scholarly journals SEC16A is a RAB10 effector required for insulin-stimulated GLUT4 trafficking in adipocytes

2016 ◽  
Vol 214 (1) ◽  
pp. 61-76 ◽  
Author(s):  
Joanne Bruno ◽  
Alexandria Brumfield ◽  
Natasha Chaudhary ◽  
David Iaea ◽  
Timothy E. McGraw
Keyword(s):  
Glucose Transporter ◽  
Whole Body ◽  
Site Formation ◽  
Coated Vesicle ◽  
Coat Proteins ◽  
Insulin Stimulation ◽  
Exit Site ◽  
Recycling Endosome ◽  
Transport Vesicles ◽  
Intracellular Compartments

RAB10 is a regulator of insulin-stimulated translocation of the GLUT4 glucose transporter to the plasma membrane (PM) of adipocytes, which is essential for whole-body glucose homeostasis. We establish SEC16A as a novel RAB10 effector in this process. Colocalization of SEC16A with RAB10 is augmented by insulin stimulation, and SEC16A knockdown attenuates insulin-induced GLUT4 translocation, phenocopying RAB10 knockdown. We show that SEC16A and RAB10 promote insulin-stimulated mobilization of GLUT4 from a perinuclear recycling endosome/TGN compartment. We propose RAB10–SEC16A functions to accelerate formation of the vesicles that ferry GLUT4 to the PM during insulin stimulation. Because GLUT4 continually cycles between the PM and intracellular compartments, the maintenance of elevated cell-surface GLUT4 in the presence of insulin requires accelerated biogenesis of the specialized GLUT4 transport vesicles. The function of SEC16A in GLUT4 trafficking is independent of its previously characterized activity in ER exit site formation and therefore independent of canonical COPII-coated vesicle function. However, our data support a role for SEC23A, but not the other COPII components SEC13, SEC23B, and SEC31, in the insulin stimulation of GLUT4 trafficking, suggesting that vesicles derived from subcomplexes of COPII coat proteins have a role in the specialized trafficking of GLUT4.

scholarly journals Insulin Stimulation of GLUT4 Exocytosis, but Not Its Inhibition of Endocytosis, Is Dependent on RabGAP AS160

2004 ◽  
Vol 15 (10) ◽  
pp. 4406-4415 ◽  
Author(s):  
Anja Zeigerer ◽  
Mary Kate McBrayer ◽  
Timothy E. McGraw
Keyword(s):  
Plasma Membrane ◽  
Blood Glucose ◽  
Insulin Signaling ◽  
Glucose Transporter ◽  
Whole Body ◽  
Insulin Stimulation ◽  
Molecular Events ◽  
Intracellular Compartments ◽  
Blood Glucose Homeostasis ◽  
Stimulation Of

Insulin maintains whole body blood glucose homeostasis, in part, by regulating the amount of the GLUT4 glucose transporter on the cell surface of fat and muscle cells. Insulin induces the redistribution of GLUT4 from intracellular compartments to the plasma membrane, by stimulating a large increase in exocytosis and a smaller inhibition of endocytosis. A considerable amount is known about the molecular events of insulin signaling and the complex itinerary of GLUT4 trafficking, but less is known about how insulin signaling is transmitted to GLUT4 trafficking. Here, we show that the AS160 RabGAP, a substrate of Akt, is required for insulin stimulation of GLUT4 exocytosis. A dominant-inhibitory mutant of AS160 blocks insulin stimulation of exocytosis at a step before the fusion of GLUT4-containing vesicles with the plasma membrane. This mutant, however, does not block insulin-induced inhibition of GLUT4 endocytosis. These data support a model in which insulin signaling to the exocytosis machinery (AS160 dependent) is distinct from its signaling to the internalization machinery (AS160 independent).

scholarly journals Specialized sorting of GLUT4 and its recruitment to the cell surface are independently regulated by distinct Rabs

2013 ◽  
Vol 24 (16) ◽  
pp. 2544-2557 ◽  
Author(s):  
L. Amanda Sadacca ◽  
Joanne Bruno ◽  
Jennifer Wen ◽  
Wenyong Xiong ◽  
Timothy E. McGraw
Keyword(s):  
Plasma Membrane ◽  
Glucose Uptake ◽  
Glucose Transporter ◽  
Receptor Activation ◽  
Glut4 Translocation ◽  
Insulin Stimulation ◽  
Transport Vesicles ◽  
Glucose Transporter Glut4

