Demonstration of insulin-responsive trafficking of GLUT4 and vpTR in fibroblasts

2000 ◽  
Vol 113 (22) ◽  
pp. 4065-4076 ◽  
Author(s):  
M.A. Lampson ◽  
A. Racz ◽  
S.W. Cushman ◽  
T.E. McGraw
Keyword(s):  
Transferrin Receptor ◽  
Glucose Transporter ◽  
Insulin Response ◽  
Physiological Role ◽  
Muscle Cells ◽  
Cell Types ◽  
Whole Body ◽  
Trafficking Pathway ◽  
Undifferentiated Cells ◽  
Regulated Trafficking

Insulin-responsive trafficking of the GLUT4 glucose transporter and the insulin-regulated aminopeptidase (IRAP) in adipose and muscle cells is well established. Insulin regulation of GLUT4 trafficking in these cells underlies the role that adipose tissue and muscle play in the maintenance of whole body glucose homeostasis. GLUT4 is expressed in a very limited number of tissues, most highly in adipose and muscle, while IRAP is expressed in many tissues. IRAP's physiological role in any of the tissues in which it is expressed, however, is unknown. The fact that IRAP, which traffics by the same insulin-regulated pathway as GLUT4, is expressed in ‘non-insulin responsive’ tissues raises the question of whether these other cell types also have a specialized insulin-regulated trafficking pathway. The existence of an insulin-responsive pathway in other cell types would allow regulation of IRAP activity at the plasma membrane as a potentially important physiological function of insulin. To address this question we use reporter molecules for both GLUT4 and IRAP trafficking to measure insulin-stimulated translocation in undifferentiated cells by quantitative fluorescence microscopy. One reporter (vpTR), a chimera between the intracellular domain of IRAP and the extracellular and transmembrane domains of the transferrin receptor, has been previously characterized. The other is a GLUT4 construct with an exofacial HA epitope and a C-terminal GFP. By comparing these reporters to the transferrin receptor, a marker for general endocytic trafficking, we demonstrate the existence of a specialized, insulin-regulated trafficking pathway in two undifferentiated cell types, neither of which normally express GLUT4. The magnitude of translocation in these undifferentiated cells (approximately threefold) is similar to that reported for the translocation of GLUT4 in muscle cells. Thus, undifferentiated cells have the necessary retention and translocation machinery for an insulin response that is large enough to be physiologically important.

scholarly journals GLUT4 Retention in Adipocytes Requires Two Intracellular Insulin-regulated Transport Steps

2002 ◽  
Vol 13 (7) ◽  
pp. 2421-2435 ◽  
Author(s):  
Anja Zeigerer ◽  
Michael A. Lampson ◽  
Ola Karylowski ◽  
David D. Sabatini ◽  
Milton Adesnik ◽  
...  
Keyword(s):  
Cell Surface ◽  
Transferrin Receptor ◽  
Glucose Transporter ◽  
Trafficking Pathway ◽  
Endosomal Recycling ◽  
Step Model ◽  
Recycling Pathway ◽  
Endosomal Pathway ◽  
Endosomal System ◽  
Regulated Trafficking

Insulin regulates glucose uptake into fat and muscle by modulating the distribution of the GLUT4 glucose transporter between the surface and interior of cells. The GLUT4 trafficking pathway overlaps with the general endocytic recycling pathway, but the degree and functional significance of the overlap are not known. In this study of intact adipocytes, we demonstrate, by using a compartment-specific fluorescence-quenching assay, that GLUT4 is equally distributed between two intracellular pools: the transferrin receptor-containing endosomes and a specialized compartment that excludes the transferrin receptor. These pools of GLUT4 are in dynamic communication with one another and with the cell surface. Insulin-induced redistribution of GLUT4 to the surface requires mobilization of both pools. These data establish a role for the general endosomal system in the specialized, insulin-regulated trafficking of GLUT4. Trafficking through the general endosomal system is regulated by rab11. Herein, we show that rab11 is required for the transport of GLUT4 from endosomes to the specialized compartment and for the insulin-induced translocation to the cell surface, emphasizing the importance of the general endosomal pathway in the specialized trafficking of GLUT4. Based on these findings we propose a two-step model for GLUT4 trafficking in which the general endosomal recycling compartment plays a specialized role in the insulin-regulated traffic of GLUT4. This compartment-based model provides the framework for understanding insulin-regulated trafficking at a molecular level.

