Amira Klip, PhD
Director, Research Training
University of Toronto
Biochemistry, Paediatrics & Physiology
Canada Research Chair
Cell Biology of Insulin Action
Dr. Amira Klip is a senior scientist in the Cell Biology Program at the SickKids Research Institute and Professor of Paediatrics, Biochemistry, and Physiology at the University of Toronto. In order to understand the process of insulin resistance, a problem for both type 1 and type 2 diabetes, Klip studies insulin action and the cellular and molecular steps involved in this process. She directs a laboratory of ten graduate students/postdoctoral fellows, one technician, and one research associate and works with several pharmaceutical companies to screen and identify modes of action of potential anti-diabetic drugs.
Klip received a PhD in Biochemistry (Mexico City, 1976), performed postdoctoral work in Toronto and Zurich, and joined The Hospital for Sick Children (SickKids) in 1979. She is a Canadian Institutes of Health Research (CIHR) Distinguished Scientist and a fellow of the Royal Society of Canada. She has been a member of the CIHR grant panels on cell physiology, cell biology, and mechanisms of disease. She is an executive member of the Banting and Best Diabetes Centre at the University of Toronto.
Klip’s research is currently funded by the CIHR operating grants and the Canadian Diabetes Association (CDA) Grant-in-Aid.
- insulin action
- glucose transport
- intracellular traffic
- signal transduction
Glucose is the major energy substrate for most cells, and it is avidly stored as glycogen in the liver and muscle tissue, as well as processed into fat in adipose tissue. The liver provides the rest of the body with glucose between meals, especially the brain. However, during a meal, insulin derived from the pancreas vigorously promotes glucose uptake into muscle and fat cells and stops the liver from releasing glucose to the blood. The mechanism whereby insulin increases glucose uptake into muscle/fat has received much attention but is not completely understood. Insulin resistance, a key defect in type 2 diabetes, involves defective responses to insulin in muscle, adipose and hepatic tissues.
Klip’s laboratory has been studying the regulation of glucose uptake by insulin and muscle contraction, using an array of rat and mouse stable muscle cell lines that they have generated. The focus is the intracellular traffic of vesicles containing glucose transporters, primarily GLUT4. Current work focuses on how a series of signal transduction pathways activated by insulin within muscle cells impinge on intracellular stores of GLUT4, how the vesicles move to the cell surface, and how they are further turned-on into faster carriers. This recruitment of transporters and their subsequent activation requires participation of the actin cytoskeleton for faithful congregation of the pertinent signals and gathering of GLUT4 below the plasma membrane.
Her lab’s studies have revealed two important bifurcations in insulin action, one defined by different outcomes of the insulin receptor substrate 1 (IRS-1) protein and insulin receptor substrate 2 (IRS-2) protein, the other defined by two signalling arms downstream of phosphatidylinositol 3-kinase (PI 3-kinase). The Akt/PKB arm, a serine/threonine protein kinase that plays a key role in multiple cellular processes such as glucose metabolism, cell proliferation, apoptosis, transcription and cell migration, leads to the inactivation of the RAb-GAP AS160 (Akt substrate of 160 kDa), which acts through distinct Rab molecules to mobilize and position GLUT4 vesicles. In muscle cells, their work identifies Rab8A, Rab13 and Rab14 as important mediators of these steps. On the other hand, the Rac activation arm functions to rapidly remodel actin filaments into a cortical mesh below the cell surface, as shown through a dynamic cycle of actin branching mediated by Arp2/3 and severing mediated by actin-binding proteins know as cofilin. Vesicles mobilized to the cell cortex interact with the actin mesh through actinin-4, possibly in preparation for optimal docking and fusion with the membrane. Klip’s team has also identified the SNARE molecules VAMP2, syntaxin4 and SNAP23 as mediators of this docking/fusion step. GLUT4 arrives at the cell surface in a state of low activity but are soon activated by a pathway that may involve the removal of ancillary inhibitors or the binding of activators of GLUT4. In particular, GAPDH binds to GLUT4 as Hexokinase II is displaced, possibly accounting for the activation of the transporter.
