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Vesicle transport in eukaryotic cells is regulated by SNARE proteins, which play an intimate role in regulating the specificity of vesicle fusion between discrete intracellular organelles. In the present study we investigated the function and plasticity of v-SNAREs in insulin-regulated GLUT4 trafficking in adipocytes. Using a combination of knockout mice,(More)
The insulin-stimulated trafficking of GLUT4 to the plasma membrane in muscle and fat tissue constitutes a central process in blood glucose homeostasis. The tethering, docking, and fusion of GLUT4 vesicles with the plasma membrane (PM) represent the most distal steps in this pathway and have been recently shown to be key targets of insulin action. However,(More)
This paper presents a novel computer vision system for automated identification of vesicle-plasma membrane fusion events in image sequences obtained from Total Internal Reflection Fluorescence (TIRF) microscopes. Identification of such events is important in order to better understand the process of exocytosis in cells. Manual analysis of thousands of(More)
In type 2 diabetes, pancreatic beta cells fail to secrete sufficient insulin to overcome peripheral insulin resistance. Intracellular lipid accumulation contributes to beta cell failure through poorly defined mechanisms. Here we report a role for the lipid-regulated protein kinase C isoform PKCepsilon in beta cell dysfunction. Deletion of PKCepsilon(More)
The protein kinase Akt mediates several metabolic and mitogenic effects of insulin, whereas activation of protein kinase C (PKC) isoforms has been implicated in the inhibition of insulin action. We have previously shown that both PKC and PKCepsilon are activated in skeletal muscle of insulin-resistant high fat-fed rats, and to identify potential substrates(More)
The adipocyte is a key regulator of mammalian metabolism. To advance our understanding of this important cell, we have used quantitative proteomics to define the protein composition of the adipocyte plasma membrane (PM) in the presence and absence of insulin. Using this approach, we have identified a high confidence list of 486 PM proteins, 52 of which(More)
The regulated trafficking or exocytosis of cargo-containing vesicles to the cell surface is fundamental to all cells. By coupling the technology of fluorescently tagged fusion proteins with total internal reflection fluorescence microscopy (TIRFM), it is possible to achieve the high spatio-temporal resolution required to study the dynamics of sub-plasma(More)
AIMS/HYPOTHESIS Insulin resistance in skeletal muscle is strongly associated with lipid oversupply, but the intracellular metabolites and underlying mechanisms are unclear. We therefore sought to identify the lipid intermediates through which the common unsaturated fatty acid linoleate causes defects in IRS-1 signalling in L6 myotubes and mouse skeletal(More)
OBJECTIVE—Insufficient insulin secretion is a hallmark of type 2 diabetes, and exposure of ␤-cells to elevated lipid levels (lipotoxicity) contributes to secretory dysfunction. Functional ablation of protein kinase C ε (PKCε) has been shown to improve glucose homeostasis in models of type 2 diabetes and, in particular, to enhance glucose-stimulated insulin(More)
The Niemann-Pick disease, type C1 (NPC1) gene encodes a transmembrane protein involved in cholesterol efflux from the lysosome. SNPs within NPC1 have been associated with obesity and type 2 diabetes, and mice heterozygous or null for NPC1 are insulin resistant. However, the molecular mechanism underpinning this association is currently undefined. This study(More)