Because endocytic protein patches contain a number of other membrane proteins, we use a conservative estimate of a 10nm center-to-center spacing, corresponding to a channel densityof 1012cm2

Because endocytic protein patches contain a number of other membrane proteins, we use a conservative estimate of a 10nm center-to-center spacing, corresponding to a channel densityof 1012cm2. plants, where the mechanisms that allow endocytosis to proceed despite high turgor pressure are largely unknown. == Introduction == Clathrin-mediated endocytosis (CME) contributes to numerous biological processes, such as cell growth, viral invasion, and neural signaling (1). It involves the inward bending of a portion of the lipid bilayer, which subsequently pinches off to form a vesicle that moves into Rabbit Polyclonal to CATZ (Cleaved-Leu62) the cytoplasm. CME has been studied extensively in yeast, due to the ease of genetic manipulation and fluorescent labeling. In yeast, CME involves actin polymerization as well as clathrin and BAR-domain proteins, which help drive invagination through a nonzero preferred curvature. It is believed (2, 3, 4, 5) that growing actin filaments exert pushing forces on the membrane, which drive the actin network into the cytoplasm. This motion of the actin network exerts A-1155463 pulling forces to drive invagination, through a coupling via the adaptor protein Sla2. Recent theoretical work A-1155463 on CME in yeast has treated the mechanics of forces generated by actin polymerization, as well as the intrinsic curvature of the clathrin coat that accumulates on the cell membrane (5, 6, 7, 8). To date, however , it is not known how actin and other curvature-generating proteins can produce enough force to overcome the large turgor pressure in yeast (9, 10). Here we explore the hypothesis that increasing the membrane permeability at the endocytic site can locally reduce the turgor pressure and thus facilitate endocytosis. The turgor pressure (10) pushes the plasma membrane against the cell wall and thus helps the cell maintain its shape and rigidity. It is generated through the larger internal concentration of glycerol, the main osmolyte in yeast, relative to the outside. The magnitude of is determined by the requirement that the chemical potential of the solvent (water) be constant across the membrane. For a dilute solution, the solvent chemical potential is(11), wherePis pressure; Tis temperature; andCandare the solute and solvent concentrations, respectively. The chemical potential varies with pressure according to(11), whereis Avogadros number andis given in molar units. Thus the constancy ofimplies (here we have ignored possible changes in the concentrations resulting from pressure variations, because pressure changes in cells are much smaller than the gigapascal bulk modulus of water) that between any two points, If the points are on opposite sides of the membrane, this implies that the turgor pressure is whereis the concentration difference across the membrane. In the scenario treated here, where a permeable patch leads to large concentration gradients inside the cell, Eq. 1 implies corresponding internal gradients of the pressure. Thus pressure does not equilibrate, even at long times, as long as the concentration gradient is maintained by the continuing production of osmolyte inside the cell. The pressure gradient does not cause macroscopic fluid flow, because there are additional forces on water molecules due to the osmolyte concentration gradient. The situation is analogous to that of water in a swimming pool. The pressure is A-1155463 greater at the bottom, but there is no macroscopic flow because the water molecules at the bottom have lower gravitational potential A-1155463 energy. In this case, the gravitational potential energy variation is replaced by variation in the free energy of water molecules resulting from the inhomogeneous osmolyte concentration. Can established mechanisms, including curvature-generating proteins and actin polymerization, provide enough force to overcome the turgor pressure barrier? The driving forces from curvature-generating proteins and actin, along with opposing forces from surface tension and turgor pressure, are encapsulated in the Helfrich membrane deformation energy: Hereis the bending modulus, is the turgor pressure, is an element of membrane area, Vis volume of the invagination, His the mean curvature, is the spontaneous curvature, is the surface tension, zis the inward displacement of the membrane, andis the pulling force density from actin. We estimate whether it is energetically favorable for a hemispherical invagination of radius of(12) to form in the presence of accepted values of the turgor pressure. We take(13), and = 0. 6 MPa as an average over a number of measurements (14, 15, 16, 17, 18, 19). Then the stabilizing contribution from the curvature-generating proteins is(the negative of the curvature energy of the flattened membrane), while the opposing contribution from the turgor pressure is, leavingto be supplied by actin pulling forces. It is unlikely that actin polymerization can.