In a new series of studies, the Klip lab has also established cellular models to study insulin resistance:
- Exposure of muscle cell cultures to high glucose and high insulin, emulating the environment in type 2 diabetes;
- Exposure to high saturated fats or their derivative ceramides and reactive oxygen species, emulating the lipotoxic component of the metabolic syndrome; and
- Exposure to conditioned media from palmitate-treated macrophages, emulating the inflammation component of insulin resistance.
In each case, GLUT4 translocation in response to insulin was dampened, but interestingly through distinct signalling defects, respectively: reduction in IRS-1 phosphorylation, in Rac activation and in Akt and AS160 phosphorylation. These findings suggest that insulin resistance in vivo must be analyzed at each signalling level and may require distinct and specific interventional therapies.
- Intracellular compartment plasticity and signal transmission in insulin action.
Canadian Institutes of Health Research (2005-10)
- The GLUT4 interactomes and their metabolic implications.
Canadian Institutes of Health Research (2007-12)
- Regulation of muscle glucose transporters by Rac and Rabs during insulin action and lipotoxicity.
Canadian Institutes of Health Research (2010-15)
- Interplay between fatty acids and macrophages in the genesis and relief of muscle cell insulin resistance.
Canadian Diabetes Association (2009-12)
Fink LN, Oberbach A, Costford SR, Chan Kenny, Sams A, Blüher M, Klip A. (2013) Expression of anti-inflammatory macrophage genes within skeletal muscle correlates with insulin sensitivity in human obesity and type 2 diabetes. Diabetologia, Manuscript Diab-12-1975.R1, accepted/in press.
Zhu Y, Pereira RO, O’Neill BT, Riehle C, Ilkun O, Wende AR, Rawlings TA, Zhang YC, Zhang Q, Klip A, Shiojima I, Walsh K, Abel ED. (2013) Cardiac PI3K-Akt Impairs Insulin-Stimulated Glucose Uptake Independent of mTORC1 and GLUT4 Translocation. Mol. Endocrinol. 27: 1: 172-184.
Teixeira SS, Tamrakar AK, Goulart-Silva F, Serrano-Nascimento C, Klip A, Nunes MT. (2012) Triiodothyronine Acutely Stimulates Glucose Transport into L6 Muscle Cells Without Increasing Surface GLUT4, GLUT1, or GLUT3. Thyroid 22: 747-54.
Boguslavsky S, Chiu T, Foley KP, Osorio-Fuentealba C, Antonescu CN, Bayer KU, Bilan PJ, Klip A. (2012) Myo1c binding to submembranous actin mediates insulin-induced tethering of GLUT4 vesicles. Mol. Biol. Cell 23: 4065-78.
Hussey SE, Liang H, Costford SR, Klip A, DeFronzo RA, Sanchez-Avila A, Ely B, Musi N. (2012) TAK-242, a small-molecule inhibitor of Toll-like receptor 4 signaling, unveils similarities and differences in lipopolysaccharide- and lipid-induced inflammation and insulin resistance in muscle cells. Biosci. Rep. 33: 37-47.
Pillon N, Arane K, Bilan PJ, Chiu TT, Klip A.(2012) Muscle cells challenged with saturated fatty acids mount an autonomous inflammatory response that activates macrophages. Cell Commun. Signal. 10: 1: 30.
Osorio-Fuentealba C, Contreras-Ferrat AE, Altamirano F, Espinosa A, Li Q, Niu W, Lavandero S, Klip A, Jaimovich E. (2012) Electrical Stimuli Release ATP to Increase Glucose Uptake and GLUT4 Translocation via PI3Kγ-Akt-AS160 in Skeletal Muscle Cells. Diabetes, Dec 28, doi. 10.2337/db12-0491.Epub ahead of print.
Sylow L, Jensen TE, Kleinert M, Mouatt JR, Maarbjerg SJ, Jeppesen J, Prats C, Chiu TT, Boguslavsky S, Klip A, Schjerling P, Richter EA. (2012) Rac1 Is a Novel Regulator of Contraction-Stimulated Glucose Uptake in Skeletal Muscle. Diabetes, Dec 28, doi. 10.2337/db12-0491. Doi: 10.2337/db12-1066. Epub ahead of print.