Adipocyte glucose uptake in response to insulin is essential for physiological glucose homeostasis: stimulation of adipocytes with insulin results in insertion of the glucose transporter GLUT4 into the plasma membrane and subsequent glucose uptake. Here we establish that RAB10 and RAB14 are key regulators of GLUT4 trafficking that function at independent, sequential steps of GLUT4 translocation. RAB14 functions upstream of RAB10 in the sorting of GLUT4 to the specialized transport vesicles that ferry GLUT4 to the plasma membrane. RAB10 and its GTPase-activating protein (GAP) AS160 comprise the principal signaling module downstream of insulin receptor activation that regulates the accumulation of GLUT4 transport vesicles at the plasma membrane. Although both RAB10 and RAB14 are regulated by the GAP activity of AS160 in vitro, only RAB10 is under the control of AS160 in vivo. Insulin regulation of the pool of RAB10 required for GLUT4 translocation occurs through regulation of AS160, since activation of RAB10 by DENND4C, its GTP exchange factor, does not require insulin stimulation.

scholarly journals Multivariate proteomic profiling identifies novel accessory proteins of coated vesicles

2012 ◽  
Vol 197 (1) ◽  
pp. 141-160 ◽  
Author(s):  
Georg H.H. Borner ◽  
Robin Antrobus ◽  
Jennifer Hirst ◽  
Gary S. Bhumbra ◽  
Patrycja Kozik ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Isotope Labeling ◽  
Principal Component ◽  
Coated Vesicles ◽  
Coated Vesicle ◽  
Coat Proteins ◽  
Proteomic Profiling ◽  
Transport Vesicles ◽  
Associated Proteins ◽  
Sirna Knockdown

Despite recent advances in mass spectrometry, proteomic characterization of transport vesicles remains challenging. Here, we describe a multivariate proteomics approach to analyzing clathrin-coated vesicles (CCVs) from HeLa cells. siRNA knockdown of coat components and different fractionation protocols were used to obtain modified coated vesicle-enriched fractions, which were compared by stable isotope labeling of amino acids in cell culture (SILAC)-based quantitative mass spectrometry. 10 datasets were combined through principal component analysis into a “profiling” cluster analysis. Overall, 136 CCV-associated proteins were predicted, including 36 new proteins. The method identified >93% of established CCV coat proteins and assigned >91% correctly to intracellular or endocytic CCVs. Furthermore, the profiling analysis extends to less well characterized types of coated vesicles, and we identify and characterize the first AP-4 accessory protein, which we have named tepsin. Finally, our data explain how sequestration of TACC3 in cytosolic clathrin cages causes the severe mitotic defects observed in auxilin-depleted cells. The profiling approach can be adapted to address related cell and systems biological questions.

Assembly of COPI and COPII Vesicular Coat Proteins on Membranes

2018 ◽  
Vol 47 (1) ◽  
pp. 63-83 ◽  
Author(s):  
Julien Béthune ◽  
Felix T. Wieland
Keyword(s):  
Electron Tomography ◽  
Coat Proteins ◽  
Structural Elements ◽  
X Ray Crystallography ◽  
Transport Vesicles ◽  
Cryo Electron Tomography ◽  
Functional Implications ◽  
Cargo Proteins ◽  
Intracellular Compartments ◽  
Proteins And Peptides

In eukaryotes, distinct transport vesicles functionally connect various intracellular compartments. These carriers mediate transport of membranes for the biogenesis and maintenance of organelles, secretion of cargo proteins and peptides, and uptake of cargo into the cell. Transport vesicles have distinct protein coats that assemble on a donor membrane where they can select cargo and curve the membrane to form a bud. A multitude of structural elements of coat proteins have been solved by X-ray crystallography. More recently, the architectures of the COPI and COPII coats were elucidated in context with their membrane by cryo-electron tomography. Here, we describe insights gained from the structures of these two coat lattices and discuss the resulting functional implications.

scholarly journals Insulin Increases Cell Surface GLUT4 Levels by Dose Dependently Discharging GLUT4 into a Cell Surface Recycling Pathway

