The organisation and dynamic regulation of ion transporters and membrane microdomains in cardiac muscle during health and disease, with particular reference to the impact of palmitoylation
Division of Molecular & Clinical Medicine
Level 5, Mailbox 12
School of Medicine, University of Dundee
Ninewells Hospital and Medical School
+(44) 01382 383089
Dr Fuller gained his BA in Pharmacology from University of Cambridge (Emmanuel College) and remained in the Department of Pharmacology in Cambridge to study for a PhD investigating the misfolding of the F508 mutant of CFTR under the supervision of the then head of department Professor AW Cuthbert. He then moved on to King's College London (St Thomas Hospital campus) where he worked on regulation of the cardiac sodium pump in the laboratory of Professor Michael Shattock. During 8 years in this lab Dr Fuller developed interests in the hormonal regulation of cardiac ion transport - working first on the sodium pump, and later on its cardiac accessory protein phospholemman. He moved to Dundee in 2006 to establish an independent research lab with a specific interest in protein-protein interactions and post-translational modifications in cardiac muscle.
Heart disease is the biggest killer in the western world. The underlying cause of heart disease is varied, but it is often characterised by major changes in ion channel expression and function in cardiac myocytes that compromise contractility and predispose to the development of lethal arrhythmias.
Research in my laboratory is split along 3 major themes:
1. Regulation of the cardiac sodium pump
The primary active means of ion transport in the cardiomyocyte sarcolemma is the sodium pump (Na/K ATPase). In cardiac muscle, the transarcolemmal sodium (Na) gradient established by Na/K ATPase activity is essential not only for generating the rapid upstroke of the action potential but also for driving a number of ion exchange and transport processes that are crucial for normal cellular function, excitation contraction coupling, ion homeostasis, mitochondrial homeostasis and the control of cell volume. By determining the set point for the sodium-calcium exchanger, the Na/K ATPase controls the predominant mechanism of transmembrane calcium flux, and hence indirectly controls intracellular calcium load and myocardial contractility. Interventions that influence either the activity of the Na/K ATPase, or indirectly the transmembrane sodium gradient, can therefore profoundly affect normal cardiac function. Misregulation of the Na/K ATPase in human heart failure not only leads to abnormalities in the Na gradient, but also knock-on effects on intracellular calcium stores and therefore alterations in cardiac contractility, as well as supply-demand mismatching at the level of mitochondria.
Regulation of the catalytic activity of the Na/K ATPase by protein kinases is through phosphorylation of phospholemman by protein kinase A and protein kinase C. Current research is directed towards understanding the relationship between these proteins in health and disease. This will increase our understanding of cardiac Na/K ATPase function and dysfunction in cardiac physiology and heart disease, and has the potential to identify new therapeutic targets in heart failure.
2. Protein Palmitoylation
S-palmitoylation is the reversible covalent post-translational attachment of the fatty acid palmitic acid to the thiol group of cysteine, via an acyl-thioester linkage. While the technology to study this post-translational modification has lagged behind commonly studied modifications such as protein phosphorylation, protein S-palmitoylation is now emerging as an important and common post-translational modification in a variety of tissues. Protein S-palmitoylation is catalysed by palmitoyl acyltransferases and reversed by protein thioesterases, and occurs dynamically and reversibly in a manner analogous to protein phosphorylation. Many different classes of protein have been identified as targets for palmitoylation, including G-proteins, ion channels, transporters, receptors and protein kinases. Palmitoylation can alter enzymatic/ion channel activity, stability or subcellular localisation of the target protein, and this is usually achieved by the recruitment of the palmitoylated cysteine to the lipid bilayer. As such, palmitoylation is largely specific for membrane-associated and integral membrane proteins, and has the potential to induce substantial changes in protein secondary structure and therefore function, through the recruitment of intracellular loops to the inner surface of the membrane bilayer. Every single route by which sodium and calcium enters and leaves a cardiac myocyte is subjected to palmitoylation, so dynamic protein palmitoylation has the potential to be as important as protein phosphorylation in the regulation of cardiac function. Indeed, since palmitoylation is restricted to integral membrane and membrane-associated proteins (i.e proteins connected with ion transport at the cell surface), it is likely to play a key role in the regulation of the cardiac output by regulating cardiac ion transporters. Palmitoylation of phospholemman contributes to its regulation of the cardiac sodium pump (see above).