Niu W, Bilan PJ, Yu J, Gao J, Boguslavsky S, Schertzer JD, Chu G, Yao Z, Klip A. (2011) PKCε Regulates Contraction-Stimulated GLUT4 Traffic in Skeletal Muscle Cells. J. Cell. Physiol. 226: 173-80.
Schertzer JD, Tamrakar AK, Magalhães JG, Pereira S, Bilan PJ, Fullerton MD, Liu Z, Steinberg GR, Giacca A, Philpott DJ, Klip A. (2011) NOD1 Activators Link Innate Immunity to Insulin Resistance. Diabetes 60: 2206-15.
Yu J, Shi L, Wang H, Bilan PJ, Yao Z, Samaan MC, He Q, Klip A, Niu W. (2011) Conditioned medium from hypoxia-treated adipocytes renders muscle cells insulin resistant. Eur. J. Cell Biol. 90: 1000-15.
Kewalramani G, Fink LN, Asadi F, Klip A. (2011) Palmitate-Activated Macrophages Confer Insulin Resistance to Muscle Cells by a Mechanism Involving Protein Kinase C θ and ε. PLoS One 6: e26947.
Tamrakar AK, Schertzer JD, Chiu TT, Foley KP, Bilan PJ, Philpott DJ, Klip A. (2010) NOD2 Activation Induces Muscle Cell-Autonomous Innate Immune Responses and Insulin Resistance. Endocrinol. 151: 5624-37.
Sun Y, Bilan PJ, Liu Z, Klip A. (2010) Rab8A and Rab13 are activated by insulin and regulate GLUT4 translocation in muscle cells. Proc. Natl. Acad. Sci. USA 107: 46: 19909-14.
Chiu TT, Patel N, Shaw AE, Bamburg JR, Klip A: (2010) Arp2/3- and cofilin-coordinated actin dynamics is required for insulin-mediated GLUT4 translocation to the surface of muscle cells. Mol. Biol. Cell, Epub ahead of print Sept 1.
Contreras-Ferrat AE, Toro B, Bravo R, Parra V, Vásquez C, Ibarra C, Mears D, Chiong M, Jaimovich E, Klip A, Lavandero S: (2010) An inositol 1,4,5-triphosphate (IP3)-IP3 receptor pathway Is required for insulin-stimulated glucose transporter 4 translocation and glucose uptake in cardiomyocytes. Endocrinol., 151: 10: Epub ahead of print Aug 4.
Niu W, Bilan PJ, Yu J, Gao J, Boguslavsky S, Schertzer JD, Chu G, Yao Z, Klip A: (2010) PKCe regulates contraction-stimulated GLUT4 traffic in skeletal muscle cells. J. Cell. Physiol., Epub ahead of print July 23.
Schwenk RW, Dirkx E, Coumans WA, Bonen A, Klip A, Glatz JFC, Luiken JJFP: (2010) Requirement for distinct vesicle-associated membrane proteins in insulin- and AMP-activated protein kinase (AMPK)-induced translocation of GLUT4 and CD36 in cultured cardiomyocytes. Diabetologia, 53: 10: 2209-19.
Niu W, Bilan PJ, Ishikura S, Schertzer JD, Contreras-Ferrat A, Fu Z, Liu J, Boguslavsky S, Foley KP, Liu Z, Li J, Chu G, Panakkezhum T, Lopaschuk GD, Lavandero S, Yao Z, Klip A: (2010) Contraction-related stimuli regulate GLUT4 traffic in C2C12-GLUT4myc skeletal muscle cells. Am. J. Physiol. Endocrinol. Metab., 298: E1058-71.
Hartig SM, Ishikura S, Hicklen RS, Feng Y, Blanchard EG, Voelker KA, Pichot CS, Grange RW, Raphael RM, Klip A, Corey SJ: (2009) The F-BAR protein CIP4 promotes GLUT4 endocytosis through bidirectional interactions with N-WASp and Dynamin-2. J. Cell Sci., 122: 2283-91.