2004 ◽  
Vol 24 (14) ◽  
pp. 6456-6466 ◽  
Author(s):  
Roland Govers ◽  
Adelle C. F. Coster ◽  
David E. James
Keyword(s):  
Plasma Membrane ◽  
Cell Surface ◽  
Glucose Transporter ◽  
Insulin Dose ◽  
Cell Types ◽  
Insulin Stimulation ◽  
Intracellular Compartments ◽  
A Cell ◽  
Recycling Pathway ◽  
Glucose Transporter Glut4

ABSTRACT The insulin-responsive glucose transporter GLUT4 plays an essential role in glucose homeostasis. A novel assay was used to study GLUT4 trafficking in 3T3-L1 fibroblasts/preadipocytes and adipocytes. Whereas insulin stimulated GLUT4 translocation to the plasma membrane in both cell types, in nonstimulated fibroblasts GLUT4 readily cycled between endosomes and the plasma membrane, while this was not the case in adipocytes. This efficient retention in basal adipocytes was mediated in part by a C-terminal targeting motif in GLUT4. Insulin caused a sevenfold increase in the amount of GLUT4 molecules present in a trafficking cycle that included the plasma membrane. Strikingly, the magnitude of this increase correlated with the insulin dose, indicating that the insulin-induced appearance of GLUT4 at the plasma membrane cannot be explained solely by a kinetic change in the recycling of a fixed intracellular GLUT4 pool. These data are consistent with a model in which GLUT4 is present in a storage compartment, from where it is released in a graded or quantal manner upon insulin stimulation and in which released GLUT4 continuously cycles between intracellular compartments and the cell surface independently of the nonreleased pool.

Turning Signals On and Off: GLUT4 Traffic in the Insulin-Signaling Highway

Physiology ◽  
2005 ◽  
Vol 20 (4) ◽  
pp. 271-284 ◽  
Author(s):  
Farah S. L. Thong ◽  
Chandrasagar B. Dugani ◽  
Amira Klip
Keyword(s):  
Skeletal Muscle ◽  
Insulin Signaling ◽  
Glucose Transporter ◽  
Round Trip ◽  
Insulin Stimulation ◽  
Adipose Tissues ◽  
Intracellular Compartments ◽  
Glucose Transporter Glut4 ◽  
Molecular Signals ◽  
Stimulation Of

Insulin stimulation of glucose uptake into skeletal muscle and adipose tissues is achieved by accelerating glucose transporter GLUT4 exocytosis from intracellular compartments to the plasma membrane and minimally reducing its endocytosis. The round trip of GLUT4 is intricately regulated by diverse signaling molecules impinging on specific compartments. Here we highlight the key molecular signals that are turned on and off by insulin to accomplish this task.

A new degree of complexi(n)ty in the regulation of GLUT4 trafficking

2021 ◽  
Vol 478 (7) ◽  
pp. 1315-1319
Author(s):  
Luc Bertrand ◽  
Marine De Loof ◽  
Christophe Beauloye ◽  
Sandrine Horman ◽  
Laurent Bultot
Keyword(s):  
Glucose Transporter ◽  
Whole Body ◽  
Glucose Transporter 4 ◽  
Insulin Stimulation ◽  
Cell Plasma Membrane ◽  
Cell Plasma ◽  
Challenges And Opportunities ◽  
Future Challenges ◽  
Whole Body Metabolism

Loss of the insulin-stimulated glucose uptake in muscle is a crucial event participating in the defect of whole-body metabolism in type 2 diabetes. Therefore, identification by Pavarotti et al. (Biochem. J (2021) 478 (2): 407–422) of complexin-2 as an important contributor to glucose transporter 4 (GLUT4) translocation to muscle cell plasma membrane upon insulin stimulation is essential. The present commentary discusses the biological importance of the findings and proposes future challenges and opportunities.

scholarly journals Expression and compartmentalization of caveolin in adipose cells: coordinate regulation with and structural segregation from GLUT4.