Caveolae are small flask-like invaginations of the cell membrane around 50-100 nm in diameter, found in almost all cells of the body. They represent a specialised form of lipid raft, an area of the cell membrane enriched in cholesterol and sphingolipids, characterised by the presence of the protein caveolin. The lipid environment, caveolin content and morphology of caveolae are central to their diverse functional roles, which include co-ordination of signal transduction, cholesterol homeostasis, and endocytosis.
One of caveolae’s best-characterised roles is as a signalosome, a compartment that brings together components of signal transduction cascades (including receptors, effectors and targets). Caveolae have been assigned a key role in regulation of signalling in the heart. For example, adrenoceptors and their effector molecules and downstream targets such as phospholemman are found in caveolae-containing membrane fractions of the adult heart. The distribution of receptors, effectors and their targets is key to the efficiency and fidelity of their coupling.
A considerable number of cardiac ion transporters are resident in cardiac caveolae. Physical co-localisation of ion transporters in the caveolar compartment may functionally link ion flow by providing a restricted diffusional space and facilitates hormonal regulation of these transporters by placing them physically adjacent to signalling molecules. Furthermore, the presence of ion transporters in caveolae is likely to have functional relevance beyond signal transduction since the lipid composition of the bilayer in which an ion transporter resides is likely to influence its activity. We are currently evaluating dynamic changes in the caveolar compartment during adrenergic signalling in cardiac muscle.
- Pavlovic, D., Fuller, W., & Shattock M.J. (2013). Novel regulation of cardiac Na pump via phospholemman. J Mol Cell Cardiol 61 pp 83-93.
- Pavlovic, D., Hall, A., Kennington, E.J., Aughton, K., Boguslavskyii, A., Fuller, W., Despa, S., Bers, D.M. & Shattock M.J. (2013). Rapid Pacing Stimulates Na/K ATPase in Rat Ventricular Myocytes via a Nitric Oxide and Phospholemman-dependent Mechanism. J Mol Cell Cardiol 61 pp 164-71.
- Wypijewski K., Howie, J., Reilly, L., Tulloch, L.B., Aughton, K.L., McLatchie, L.M., Shattock, M.J., Calaghan SC, &. Fuller, W. (2013). A separate pool of cardiac phospholemman that does not regulate or associate with the sodium pump: multimers of phospholemman in ventricular muscle. J Biol Chem 288 pp 13808-20. Corresponding author.
- Howie, J., Tulloch, L., Shattock, M.J. & Fuller W. (2013). Regulation of the Cardiac Sodium Pump by Palmitoylation of its Catalytic and Regulatory Subunits. Biochemical Society Transactions 41 pp 95-100. Corresponding author.
- Fuller, W., Tulloch, L., Shattock, M.J., Calaghan, S.C., Howie, J. & Wypijewski K. (2013). Regulation of the cardiac sodium pump. Cellular & Molecular Life Sciences 70 pp 1357-80. Corresponding author.
- Tulloch, L., Howie, J., Wilson, C., Wypijewski, K., Bernard, W., Shattock, M.J. & Fuller W. (2011). The inhibitory effect of phospholemman on the sodium pump requires its palmitoylation. J. Biol Chem 286 pp 36020-36031. Corresponding author.
- El Armouche, A., Wittköpper, K., Howie, J., Fuller, W., Shattock, M.J. & Pavlovic, D. (2011). Phospholemman-dependent regulation of the cardiac Na/K-ATPase activity is modulated by inhibitor-1 sensitive type-1 phosphatase. FASEB J 25 pp 4467-4475.
- Wypijewski K, Shattock MJ and Fuller W (2010). Phospholemman Migrates out of Cardiomyocyte Caveolae upon Protein Kinase C Activation. Circulation 122 A21319. Selected for oral presentation at American Heart Association annual meeting, Chicago, 2010.