Schertzer J, Antonescu CN, Bilan PJ, Jain S, Huang X, Liu Z, Bonen A, Klip A: (2009) A transgenic mouse model to study glucose transporter 4myc regulation in skeletal muscle. Endocrinol.,150: 1935-40.
Zaid H, Talior-Volodarsky I, Antonescu C, Liu Z, Klip A: (2009) GAPDH binds GLUT4 reciprocally to hexokinase-II and regulates glucose transport activity. Biochem. J., 419: 475-84.
Samokhvalov V, Bilan PJ, Schertzer JD, Antonescu CN, Klip A: (2009) Palmitate- and lipopolysaccharide-activated macrophages evoke contrasting insulin responses in muscle cells. Am. J. Physiol. Endocrinol. Metab., 296: E37–46.
Wood TE, Dalili S, Simpson CD, Hurren R, Mao X, Saiz FS, Gronda M, Eberhard Y, Minden MD, Bilan PJ, Klip A, Batey RA, Schimmer AD: (2008) A novel inhibitor of glucose uptake sensitizes cells to FAS-induced cell death. Mol. Cancer Ther., 7: 3546-55.
Ishikura S, Klip A: (2008) Muscle cells engage Rab8A and myosin Vb in insulin-dependent GLUT4 translocation. Am. J. Physiol. Cell Physiol., 295: C1016-25.
Randhawa VK, Ishikura S, Talior-Volodarksy I, Cheng AW, Patel N, Hartwig JH, Klip A: (2008) GLUT4 vesicle recruitment and fusion are differentially regulated by Rac, AS160, and Rab8A in muscle cells. J. Biol. Chem., 283: 27208-19.
Talior-Volodarsky I, Randhawa VK, Zaid H, Klip A: (2008) α-Actinin-4 is selectively required for insulin-induced GLUT4 translocation. J. Biol. Chem., 283: 25115-23.
Antonescu CN, Díaz M, Femia G, Planas JV, and Klip A: (2008) Clathrin-dependent and independent endocytosis of glucose transporter 4 (GLUT4) in myoblasts: regulation by mitochondrial uncoupling. Traffic, 9: 1173-90.
Dugani CB, Randhawa VR, Cheng AW, Patel N, and Klip A: (2008) Selective regulation of the perinuclear distribution of glucose transporter 4 (GLUT4) by insulin signals in muscle cells. Eur. J. Cell Biol., 87: 337-51.
Roher N, Samokhvalov V, Díaz M, MacKenzie S, Klip A, and Planas JV: (2008) The proinflammatory cytokine tumor necrosis factor-a increases the amount of glucose transporter-4 at the surface of muscle cells independently of changes in interleukin-6. Endocrinol., 149: 1880-9.
Ishikura S, Bilan PJ, and Klip A: (2007) Rabs 8A and 14 are targets of the insulin-regulated Rab-GAP AS160 regulating GLUT4 traffic in muscle cells. Biochem. Biophys. Res. Commun., 353: 1074-9.
Thong FSL, Bilan PJ, and Klip A: (2007) The Rab GTPase-activating protein AS160 integrates Akt, protein kinase C, and AMP-activated protein kinase signals regulating GLUT4 traffic. Diabetes, 56: 414-23.
JeBailey L, Wanono O, Niu W, Roessler J, Rudich A, and Klip A: (2007) Ceramide- and oxidant-Induced insulin resistance involve loss of insulin-dependent Rac-activation and actin remodeling in muscle cells. Diabetes, 56: 394-403.
Wijesekara N, Tung A, Thong F, and Klip A: (2006) Muscle cell depolarization induces a gain in surface GLUT4 via reduced endocytosis independently of AMPK. Am. J. Physiol. Endocrinol. Metab., 290: E1276-86.
Thirone ACP, JeBailey L, Bilan PJ, and Klip A: (2006) Opposite effect of JAK2 on insulin-dependent activation of mitogen-activated protein kinases and Akt in muscle cells: possible target to ameliorate insulin resistance. Diabetes, 55: 942-51.
