We are interested understanding how insulin stimulates glucose transport into fat and muscle cells, and how this process breaks down in disease.
One of the ways in which insulin lowers blood glucose is through stimulating glucose transport into adipose and muscle tissue. Insulin activates a signal transduction cascade in these tissues to promote the translocation of the glucose transporter GLUT4 from specialised intracellular storage vesicles to the plasma membrane, facilitating glucose uptake. We currently have an incomplete understanding of the signalling events and trafficking processes that control the redistribution of GLUT4. One of the objectives of our work is to fill in these knowledge gaps.
The reason we want to increase our understanding in this area is that impaired insulin-stimulated glucose transport in muscle and fat is a major contributor to whole body insulin resistance – a state where insulin no longer efficiently lowers blood glucose and a risk factor for type II diabetes. There is currently no consensus on the molecular basis for impaired insulin responses in these tissues. We aim to shed light on how the insulin signalling network and GLUT4 trafficking apparatus are altered in insulin resistance.
We take an interdisciplinary approach using cell culture and in vivo models to study insulin action, GLUT4 trafficking and glucose metabolism. We design and perform unbiased mass spectrometry-based proteomics studies to uncover proteins or protein post-translational modifications (e.g. phosphorylation) that may play a role in insulin-stimulated GLUT4 trafficking and/or insulin resistance.
A current focus of our work is to develop techniques that allow us to screen many genes-of-interest for a role in insulin-stimulated GLUT4 trafficking, and that allow us to study district aspects of GLUT4 trafficking in cells (e.g. delivery to the cell surface, internalisation). We use our expertise in cell biology (e.g. microscopy) and biochemistry (e.g. subcellular fractionation, immunoprecipitation) techniques to study the role that proteins-of-interest play in the insulin signalling-GLUT4 pathway in health and disease.
Krycer JR, Quek L-E, Francis D, Zadoorian A, Weiss F.C, Cooke KC, Nelson ME, Diaz-Vegas A, Humphrey SJ, Scalzo R, Hirayama A, Ikeda S, Shoji F, Suzuki K, Huynh K, Giles C, Varney B, Nagarajan SR, Hoy A.J, Soga T, Meikle PJ, Cooney GJ, Fazakerley DJ, James DE. Insulin signalling requires glucose to promote lipid anabolism in adipocytes. J Biol Chem. 2020 doi: 10.1074/jbc.RA120.014907
Krycer JR, Elkington SD, Diaz-Vegas A, Cooke KC, Burchfield JG, Fisher-Wellman KH, Cooney GJ, Fazakerley DJ, James DE. Mitochondrial oxidants, but not respiration, are sensitive to glucose in adipocytes. J Biol Chem. 2020 doi: 10.1074/jbc.RA119.011695.
Kearney AL, Cooke KC, Norris DM, Zadoorian A, Krycer JR, Fazakerley DJ, Burchfield JG, James DE. Serine 474 phosphorylation is essential for maximal Akt2 kinase activity in adipocytes. J Biol Chem. 2019 doi: 10.1074/jbc.RA119.010036.
Fazakerley DJ, Krycer JR, Kearney AL, Hocking SL, James DE. Muscle and adipose tissue insulin resistance: malady without mechanism? J Lipid Res. 2018 doi: 10.1194/jlr.R087510.
Duan X, Krycer JR, Cooke KC, Yang G, James DE, Fazakerley DJ. Membrane Topology of Trafficking Regulator of GLUT4 1 (TRARG1). Biochemistry. 2018 doi:10.1021/acs.biochem.8b00361.
Fazakerley DJ, Minard AY, Krycer JR, Thomas KC, Stöckli J, Harney DJ, Burchfield JG, Maghzal GJ, Caldwell ST, Hartley RC, Stocker R, Murphy MP, James DE. Mitochondrial oxidative stress causes insulin resistance without disrupting oxidative phosphorylation.
J Biol Chem. 2018 doi: 10.1074/jbc.RA117.001254.
Burchfield JG, Kebede MA, Meoli CC, Stöckli J, Whitworth PT, Wright AL, Hoffman NJ, Minard AY, Ma X, Krycer JR, Nelson ME, Tan SX, Yau B, Thomas KC, Wee NKY, Khor EC, Enriquez RF, Vissel B, Biden TJ, Baldock PA, Hoehn KL, Cantley J, Cooney GJ, James DE, Fazakerley DJ. High dietary fat and sucrose results in an extensive and time-dependent deterioration in health of multiple physiological systems in mice. J Biol Chem. 2018 doi: 10.1074/jbc.RA117.000808.
Fazakerley DJ, Chaudhuri R, Yang P, Maghzal GJ, Thomas KC, Krycer JR, Humphrey SJ, Parker BL, Fisher-Wellman KH, Meoli CC, Hoffman NJ, Diskin C, Burchfield JG, Cowley MJ, Kaplan W, Modrusan Z, Kolumam G, Yang JY, Chen DL, Samocha-Bonet D, Greenfield JR, Hoehn KL, Stocker R, James DE. Mitochondrial CoQ deficiency is a common driver of mitochondrial oxidants and insulin resistance. Elife. 2018 doi: 10.7554/eLife.32111.
Fazakerley DJ, Naghiloo S, Chaudhuri R, Koumanov F, Burchfield JG, Thomas KC, Krycer JR, Prior MJ, Parker BL, Murrow BA, Stöckli J, Meoli CC, Holman GD, James DE. Proteomic Analysis of GLUT4 Storage Vesicles Reveals Tumor Suppressor Candidate 5 (TUSC5) as a Novel Regulator of Insulin Action in Adipocytes. J Biol Chem. 2015 doi: 10.1074/jbc.M115.657361.
Stöckli J, Fazakerley DJ, James DE. GLUT4 exocytosis. J Cell Sci. 2011 doi: 10.1242/jcs.097063.
Rowland AF, Fazakerley DJ, James DE. Mapping insulin/GLUT4 circuitry. Traffic. 2011 doi: 10.1111/j.1600-0854.2011.01178.x.
Fazakerley DJ, Holman GD, Marley A, James DE, Stöckli J, Coster AC.
Kinetic evidence for unique regulation of GLUT4 trafficking by insulin and AMP-activated protein kinase activators in L6 myotubes. J Biol Chem. 2010 doi: 10.1074/jbc.M109.051185.
Fazakerley DJ, Lawrence SP, Lizunov VA, Cushman SW, Holman GD. A common trafficking route for GLUT4 in cardiomyocytes in response to insulin, contraction and energy-status signalling. J Cell Sci. 2009 doi: 10.1242/jcs.041178.