Studies of the regulated assembly of SNARE complexes in adipocytes

2014 ◽  
Vol 42 (5) ◽  
pp. 1396-1400 ◽  
Author(s):  
Dimitrios Kioumourtzoglou ◽  
Jessica B.A. Sadler ◽  
Hannah L. Black ◽  
Rebecca Berends ◽  
Cassie Wellburn ◽  
...  
Keyword(s):  
Membrane Trafficking ◽  
Glucose Transporter ◽  
Cell Types ◽  
Whole Body ◽  
Secretory Cells ◽  
Glut4 Translocation ◽  
Glucose Transporter Proteins ◽  
Regulated Trafficking ◽  
Insight Into ◽  
Signalling Cascade

Insulin plays a fundamental role in whole-body glucose homeostasis. Central to this is the hormone's ability to rapidly stimulate the rate of glucose transport into adipocytes and muscle cells [1]. Upon binding its receptor, insulin stimulates an intracellular signalling cascade that culminates in redistribution of glucose transporter proteins, specifically the GLUT4 isoform, from intracellular stores to the plasma membrane, a process termed ‘translocation’ [1,2]. This is an example of regulated membrane trafficking [3], a process that also underpins other aspects of physiology in a number of specialized cell types, for example neurotransmission in brain/neurons and release of hormone-containing vesicles from specialized secretory cells such as those found in pancreatic islets. These processes invoke a number of intriguing biological questions as follows. How is the machinery involved in these membrane trafficking events mobilized in response to a stimulus? How do the signalling pathways that detect the external stimulus interface with the trafficking machinery? Recent studies of insulin-stimulated GLUT4 translocation offer insight into such questions. In the present paper, we have reviewed these studies and draw parallels with other regulated trafficking systems.

Erythropoietin receptor in human skeletal muscle and the effects of acute and long-term injections with recombinant human erythropoietin on the skeletal muscle

2008 ◽  
Vol 104 (4) ◽  
pp. 1154-1160 ◽  
Author(s):  
Carsten Lundby ◽  
Ylva Hellsten ◽  
Mie B. F. Jensen ◽  
Anders S. Munch ◽  
Henriette Pilegaard
Keyword(s):  
Skeletal Muscle ◽  
Transferrin Receptor ◽  
Human Skeletal Muscle ◽  
Physiological Role ◽  
Cell Types ◽  
Erythropoietin Receptor ◽  
Mrna Levels ◽  
Human Erythropoietin ◽  
Plasma Hormones

The presence and potential physiological role of the erythropoietin receptor (Epo-R) were examined in human skeletal muscle. In this study we demonstrate that Epo-R is present in the endothelium, smooth muscle cells, and in fractions of the sarcolemma of skeletal muscle fibers. To study the potential effects of Epo in human skeletal muscle, two separate studies were conducted: one to study the acute effects of a single Epo injection on skeletal muscle gene expression and plasma hormones and another to study the effects of long-term (14 wk) Epo treatment on skeletal muscle structure. Subjects ( n = 11) received a single Epo injection of 15,000 IU (double blinded, cross over, placebo). A single Epo injection reduced myoglobin and increased transferrin receptor and MRF-4 mRNA content within 10 h after injection. Plasma hormones remained unaltered. Capillarization and fiber hypertrophy was studied in subjects ( n = 8) who received long-term Epo administration, and muscle biopsies were obtained before and after. Epo treatment did not alter mean fiber area (0.84 ± 0.2 vs. 0.72 ± 0.3 mm2), capillaries per fiber (4.3 ± 0.5 vs. 4.4 ± 1.3), or number of proliferating endothelial cells. In conclusion, the Epo-R is present in the vasculature and myocytes in human skeletal muscle, suggesting a role in both cell types. In accordance, a single injection of Epo regulates myoglobin, MRF-4, and transferrin receptor mRNA levels. However, in contrast to our hypothesis, prolonged Epo administration had no apparent effect on capillarization or muscle fiber hypertrophy.

scholarly journals GLUT4 Is Retained by an Intracellular Cycle of Vesicle Formation and Fusion with Endosomes

2004 ◽  
Vol 15 (2) ◽  
pp. 870-882 ◽  
Author(s):  
Ola Karylowski ◽  
Anja Zeigerer ◽  
Alona Cohen ◽  
Timothy E. McGraw
Keyword(s):  
Plasma Membrane ◽  
Cell Surface ◽  
Glucose Transporter ◽  
Kinetic Studies ◽  
Whole Body ◽  
Microtubule Cytoskeleton ◽  
Vesicle Budding ◽  
Trafficking Pathway ◽  
Retrieval Mechanism ◽  
Golgi Network