Foster LJ, Rudich A, Talior I, Patel N, Huang X, Furtado LM, Bilan PJ, Mann M, and Klip A: (2006) Insulin-dependent interactions of proteins with GLUT4 revealed through stable isotope labeling by amino acids in cell culture (SILAC). J. Proteome Res., 5: 64-75.
Ishiki M, Randhawa VK, Poon V, JeBailey L, and Klip A: (2005) Insulin regulates the membrane arrival, fusion, and C-terminal unmasking of glucose transporter-4 via distinct phosphoinositides. J. Biol. Chem., 280: 28792-802.
Antonescu CN, Huang C, Niu W, Liu Z, Eyers PA, Heidenreich KA, Bilan PJ, and Klip A: (2005) Reduction of insulin-stimulated glucose uptake in L6 myotubes by the protein kinase inhibitor SB203580 is independent of p38 MAPK activity. Endocrinol., 146: 3773-81.
Huang C, Thirone ACP, Huang X, and Klip A: (2005) Differential contribution of insulin receptor substrates 1 versus 2 to insulin signaling and glucose uptake in L6 myotubes. J. Biol. Chem., 280: 19426-35.
Konrad D, Rudich A, Bilan P J, Patel N, Richardson C, Witters LA, and Klip A: (2005) Troglitazone causes acute mitochondrial membrane depolarization and an AMPK-mediated increase in glucose phosphorylation in muscle cells. Diabetologia, 48: 954-66.
Wijesekara N, Konrad D, Eweida M, Jefferies C, Liadis N, Giacca A, Crackower M, Suzuki A, Mak TW, Kahn CR, Klip A, and Woo M: (2005) Muscle-specific pten deletion protects against insulin resistance and diabetes. Mol. Cell. Biol., 25: 1135-45.
Randhawa VK, Thong FSL, Lim DY, Li D, Garg RR, Rudge R, Galli T, Rudich A, and Klip A: (2004) Insulin and hypertonicity recruit GLUT4 to the plasma membrane of muscle cells by Using N-Ethylmaleimide-sensitive factor-dependent SNARE mechanisms but different v-SNAREs: role of TI-VAMP. Mol. Biol. Cell, 15: 5565-73.
Török D, Patel N, JeBailey L, Thong FSL, Randhawa VK, Klip A, and Rudich A: (2004) Insulin but not PDGF rely on actin remodeling and on VAMP2 for GLUT4 translocation in myoblasts. J. Cell Sci., 117: 5447-55.
Sweeney G, Garg RR, Ceddia RB, Li D, Ishiki M, Somwar R, Foster LJ, Neilsen PO, Prestwich GD, Rudich A, and Klip A: (2004) Intracellular delivery of phosphatidylinositol (3,4,5)-trisphosphate causes incorporation of glucose transporter 4 into the plasma membrane of muscle and fat cells without increasing glucose uptake. J. Biol. Chem., 279: 32233-42.
JeBailey L, Rudich A, Huang X, Di Ciano-Oliveira C, Kapus A, and Klip A: (2004) Skeletal muscle cells and adipocytes differ in their reliance on TC10 and Rac for insulin-induced actin remodeling. Mol. Endocrinol., 18: 359-72.
Patel N, Rudich A, Khayat Z, Garg R, and Klip A: (2003) Intracellular segregation of phosphatidylinositol-3,4,5-trisphosphate by insulin-dependent actin remodeling in L6 skeletal muscle cells. Mol. Cell. Biol., 23: 4611-26.
Rudich A, Konrad D, Török D, Ben-Romano R, Huang C, Niu W, Garg RR, Wijesekara N, Germinario RJ, Bilan PJ, and Klip A: (2003) Indinavir uncovers different contributions of GLUT4 and GLUT1 towards glucose uptake in muscle and fat cells and tissues. Diabetologia, 46: 649-58.
Pillon NJ, Bilan PJ, Fink LN, Klip A. (2013) Cross-talk between skeletal muscle and immune cells: muscle-derived mediators and metabolic implications. Am. J. Physiol. Endocrinol. Metab. 304: E453-65.