- Howie J, Tulloch LB, Beddie E, Shattock, MJ and Fuller W (2010) Peroxiredoxin 6 is Recruited to the Cardiac Sodium Pump Complex by Phospholemman. Circulation 122 A21311. Selected for poster presentation at American Heart Association annual meeting, Chicago, 2010.
- Madhani M, Hall AR, Cuello F, Charles RL, Burgoyne JR, Fuller W, Hobbs AJ, Shattock MJ, Eaton P. (2010) Phospholemman Ser-69 phosphorylation contributes to sildenafil-induced cardioprotection against reperfusion injury. Am J Physiol 299(3):H827-3.
- Fuller W, Howie J, McLatchie LM, Weber RJ, Hastie CJ, Burness K, Pavlovic D, Shattock MJ (2009) FXYD1 phosphorylation in vitro and in adult rat cardiac myocytes: threonine 69 is a novel substrate for protein kinase C. Am J Physiol Cell Physiol. 296:C1346-55.
- Bell JR, Kennington E, Fuller W, Dighe K, Donoghue P, Clark JE, Jia LG, Tucker AL, Moorman JR, Marber MS, Eaton P, Dunn MJ, Shattock MJ (2008) Characterization of the phospholemman knockout mouse heart: depressed left ventricular function with increased Na-K-ATPase activity. Am J Physiol Heart Circ Physiol. 294:H613-21.
- Pavlović D, Fuller W, Shattock MJ (2007) The intracellular region of FXYD1 is sufficient to regulate cardiac Na/K ATPase. FASEB J. 21:1539-46.
- Berry RG, Despa S, Fuller W, Bers DM, Shattock MJ (2007) Differential distribution and regulation of mouse cardiac Na/K-ATPase a1 and a2 subunits in T-tubule and surface sarcolemmal membranes. Cardiovasc Res. 73:92-100.
- Fuller W, Shattock MJ (2006) Phospholemman and the cardiac sodium pump: protein kinase C, take a bow. Circ Res. 99:1290-2.
Final year honours, University of Dundee - Course 4D09 Regulation of Cardiac Function.
2013: Supervisor of Fiona Ashford, BHF PhD studentship. Thesis title ‘Interaction between palmitoylation and glutathionylation in the regulation of cardiac function’.
2011: Supervisor of Louise Reilly, MRC DTA PhD student. Thesis title ‘Regulation of Cardiac Ion Transport by Protein Palmitoylation’.
2012 Tenth International Annual FXYD Symposium, Dalmunzie Castle, Scotland, 28-30 September 2012. Sole organiser.
2012 Scottish Cardiovascular Forum 15th annual meeting, February 4 2012, Apex Hotel Dundee. Principal organiser.
Invited speaker engagements:
2014 Regulation of the sodium pump in excitable tissue by phospholemman. Plenary Lecture at 14th P-type ATPase meeting, The Netherlands.
2013 Palmitoylation and the control of cardiac ion transporters in Symposium P222, Physiology of Protein Palmitoylation, IUPS 2013.
2013 New tricks for an old ion transporter: regulation of the cardiac sodium pump in Faculty of Biological Sciences Seminar Series, University of Leeds, April 2013.
2013 New tricks for an old ion transporter: regulation of the cardiac sodium pump in the Spinal Cord and Movement Group Seminar Series, University of St Andrews, February 2013.
2011 Phospholemman Palmitoylation: mechanisms, and functional effect on the pump at the Ninth Annual FXYD Symposium, UC Davis, 2011.
2010 Another FXYD Meeting, Another New Post-Translational Modification for Phospholemman at the Eighth Annual FXYD Symposium, Harvard University, Boston, 2010.
2009 The Hunt for Phospholemman interacting proteins in cardiac myocytes at the Seventh Annual FXYD Symposium, Thomas Jefferson University, Philadelphia, 2009.
2008 Phospholemman: Master Regulator of the Cardiac Na/K ATPase at Experimental Biology (American Physiological Society, Muscle Biology Group Symposium), San Diego, USA, 2008.