The intracellularly stored GLUT4 glucose transporter is rapidly translocated to the cell surface upon insulin stimulation. Regulation of GLUT4 distribution is key for the maintenance of whole body glucose homeostasis. We find that GLUT4 is excluded from the plasma membrane of adipocytes by a dynamic retention/retrieval mechanism. Our kinetic studies indicate that GLUT4-containing vesicles continually bud and fuse with endosomes in the absence of insulin and that these GLUT4 vesicles are 5 times as likely to fuse with an endosome as with the plasma membrane. We hypothesize that this intracellular cycle of vesicle budding and fusion is an element of the active mechanism by which GLUT4 is retained. The GLUT4 trafficking pathway does not extensively overlap with that of furin, indicating that the trans-Golgi network, a compartment in which furin accumulates, is not a significant storage reservoir of GLUT4. An intact microtubule cytoskeleton is required for insulin-stimulated recruitment to the cell surface, although it is not required for the basal budding/fusion cycle. Nocodazole disruption of the microtubule cytoskeleton reduces the insulin-stimulated exocytosis of GLUT4, accounting for the reduced insulin-stimulated translocation of GLUT4 to the cell surface.

Transferrin receptor 2 mediates a biphasic pattern of transferrin uptake associated with ligand delivery to multivesicular bodies

2004 ◽  
Vol 287 (6) ◽  
pp. C1769-C1775 ◽  
Author(s):  
Aeisha D. Robb ◽  
Maria Ericsson ◽  
Marianne Wessling-Resnick
Keyword(s):  
Plasma Membrane ◽  
Transferrin Receptor ◽  
Physiological Role ◽  
Cell Types ◽  
Multivesicular Bodies ◽  
Unique Role ◽  
Biphasic Pattern ◽  
Exogenous Expression ◽  
Transferrin Uptake

The physiological role of transferrin (Tf) receptor 2 (TfR2), a homolog of the well-characterized TfR1, is unclear. Mutations in TfR2 result in hemochromatosis, indicating that this receptor has a unique role in iron metabolism. We report that HepG2 cells, which endogenously express TfR2, display a biphasic pattern of Tf uptake when presented with ligand concentrations up to 2 μM. The apparently nonsaturating pathway of Tf endocytosis resembles TfR1-independent Tf uptake, a process previously characterized in some liver cell types. Exogenous expression of TfR2 but not TfR1 induces a similar biphasic pattern of Tf uptake in HeLa cells, supporting a role for TfR2 in this process. Immunoelectron microscopy reveals that while Tf, TfR1, and TfR2 are localized in the plasma membrane and tubulovesicular endosomes, TfR2 expression is associated with the additional appearance of Tf in multivesicular bodies. These combined results imply that unlike TfR1, which recycles apo-Tf back to the cell surface after the release of iron, TfR2 promotes the intracellular deposition of ligand. Tf delivered by TfR2 does not appear to be degraded, which suggests that its delivery to this organelle may be functionally relevant to the storage of iron in overloaded states.

scholarly journals Rab-GAP TBC1D4 (AS160) is dispensable for the renal control of sodium and water homeostasis but regulates GLUT4 in mouse kidney

2015 ◽  
Vol 309 (9) ◽  
pp. F779-F790 ◽  
Author(s):  
Marianna Di Chiara ◽  
Bob Glaudemans ◽  
Dominique Loffing-Cueni ◽  
Alex Odermatt ◽  
Hadi Al-Hasani ◽  
...  
Keyword(s):  
Glucose Uptake ◽  
Glucose Transporter ◽  
Insulin Response ◽  
Physiological Role ◽  
Distal Tubule ◽  
Rab Gtpase ◽  
Thick Ascending Limb ◽  
Water Restriction ◽  
Distal Tubules

