Dynamics of Dynamin during Clathrin Mediated Endocytosis in PC12 Cells

Members of the dynamin super-family of GTPases are involved in disparate cellular pathways. Dynamin1 and dynamin2 have been implicated in clathrin-mediated endocytosis. While some models suggest that dynamin functions specifically at the point of vesicle fission, evidence also exists for a role prior to fission during vesicle formation and it is unknown if there is a role for dynamin after vesicle fission. Although dynamin2 is ubiquitously expressed, dynamin1 is restricted to the nervous system. These two structurally similar endocytic accessory proteins have not been studied in cells that endogenously express both.The present study quantitatively assesses the dynamics of dynamin1 and dynamin2 during clathrin-mediated endocytosis in PC12 cells, which endogenously express both proteins. Both dynamin isoforms co-localized with clathrin and showed sharp increases in fluorescence intensity immediately prior to internalization of the nascent clathrin-coated vesicle. The fluorescence intensity of both proteins then decreased with two time constants. The slower time constant closely matched the time constant for the decrease of clathrin intensity and likely represents vesicle movement away from the membrane. The faster rate may reflect release of dynamin at the neck of nascent vesicle following GTP hydrolysis.This study analyses the role of dynamin in clathrin-mediated endocytosis in a model for cellular neuroscience and these results may provide direct evidence for the existence of two populations of dynamin associated with nascent clathrin-coated vesicles

Arene oxidation with malonoyl peroxides

Malonoyl peroxide 7, prepared in a single step from the commercially available diacid, is an effective reagent for the oxidation of aromatics. Reaction of an arene with peroxide 7 at room temperature leads to the corresponding protected phenol which can be unmasked by aminolysis. An ionic mechanism consistent with the experimental findings and supported by isotopic labeling, Hammett analysis, EPR investigations and reactivity profile studies is proposed

RhoJ interacts with the GIT-PIX complex and regulates focal adhesion disassembly

RhoJ is a Rho GTPase expressed in endothelial cells and tumour cells, which regulates cell motility, invasion, endothelial tube formation and focal adhesion numbers. This study aimed to further delineate the molecular function of RhoJ. Using timelapse microscopy RhoJ was found to regulate focal adhesion disassembly; small interfering RNA (siRNA)-mediated knockdown of RhoJ increased focal adhesion disassembly time, whereas expression of an active mutant (daRhoJ) decreased it. Furthermore, daRhoJ co-precipitated with the GIT–PIX complex, a regulator of focal adhesion disassembly. An interaction between daRhoJ and GIT1 was confirmed using yeast two-hybrid experiments, and this depended on the Spa homology domain of GIT1. GIT1, GIT2, β-PIX (also known as ARHGEF7) and RhoJ all colocalised in focal adhesions and depended on each other for their recruitment to focal adhesions. Functionally, the GIT–PIX complex regulated endothelial tube formation, with knockdown of both GIT1 and GIT2, or β-PIX phenocopying RhoJ knockdown. RhoJ-knockout mice showed reduced tumour growth and diminished tumour vessel density, identifying a role for RhoJ in mediating tumour angiogenesis. These studies give new insight into the molecular function of RhoJ in regulating cell motility and tumour vessel formation

PIP5KIβ Selectively Modulates Apical Endocytosis in Polarized Renal Epithelial Cells

