Gail A. Robertson, PhD Associate Professor of Physiology Room 237 MSC 1300 University Avenue Madison, WI 53706 Phone: (608) 265-3339 Fax: (608) 265-5512 Email: robertson@physiology.wisc.edu

The electrical signals underlying neuronal communication and cardiac rhythmicity depend on potassium channels, proteins that regulate the movement of potassium ions across cell membranes. The disruption of these channels by inherited diseases or drugs can lead to neurological defects or cardiac arrhythmias. Our studies focus on potassium channels encoded by the Human Ether-a-go-go Related Gene (HERG) in human heart and brain. In the heart, HERG ion channels give rise to a potassium current that repolarizes the ventricular action potential. Mutations in HERG cause congenital Long QT Syndrome (LQTS) by disrupting IKr, increasing cardiac excitability and, in some cases, triggering catastrophic torsades de pointes arrhythmias. An acquired form of LQTS is a side effect of a surprisingly large and diverse collection of pharmaceutics that block HERG channels. In either its inherited or acquired forms, LQTS can lead to life-threatening ventricular arrhythmias and sudden death.

Genetic Modifiers of HERG Channel Function and Expression

To date, more than 90 inherited mutations in HERG have been linked with LQTS. Several mutations prevent HERG channels from trafficking to the membrane, but the details of these trafficking processes are completely unknown. To elucidate the transport pathway, we isolated proteins interacting with HERG using a yeast two-hybrid screen of a human heart library. Our screen uncovered five HERG-interacting proteins, or 'HIPs', including three novel proteins. One of the HIPs is a Golgi-resident protein. Another is an actin-binding protein proposed to stabilize HERG expression at the membrane. A third is found in endosomes, where it may play a role in recycling and regulating channel density at the membrane. We are studying the HIPs using electrophysiology to assay their effects on HERG channel function or expression levels, immunocytochemistry with confocal microscopy to localize the proteins to their subcellular compartments, and yeast two-hybrid and biochemistry techniques to map interaction domains. In addition, we are screening families who carry LQTS but do not map to any of the known LQTS loci for mutations in the genes encoding these interacting proteins. With this multipdisciplinary approach we hope to uncover novel biological processes regulating ion channel function and to define additional molecular mechanisms of ion channel disease.

Gating mechanisms of HERG ion channels

A second line of research focuses on how HERG ion channels open and close, a process known as gating. For these experiments we use a structure-function approach of targeting specific amino acids for mutagenesis or modification and measuring microscopic changes in ion channel function using voltage- and patch-clamp techniques and biophysical analysis tools. Although HERG channels are structurally similar to a wide range of voltage-gated ion channels, we have identified a novel gating process that slows channel closing during cardiac repolarization and ensures that HERG channels will contribute to the final repolarization process, their key physiological role in the heart. How this process controls the channel 'gate' is the focus of current research.

Selected Publications

Roti Roti, E.C., Myers, C.D., Ayers, R.A., Boatman, D.E., Delfosse, S.A., Chan, E.K.L., Ackerman, M.J.,January, C.T. and Robertson, G.A. (2002). Interaction with GM130 during HERG ion channel trafficking Disruption by LQT2 mutations. J. Biol. Chem. 277;47779-47785.

Robertson, G.A. (2000) LQT2: Amplitude reduction and loss of selectivity in the tail that wags the HERG channel. [Editorial] Circulation Research 86(5):492-493, 2000 Mar 17.

Wang, J., Myers, C.D. and Robertson, G.A. (2000). Dynamic Control of Deactivation Gating by a Soluble Amino-Terminal Domain in HERG K+ Channels. J. Gen. Physiol. 115: 749-758.

Trudeau, M.C., Titus, S.A., Branchaw, J.L., Ganetzky, B. and Robertson, G.A. (1999). Functional analysis of a mouse Elk-type K+ channel. J. Neurosci., 19:2906-2918.

Wang, J., Trudeau, M.C., Zappia, A. and Robertson, G.A. (1998). Regulation of deactivation by an amino terminal domain in HERG potassium channels. J. Gen. Physiol. 112:637-647.

Herzberg*, I.M., Trudeau*, M.C. and Robertson, G.A. (1998). Transfer of rapid inactivation and E-4031 sensitivity from HERG to M-EAG Channels. J. Physiol. 511:3-14.

Robertson, G.A., J.W. Warmke, and B. Ganetzky. (1996). Potassium currents expressed from Drosophila and mouse eag cDNAs in Xenopus oocytes. Neuropharmacology 35: 841-850.

Trudeau, M. C., J.W. Warmke, B. Ganetzky, and G.A. Robertson. (1995). HERG, a human inward rectifier in the voltage-gated potassium channel family. Science 269: 92-95.