The Rab GTPase-activating protein TBC1D4 (AS160) controls trafficking of the glucose transporter GLUT4 in adipocytes and skeletal muscle cells. TBC1D4 is also highly abundant in the renal distal tubule, although its role in this tubule is so far unknown. In vitro studies suggest that it is involved in the regulation of renal transporters and channels such as the epithelial sodium channel (ENaC), aquaporin-2 (AQP2), and the Na+-K+-ATPase. To assess the physiological role of TBC1D4 in the kidney, wild-type (TBC1D4+/+) and TBC1D4-deficient (TBC1D4−/−) mice were studied. Unexpectedly, neither under standard nor under challenging conditions (low Na+/high K+, water restriction) did TBC1D4−/−mice show any difference in urinary Na+and K+excretion, urine osmolarity, plasma ion and aldosterone levels, and blood pressure compared with TBC1D4+/+mice. Also, immunoblotting did not reveal any change in the abundance of major renal sodium- and water-transporting proteins [Na-K-2Cl cotransporter (NKCC2) NKCC2, NaCl cotransporter (NCC), ENaC, AQP2, and the Na+-K+-ATPase]. However, the abundance of GLUT4, which colocalizes with TBC1D4 along the distal nephron of TBC1D4+/+mice, was lower in whole kidney lysates of TBC1D4−/−mice than in TBC1D4+/+mice. Likewise, primary thick ascending limb (TAL) cells isolated from TBC1D4−/−mice showed an increased basal glucose uptake and an abrogated insulin response compared with TAL cells from TBC1D4+/+mice. Thus, TBC1D4 is dispensable for the regulation of renal Na+and water transport, but may play a role for GLUT4-mediated basolateral glucose uptake in distal tubules. The latter may contribute to the known anaerobic glycolytic capacity of distal tubules during renal ischemia.

Hemostatic Factors and Inflammatory Disease

1999 ◽  
Vol 82 (08) ◽  
pp. 858-864 ◽  
Author(s):  
Jay Degen
Keyword(s):  
Endothelial Cells ◽  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Biological Activities ◽  
Bacterial Species ◽  
Physiological Role ◽  
Muscle Cells ◽  
Cell Types ◽  
Hemostatic Factors

IntroductionVascular integrity is preserved by a sophisticated system of circulating and cell-associated hemostatic factors that control local thrombin generation, platelet deposition, and the conversion of soluble fibrinogen to an insoluble fibrin matrix.1,2 However, there is considerable evidence that hemostatic factors play both a wider physiological role than simply controlling blood loss, and a wider pathological role than simply triggering inopportune thrombotic events, such as myocardial infarction and stroke. In tissue repair, a crucial physiological process, fibrin(ogen) is thought to provide a critical provisional matrix on which cells can proliferate, organize, and carry out specialized functions. A variety of cell types specifically bind to and migrate on fibrin(ogen) matrices. These include endothelial cells, macrophages, neutrophils, smooth muscle cells, fibroblasts, and keratinocytes.3-8 Direct binding to fibrin(ogen) through both integrin [e.g., αvβ3, α1β5, αMβ2 (CD11b/CD18, Mac-1)] and non-integrin receptors (e.g., intercellular adhesion molecule (ICAM-1)) appears to contribute to these cell-fibrin interactions.8-11 Fibrin(ogen) degradation products have also been reported to have an impressive array of biological activities, including mitogenic, angiogenic, chemotactic, and immunosuppressive activities.12-14 There are now substantial data indicating that fibrin(ogen) may plays an important role in the inflammatory response15,16 and that it may, in fact, direct leukocyte transendothelial cell migration.11 Similarly, through several G-protein coupled protease-activated receptors on fibroblasts, endothelial cells, leukocytes, smooth muscle cells, and other cell types, thrombin is thought to play an important role in inflammatory and fibroproliferative responses.17 Fibrinolytic factors, such as plasmin(ogen), also appear to be important modulators of inflammation.18 Finally, host fibrinogen, prothrombin, plasminogen, plasminogen activator, and other hemostatic factors appear to be crucial to the pathogenesis and virulence of many bacterial species.19-21 Unfortunately, despite a myriad of provocative observations made using in vitro systems, there is little direct in vivo evidence supporting an important role of fibrin(ogen) or other hemostatic factors in either the inflammatory response or disease progression. Direct and definitive analyses have been hampered by the lack of an experimental means to specifically manipulate the level or structure of selected hemostatic factors in vivo. Fortunately, this experimental roadblock has been effectively removed by the development of gene-targeting and gene transfer technologies in mice (see below).

scholarly journals The clathrin heavy chain isoform CHC22 functions in a novel endosomal sorting step