Lee WL, Klip A. (2012) Shuttling glucose across brain microvessels, with a little help from GLUT1 and AMP Kinase. Focus on “AMP kinase regulation of sugar transport in brain capillary endothelial cells during acute metabolic stress”. Am. J. Physiol. Cell Physiol. 303: C803-5.
Foley K, Boguslavsky S, Klip A. (2011) Endocytosis, Recycling, and Regulated Exocytosis of Glucose Transporter 4. Biochemistry 50: 3048-61.
Chiu TT, Jensen TE, Sylow L, Richter EA, Klip A. (2011) Rac1 signalling towards GLUT4/glucose uptake in skeletal muscle. Cell. Signal. 23: 1546-54.
Schertzer JD, Klip A. (2011) Give a NOD to Insulin Resistance. Am. J. Physiol. Endocrinol. Metab. 301: E585-6.
Badawi A, Klip A, Haddad P, Cole DEC, Bailo BG, El-Sohemy A, Karmali M. (2010) Type 2 diabetes mellitus and inflammation: Prospects for biomarkers of risk and nutritional intervention. Diabetes, Metabolic Syndrome and Obesity: Targets and Therapy, Dove Medical Press Ltd., 3: 173-86.
Kewalramani G, Bilan PJ, Klip A: (2010) Muscle insulin resistance: assault by lipids, cytokines and local macrophages. Curr. Opin. Clin. Nutr. Metab. Care, 13: 382-90.
Ishikura S, Antonescu CN, Klip A: (2010) Documenting GLUT4 exocytosis and endocytosis in muscle cell monolayers. Curr. Protoc. Cell Biol., 46: Unit 15.15: 1-9.
Bilan PJ, Samokhvalov V, Koshkina A, Schertzer JD, Samaan CM, Klip A: (2009) Direct and macrophage-mediated actions of fatty acids causing insulin resistance in muscle cells. Arch. Physiol. Biochem., 115: 4: 176-90.
Klip A: (2009) The many ways to regulate glucose transporter 4. Appl. Physiol. Nutr. Metab., 34: 481-7.
Klip A, Schertzer JD, Bilan PJ, Thong F, Antonescu C: (2009) Regulation of glucose transporter 4 traffic by energy deprivation from mitochondrial compromise. Acta Physiol., 196: 1: 27-35.
Antonescu CN, Foti M, Sauvonnet N, Klip A: (2009) Ready, set, internalize: mechanisms and regulation of GLUT4 endocytosis. Biosci. Rep., 29: 1: 1-11.
Antonescu CN, Randhawa VK, Klip A: (2008) Dissecting GLUT4 traffic components in L6 myocytes by fluorescence-based, single-cell assays. Meth. Mol. Biol., Membrane Trafficking, Vancura A (Editor), Humana Press, Totowa, New Jersey, 457: 367-78.
Samaan MC, Klip A: (2008) Obesity, insulin resistance, and type 2 diabetes: the fat-muscle connection. Endocrinology Rounds (As presented in the Rounds of the Division of Endocrinology and Metabolism, St. Michael’s Hospital), 8: 1-6.
Zaid H, Antonescu CN, Randhawa VK, and Klip A: (2008) Insulin action on glucose transporters through molecular switches, tracks and tethers. Biochem. J., 413: 201-15.
Ishikura S, Koshkina A, and Klip A: (2008) Small G proteins in insulin action: Rab and Rho families at the crossroads of signal transduction and GLUT4 vesicle traffic. Acta Physiol., 192: 61-74.
Dugani CB, and Klip A: (2005) Glucose transporter 4: cycling, compartments and controversies. EMBO Reports, 6: 1137-42.
Ishiki M, and Klip A: (2005) Minireview: Recent developments in the regulation of glucose transporter-4 traffic: new signals, locations, and partners. Endocrinol., 146: 5071-8.
Thong FSL, Dugani CB, and Klip A: (2005) Turning signals on and off: GLUT4 traffic in the insulin-signalling highway. Physiol., 20: 271-84.
Use of R-(+)-.alpha.-lipoic acid, R-(-)dihydrolipoic acid and metabolites for the treatment of Diabetes Mellitus (US patent 5,693,664)