1995 ◽  
Vol 129 (4) ◽  
pp. 999-1006 ◽  
Author(s):  
K V Kandror ◽  
J M Stephens ◽  
P F Pilch
Keyword(s):  
Cell Surface ◽  
Glucose Transporter ◽  
Buoyant Density ◽  
Fat Cells ◽  
Structural Protein ◽  
Coordinate Regulation ◽  
Rat Adipocytes ◽  
Transport Vesicles ◽  
Adipocyte Cell ◽  
Insulin Dependent

Native rat adipocytes and the mouse adipocyte cell line, 3T3-L1, possess transport vesicles of apparently uniform composition and size which translocate the tissue-specific glucose transporter isoform, GLUT4, from an intracellular pool to the cell surface in an insulin-sensitive fashion. Caveolin, the presumed structural protein of caveolae, has also been proposed to function in vesicular transport. Thus, we studied the expression and subcellular distribution of caveolin in adipocytes. We found that rat fat cells express the highest level of caveolin protein of any tissue studied, and caveolin is also expressed at high levels in cardiac muscle, another tissue possessing insulin responsive GLUT4 translocation. Both proteins are absent from 3T3-L1 fibroblasts and undergo a dramatic coordinate increase in expression upon differentiation of these cells into adipocytes. However, unlike GLUT4 in rat adipocytes not exposed to insulin, the majority of caveolin is present in the plasma membrane. In native rat adipocytes, intracellular GLUT4 and caveolin reside in vesicles practically indistinguishable by their size and buoyant density in sucrose gradients, and both proteins show insulin-dependent translocation to the cell surface. However, by immunoadsorption of GLUT4-containing vesicles with anti-GLUT4 antibody, we show that these vesicles have no detectable caveolin, and therefore, this protein is present in a distinct vesicle population. Thus, caveolin has no direct structural relation to the organization of the intracellular glucose transporting machinery in fat cells.

scholarly journals COPII-Coated Vesicle Formation Reconstituted with Purified Coat Proteins and Chemically Defined Liposomes

Cell ◽  
1998 ◽  
Vol 93 (2) ◽  
pp. 263-275 ◽  
Author(s):  
Ken Matsuoka ◽  
Lelio Orci ◽  
Mylène Amherdt ◽  
Sebastian Y Bednarek ◽  
Susan Hamamoto ◽  
...  
Keyword(s):  
Coated Vesicle ◽  
Vesicle Formation ◽  
Coat Proteins

scholarly journals Effect of insulin on the rates of synthesis and degradation of GLUT1 and GLUT4 glucose transporters in 3T3-L1 adipocytes

1993 ◽  
Vol 290 (3) ◽  
pp. 913-919 ◽  
Author(s):  
R J Sargeant ◽  
M R Pâquet
Keyword(s):  
Glucose Transporter ◽  
Insulin Treatment ◽  
Fold Increase ◽  
Glucose Transporters ◽  
Synthesis Rate ◽  
Insulin Stimulation ◽  
Transporter Proteins ◽  
Specific Manner ◽  
Glucose Transporter Proteins ◽  
Synthesis And Degradation

The effect of continuous insulin stimulation on the rates of turnover and on the total cellular contents of the glucose-transporter proteins GLUT1 and GLUT4 in 3T3-L1 adipocytes was investigated. Pulse-and-chase studies with [35S]methionine followed by immunoprecipitation of GLUT1 and GLUT4 with isoform-specific antibodies revealed the half-lives of these proteins to be 19 h and 50 h respectively. Inclusion of 100 nM insulin in the chase medium resulted in a decrease in the half-lives of both proteins to about 15.5 h. This effect of insulin was specific for the glucose-transporter proteins, as the average half-life of all proteins was found to be 55 h both with and without insulin stimulation. The effect of insulin on the rate of synthesis of the glucose transporters was determined by the rate of incorporation of [35S]methionine. After 24 h of insulin treatment, the rate of synthesis of GLUT1 and GLUT4 were elevated over control levels by 3.5-fold and 2-fold respectively. After 72 h of treatment under the same conditions, the rate of synthesis of GLUT1 remained elevated by 2.5-fold, whereas the GLUT4 synthesis rate was not different from control levels. Western-blot analysis of total cellular membranes revealed a 4.5-fold increase in total cellular GLUT1 content and a 50% decrease in total cellular GLUT4 after 72 h of insulin treatment. These observations suggest that the rates of synthesis and degradation of GLUT1 and GLUT4 in 3T3-L1 adipocytes are regulated independently and that these cells respond to prolonged insulin treatment by altering the metabolism of GLUT1 and GLUT4 proteins in a specific manner.

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