2010 ◽  
Vol 188 (1) ◽  
pp. 131-144 ◽  
Author(s):  
Christopher Esk ◽  
Chih-Ying Chen ◽  
Ludger Johannes ◽  
Frances M. Brodsky
Keyword(s):  
Heavy Chain ◽  
Glucose Transporter ◽  
Muscle Cells ◽  
Cell Types ◽  
Hek293 Cells ◽  
Glucose Transporter 4 ◽  
Organelle Biogenesis ◽  
Endosomal Sorting ◽  
Clathrin Heavy Chain ◽  
Golgi Network

Clathrin heavy chain 22 (CHC22) is an isoform of the well-characterized CHC17 clathrin heavy chain, a coat component of vesicles that mediate endocytosis and organelle biogenesis. CHC22 has a distinct role from CHC17 in trafficking glucose transporter 4 (GLUT4) in skeletal muscle and fat, though its transfection into HEK293 cells suggests functional redundancy. Here, we show that CHC22 is eightfold less abundant than CHC17 in muscle, other cell types have variably lower amounts of CHC22, and endogenous CHC22 and CHC17 function independently in nonmuscle and muscle cells. CHC22 was required for retrograde trafficking of certain cargo molecules from endosomes to the trans-Golgi network (TGN), defining a novel endosomal-sorting step distinguishable from that mediated by CHC17 and retromer. In muscle cells, depletion of syntaxin 10 as well as CHC22 affected GLUT4 targeting, establishing retrograde endosome–TGN transport as critical for GLUT4 trafficking. Like CHC22, syntaxin 10 is not expressed in mice but is present in humans and other vertebrates, implicating two species-restricted endosomal traffic proteins in GLUT4 transport.

scholarly journals Short communication: Maternal heat stress during the dry period alters postnatal whole-body insulin response of calves

2014 ◽  
Vol 97 (2) ◽  
pp. 897-901 ◽  
Author(s):  
S. Tao ◽  
A.P.A. Monteiro ◽  
M.J. Hayen ◽  
G.E. Dahl
Keyword(s):  
Heat Stress ◽  
Short Communication ◽  
Insulin Response ◽  
Whole Body ◽  
Dry Period

scholarly journals Selective deletion of SHIP-1 in hematopoietic cells in mice leads to severe lung inflammation involving ILC2 cells

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Xujun Ye ◽  
Fengrui Zhang ◽  
Li Zhou ◽  
Yadong Wei ◽  
Li Zhang ◽  
...  
Keyword(s):  
Lung Inflammation ◽  
Airway Remodeling ◽  
Pulmonary Inflammation ◽  
Hematopoietic Cells ◽  
Cell Lineage ◽  
Allergic Inflammation ◽  
Cell Types ◽  
Lymphoid Cells ◽  
Whole Body

AbstractSrc homology 2 domain–containing inositol 5-phosphatase 1 (SHIP-1) regulates the intracellular levels of phosphotidylinositol-3, 4, 5-trisphosphate, a phosphoinositide 3–kinase (PI3K) product. Emerging evidence suggests that the PI3K pathway is involved in allergic inflammation in the lung. Germline or induced whole-body deletion of SHIP-1 in mice led to spontaneous type 2-dominated pulmonary inflammation, demonstrating that SHIP-1 is essential for lung homeostasis. However, the mechanisms by which SHIP-1 regulates lung inflammation and the responsible cell types are still unclear. Deletion of SHIP-1 selectively in B cells, T cells, dendritic cells (DC) or macrophages did not lead to spontaneous allergic inflammation in mice, suggesting that innate immune cells, particularly group 2 innate lymphoid cells (ILC2 cells) may play an important role in this process. We tested this idea using mice with deletion of SHIP-1 in the hematopoietic cell lineage and examined the changes in ILC2 cells. Conditional deletion of SHIP-1 in hematopoietic cells in Tek-Cre/SHIP-1 mice resulted in spontaneous pulmonary inflammation with features of type 2 immune responses and airway remodeling like those seen in mice with global deletion of SHIP-1. Furthermore, when compared to wild-type control mice, Tek-Cre/SHIP-1 mice displayed a significant increase in the number of IL-5/IL-13 producing ILC2 cells in the lung at baseline and after stimulation by allergen Papain. These findings provide some hints that PI3K signaling may play a role in ILC2 cell development at baseline and in response to allergen stimulation. SHIP-1 is required for maintaining lung homeostasis potentially by restraining ILC2 cells and type 2 inflammation.

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