The influence of cholesterol (CHOL) level on integrin sequestration in raft-mimicking lipid mixtures forming coexisting liquid-ordered (l) and liquid-disordered (l) lipid domains is investigated using complementary, single-molecule-sensitive, confocal detection methods. Systematic analysis of membrane protein distribution in such a model membrane environment demonstrates that variation of CHOL level has a profound influence on l-l sequestration of integrins, thereby exhibiting overall l preference in the absence of ligands and l affinity upon vitronectin addition. Accompanying photon-counting histogram analysis of integrins in the different model membrane mixtures shows that the observed changes of integrin sequestration in response to variations of membrane CHOL level are not associated with altering integrin oligomerization states. Instead, our experiments suggest that the strong CHOL dependence of integrin sequestration can be attributed to CHOL-mediated changes of lipid packing and bilayer thickness in coexisting l and l domains, highlighting the significance of a biophysical mechanism of CHOL-mediated regulation of integrin sequestration. We envision that this model membrane study may help clarify the influence of CHOL in integrin functionality in plasma membranes, thus providing further insight into the role of lipid heterogeneities in membrane protein distribution and function in a cellular membrane environment.

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An interpenetrating phase composite is made by injection molding thermoplastic polymers into the voids of open-cell aluminum foam. Two types of polypropylene and an acetyl were mechanically introduced into the open cells of a Duocel® aluminum foam. Prior experimental work revealed that the combination of the polymer and the metal foam yields a hybrid that is stiffer than the polymer alone but has a reduced tensile strength. A finite element model using a tetrakaidecahedral unit cell is used to model the metal foam ligaments with the polymer occupying the remaining space. The geometric model as well as the interface between the two materials were validated against the experimental results. The resulting conclusions are that the aluminum ligaments oriented along the load direction cause an increase in stiffness but ligaments oriented laterally cause stress concentration that yield lower strength. The finite element model is used to give both qualitative and quantitative explanations of the physics of the interrelations between the metal foam and the polymer.

Quite surprisingly, N-and the C-termini remained in close proximity at high denaturant concentrations for ca. 40% of the conformations, suggesting that DrkN-SH3behaves at least partially like a disordered circular chain.

Thymidine block that arrests cells at the border between G1 and S phases. Critical temperatures were elevated in GPMVs isolated from cells in cell cycle phases that precede cell division (G2 and M) compared to other stages (G1 and S). In unsynchronized cells, critical temperatures were found to be inversely proportional to cell density, suggesting that contact inhibition and associated arrest of cell growth results in lower plasma membrane critical temperatures. Lower critical temperatures were also observed when growth was arrested through overnight serum starvation, and elevated critical temperatures were restored 24h after the addition of serum containing medium. Transition temperatures are also lowered in GPMVs prepared from cells undergoing apoptosis through the application of TRAIL, and vesicles contain a more rigid liquid-ordered or gel phase at low temperature. These results are in agreement with past studies that have indicated that plasma membrane composition varies within the cell cycle. Since GPMV critical temperatures are hypothesized to reflect on the magnitude of lipid-mediated heterogeneity in intact cells, these results suggest that membrane heterogeneity is greatest in cells undergoing rapid growth and cell division and is suppressed in cells under low growth conditions. Nano-membrane domains are hypothesized to play an integral role in many cell signaling pathways. Their transient nature and biocomplexity underlies a myriad of fundamental questions about lipid-lipid and lipid-protein interactions and their roles in cellular functions. As a result, there is a need for innovative approaches for understanding different biophysical aspects of membrane assemblies and their underlying, multiscale dynamics. Here, we integrate dynamic holographic optical trapping (HOT) and fluorescence imaging with fluorescence correlation spectroscopy (FCS) to characterize membrane domain nucleation in biomimetic planar supported bilayers. The dynamic HOT system allows for the creation of multiple traps from a single light source, each of which can be controlled individually in real time. Silica microspheres are being trapped into arbitrary patterns for system optimization. Receptor-bound microspheres associated with nano-domains in planar supported bilayers act as handles for dynamic HOT manipulation. Our hypothesis is that by trapping multiple microsphere-bound receptors, the associated heterogeneous lipid domains will nucleate a larger domain upon interaction in a manner that depends on the lipid type, cholesterol and protein content. Fluorescence imaging is used to visualize lipid domain formation, and subsequent lateral diffusion of lipid species will be measured with FCS as a function of trap-induced confinement. These results will ultimately lead to new insights into domain formation in membranes. It is now well established that the cytoplasm and plasma membrane of cells are characterized by high concentrations of proteins. Consequently, macromolecular crowding and confinement effects are believed to play impor...

Membrane-targeting domains play crucial roles in the association of signaling molecules to the plasma membrane. For most peripheral proteins, the proteinto-membrane interaction is transient; but after proteins dissociate from the membrane they are observed to rebind following a brief excursion in the bulk solution. These membrane hops can have broad implications on protein dynamics. We study the diffusion of membrane-targeting domains using single-molecule tracking in supported lipid bilayers. The ensemble-averaged mean square displacement (MSD) exhibits superdiffusive behavior. However, the time-averaged MSD of individual trajectories is found to be linear with respect to lag time, as in Brownian diffusion. These observations are explained in terms of bulk excursions that introduce jumps with a heavy-tail distribution, rapidly increasing the area explored. Furthermore, this type of motion is characterized by a pronounced scattering in the time-averaged MSD of individual trajectories.

Our focus is the effects of cholesterol on the activity of the Adenosine A 2a receptor (A 2a R), a class A GPCR. Membrane cholesterol concentrations were varied in Human embryonic kidney (HEK-293) cells by the use of methylb-cyclodextrin (MbCD), which is capable of capturing cholesterol in its inner cavity (Pucadyil et al., 2006). We tested the role that bulk cholesterol depletion played in expression, ligand binding and downstream synthesis of cyclic AMP (cAMP) in HEK-293 cells. Continuing work from our lab will study the specific interaction between cholesterol and A 2a R by making point mutations in the residues of the CCM. In A 2a R those residues include Tyr43(2.41), Ser47(2.45), Lys122(4.43), Ile125(4.46) and Trp129(4.50) (Lee et al., 2013). Our lab will investigate whether these point mutations effect the interaction between cholesterol and purified receptor, as well as the receptors activity in both yeast and mammalian cell model systems.

To know more about the hydrodynamic environments either in extreme conditions or in normal conditions, numerical simulation becomes more important due to insufficient field data. For large open sea, numerical models based on momentum balanced equation as mild slope equation or Boussinesq equation seems to be impractical. The third generation spectral numerical model was used in this discussion WAVEWATCH and SWAN to forecast wave conditions. Each model itself was nested and offered boundary conditions for smaller scale computation. WAVEWATCH provided extern boundary conditions for SWAN model computations. So wave parameter of different scale could be described so to offer wave parameters for engineering concerning. At the same time, some characteristics of third generation spectral wave model were depicted. Input winds were from NCEP analyzed data and QSCAT data respectively. The comparisons of computation with these data would show the spectral model characteristics of typical dependence on the wind condition. The output of WAVEWATCH under cyclone was also discussed in the paper.