Localized synthesis of phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2] at clathrin coated pits (CCPs) is crucial for the recruitment of adaptors and other components of the internalization machinery, as well as for regulating actin dynamics during endocytosis. PtdIns(4,5)P2 is synthesized from phosphatidylinositol 4-phosphate by any of three phosphatidylinositol 5-kinase type I (PIP5KI) isoforms (α, β or γ). PIP5KIβ localizes almost exclusively to the apical surface in polarized mouse cortical collecting duct cells, whereas the other isoforms have a less polarized membrane distribution. We therefore investigated the role of PIP5KI isoforms in endocytosis at the apical and basolateral domains. Endocytosis at the apical surface is known to occur more slowly than at the basolateral surface. Apical endocytosis was selectively stimulated by overexpression of PIP5KIβ whereas the other isoforms had no effect on either apical or basolateral internalization. We found no difference in the affinity for PtdIns(4,5)P2-containing liposomes of the PtdIns(4,5)P2 binding domains of epsin and Dab2, consistent with a generic effect of elevated PtdIns(4,5)P2 on apical endocytosis. Additionally, using apical total internal reflection fluorescence imaging and electron microscopy we found that cells overexpressing PIP5KIβ have fewer apical CCPs but more internalized coated structures than control cells, consistent with enhanced maturation of apical CCPs. Together, our results suggest that synthesis of PtdIns(4,5)P2 mediated by PIP5KIβ is rate limiting for apical but not basolateral endocytosis in polarized kidney cells. PtdIns(4,5)P2 may be required to overcome specific structural constraints that limit the efficiency of apical endocytosis. © 2013 Szalinski et al

Entanglement entropy in aperiodic singlet phases

We study the average entanglement entropy of blocks of contiguous spins in aperiodic XXZ chains which possess an aperiodic singlet phase at least in a certain limit of the coupling ratios. In this phase, where the ground state constructed by a real space renormalization group method, consists (asymptotically) of independent singlet pairs, the average entanglement entropy is found to be a piecewise linear function of the block size. The enveloping curve of this function is growing logarithmically with the block size, with an effective central charge in front of the logarithm which is characteristic for the underlying aperiodic sequence. The aperiodic sequence producing the largest effective central charge is identified, and the latter is found to exceed the central charge of the corresponding homogeneous model. For marginal aperiodic modulations, numerical investigations performed for the XX model show a logarithmic dependence, as well, with an effective central charge varying continuously with the coupling ratio.Comment: 18 pages, 9 figure

Quantum Impurity Entanglement

Entanglement in J_1-J_2, S=1/2 quantum spin chains with an impurity is studied using analytic methods as well as large scale numerical density matrix renormalization group methods. The entanglement is investigated in terms of the von Neumann entropy, S=-Tr rho_A log rho_A, for a sub-system A of size r of the chain. The impurity contribution to the uniform part of the entanglement entropy, S_{imp}, is defined and analyzed in detail in both the gapless, J_2 <= J_2^c, as well as the dimerized phase, J_2>J_2^c, of the model. This quantum impurity model is in the universality class of the single channel Kondo model and it is shown that in a quite universal way the presence of the impurity in the gapless phase, J_2 <= J_2^c, gives rise to a large length scale, xi_K, associated with the screening of the impurity, the size of the Kondo screening cloud. The universality of Kondo physics then implies scaling of the form S_{imp}(r/xi_K,r/R) for a system of size R. Numerical results are presented clearly demonstrating this scaling. At the critical point, J_2^c, an analytic Fermi liquid picture is developed and analytic results are obtained both at T=0 and T>0. In the dimerized phase an appealing picure of the entanglement is developed in terms of a thin soliton (TS) ansatz and the notions of impurity valence bonds (IVB) and single particle entanglement (SPE) are introduced. The TS-ansatz permits a variational calculation of the complete entanglement in the dimerized phase that appears to be exact in the thermodynamic limit at the Majumdar-Ghosh point, J_2=J_1/2, and surprisingly precise even close to the critical point J_2^c. In appendices the relation between the finite temperature entanglement entropy, S(T), and the thermal entropy, S_{th}(T), is discussed and and calculated at the MG-point using the TS-ansatz.Comment: 62 pages, 27 figures, JSTAT macro

Community Mental Health Centers as Human Service Organizations

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/67333/2/10.1177_000276428502800506.pd

In-depth exploration of the photophysics of a trinuclear palladium complex

Contains fulltext : 128388.pdf (publisher's version ) (Open Access