Similarly, treatment of the cells with colchicine and nocodazole results in more than a twofold increase in tether length. We found that elevation of membrane tension (through osmotic pressure, a long-term elevation of tether force, or a number of transitory increases) increased reservoir size over the whole cell. Using a tracking system to hold tether force on the bead constant near its maximal length in the exponential phase, the rate of elongation of the tethers was measured as a function of tether force (membrane tension). The rate of elongation of tethers was linearly dependent on the tether force and reflected an increase in size of the reservoir. Increases in the reservoir caused by tension increases on one side of the cell caused increases in reservoir size on the other side of the cell. Thus, we suggest that cells maintain a plasma membrane reservoir to buffer against changes in membrane tension and that the reservoir is increased with membrane tension or disruption of the cytoskeleton.     

Nucleoside diphosphatase: membrane asymmetry, membrane binding and latency of a peripheral membrane protein

Nucleoside diphosphatase is a peripheral protein of the endoplasmic reticulum in the liver. This review brings together the available information on the properties of the membrane-associated form of the enzyme found in microsomal vesicles. The data are consistent with the view that the enzyme is loosely bound to the inner surface of the vesicles and, therefore, to the luminal surface of the endoplasmic reticulum in vivo.     

Detergent-resistant membrane subfractions containing proteins of plasma membrane, mitochondrial, and internal membrane origins

HEK293 cell detergent-resistant membranes (DRMs) isolated by the standard homogenization protocol employing a Teflon pestle homogenizer yielded a prominent opaque band at approximately 16% sucrose upon density gradient ultracentrifugation. In contrast, cell disruption using a ground glass tissue homogenizer generated three distinct DRM populations migrating at approximately 10%, 14%, and 20% sucrose, named DRM subfractions A, B, and C, respectively. Separation of the DRM subfractions by mechanical disruption suggested that they are physically associated within the cellular environment, but can be dissociated by shear forces generated during vigorous homogenization. All three DRM subfractions possessed cholesterol and ganglioside G_M1, but differed in protein composition. Subfraction A was enriched in flotillin-1 and contained little caveolin-1. In contrast, subfractions B and C were enriched in caveolin-1. Subfraction C contained several mitochondrial membrane proteins, including mitofilin and porins. Only subfraction B appeared to contain significant amounts of plasma membrane-associated proteins, as revealed by cell surface labeling studies. A similar distribution of DRM subfractions, as assessed by separation of flotillin-1 and caveolin-1 immunoreactivities, was observed in CHO cells, in 3T3-L1 adipocytes, and in HEK293 cells lysed in detergent-free carbonate. Teflon pestle homogenization of HEK293 cells in the presence of the actin-disrupting agent latrunculin B generated DRM subfractions A–C. The microtubule-disrupting agent vinblastine did not facilitate DRM subfraction separation, and DRMs prepared from fibroblasts of vimentin-null mice were present as a single major band on sucrose gradients, unless pre-treated with latrunculin B. These results suggest that the DRM subfractions are interconnected by the actin cytoskeleton, and not by microtubes or vimentin intermediate filaments. The subfractions described may prove useful in studying discrete protein populations associated with detergent-resistant membranes, and their potential interactions in cell signaling.     

Palaeontological evidence for membrane fusion between a unit membrane and a half-unit membrane

Membrane fusion is of fundamental importance for many biological processes and has been a topic of intensive research in past decades with several models being proposed for it. Fossils had previously not been considered relevant to studies on membrane fusion. But here two different membrane fusion patterns are reported in the same well-preserved fossil plant from the Miocene (15–20 million years old) at Clarkia, Idaho, US. Scanning electron microscope, transmission electron microscope, and traditional studies reveal the vesicles in various states (even transient semi-fusion) of membrane fusion, and thus shed new light on their membrane structure and fusion during exocytoses. The new evidence suggests that vesicles in plant cells may have not only a unit membrane but also a half-unit membrane, and that a previously overlooked membrane fusion pattern exists in plant cells. This unexpected result from an unexpected material not only marks the first evidence of on-going physiological activities in fossil plants, but also raises questions on membrane fusion in recent plants.     

Paleontological evidence for membrane fusion between a unit membrane and a half-unit membrane

Membrane fusion is of fundamental importance for many biological processes and has been a topic of intensive research in past decades with several models being proposed for it. Fossils had previously not been considered relevant to studies on membrane fusion. But here two different membrane fusion patterns are reported in the same well-preserved fossil plant from the Miocene (15-20 million years old) at Clarkia, Idaho, US. Scanning electron microscope, transmission electron microscope, and traditional studies reveal the vesicles in various states (even transient semi-fusion) of membrane fusion, and thus shed new light on their membrane structure and fusion during exocytoses. The new evidence suggests that vesicles in plant cells may have not only a unit membrane but also a half-unit membrane, and that a previously overlooked membrane fusion pattern exists in plant cells. This unexpected result from an unexpected material not only marks the first evidence of on-going physiological activities in fossil plants, but also raises questions on membrane fusion in recent plants.     

Modulation of membrane dynamics and cell motility by membrane tension

The plasma membrane of most cells is drawn tightly over the cytoskeleton of the cell, resulting in a significant tension being developed in the membrane. The tension in the membrane can be calculated from the force required to separate it from the cytoskeleton; and the force itself can be measured rapidly by using laser tweezers. Recent observations indicate that decreasing membrane tension stimulates endocytosis and increasing tension stimulates secretion. Thus, membrane tension provides a simple physical mechanism to control the area of the plasma membrane. Here, we speculate that tension is a global parameter that the cell uses to control physically plasma membrane dynamics, cell shape and cell motility.