Provost

Steve A. N. Goldstein
'78 B.A., '78 M.A., Ph.D., F.A.A.P.

Professor of Biochemistry
Contact Information
Goldstein Lab

Research Interests

Our research is directed towards understanding how ion channels operate in health and illness. These integral membrane proteins catalyze the selective transfer of ions across membranes and, like enzymes, show exquisite specificity and tight regulation. As a class, ion channels orchestrate the electrical activity that allows operation of the heart, nervous system and skeletal muscles--even the signals in T cells require ion channels. Less sensational but equally important, ion channels mediate cellular fluid and electrolyte homeostasis. Remarkably, fundamental questions remain to be answered. How do ion channels open and close? What is their architecture? How do mutations produce cardiac arrhythmia, hypertension, seizures, or deafness? How do drugs act to produce beneficial outcomes (~20% of our current pharmacopeia targets ion channels) or to yield undesirable side effects? Our laboratory uses macroscopic and single molecule electrophysiology and spectroscopy, molecular genetics, high-throughput and structural methods to pursue five research directions:

Figure 1 Figure 1. MinK (E1) slows the movement of voltage sensors in Q1 (black traces are current recordings and red traces show sensor movement by changes in site-directed fluorescence), Ruscic 2013.

(1) Accessory Subunits—discovery, roles in health and disease, and structural basis for function. Ion channels are composed of pore-forming subunits and accessory subunits that determine, where, when and how the pores function. Accessory subunits are the power behind the throne, determining the differences in how channels operate from tissue to tissue (and from cell to cell within the same tissue) allowing the heart to beat slowly and neurons to respond in milliseconds (Figures 1 and 2). MinK (encoded by KCNE1) has just 129 amino acids and a single transmembrane domain. Nonetheless, in the heart, MinK assembles with KCNQ1 (to form IKs channels) establishing the conductance, gating, regulation and anti-arrhythmic drug sensitivity of the mixed complexes. In mutant form, MinK is associated with cardiac arrhythmia and deafness due to changes in these same attributes. It appeared that MinK was unique until we discovered a family of genes encoding MinK-related peptides (MiRPs) in 1998. Since then we have shown roles for MiRP1 (encoded by KCNE2) and MiRP2 (encoded by KCNE3) in normal and disordered function of the heart and skeletal muscle. Other accessory subunits we study include KChIPs, DPPs, 14-3-3 and KCTDs.

Illustrative citations: (Tai and Goldstein 1998), (Abbott and Goldstein 1998), (Abbott, Butler et al. 2001), (Kim, Furst et al. 2004), (O'Kelly and Goldstein 2008), (Ruscic, Miceli et al. 2013)

Figure 2. X-ray crystallography shows KCTD5 to be a tetramer. Panel shows the C module in high and low salt crystals, Dementieva 2009.

Figure 2

(2) The K2Ps—discovery of a family of potassium channels that produce background currents. Hodgkin and Huxley showed background potassium currents to central physiology but for 50 years their molecular nature was uncertain. We discovered K2P channels in yeast, worms, flies and mammals in 1995. The channels are novel in structure as well as function: they bear 2 pore-forming domains in each subunit. In humans, there are 15 KCNK genes for K2P channels and studies have begun to reveal their roles in the heart and nervous system, for example, as targets of volatile anesthetics and unique forms of channel regulation in development and across tissues, for example, modification of ion selectivity via alternative initiation translation (ATI), Figures 3 and 4.

Illustrative citations: (Ketchum, Joiner et al. 1995), (Goldstein, Price et al. 1996), (Bockenhauer, Zilberberg et al. 2001), (Thomas, Plant et al. 2008), (Plant, Zuniga et al. 2012).

figure 3

Figure 3. A homology model for ?K2PØ channels from drosophila melanogaster shows bilateral symmetry with a fourfold symmetric selectivity filter from Kollowe 2009.

figure 4 Figure 4. K2P channels have a 2P/4TM subunit topology. Active channels are open rectifiers that allow large outward currents under normal conditions of high internal and low external K+ (Phys) whereas they show a nearly linear current-voltage relationship in symmetrical K+ (Sym) from Goldstein 2001.
figure 5

Figure 5. Sumoylation of Kv2.1 in rat hippocampal neurons shown by antibody-mediated FRET (left). Each channel tetramer carries two SUMOs on non-adjacent subunits (right); stoichiometry is determined by expression of Kv2.1 and GFP-tagged SUMO1 in CHO cells, TIRF visualization of the cell surface and stepwise photobleaching from Plant 2011.

(3) SUMO—a pathway is discovered to control the activity of ion channels at the cell surface. Recently, we identified a novel enzymatic pathway at the cell surface that regulates the opening and closing of ion channels: post-translational modification with the protein called SUMO. SUMO was previously known only to determine the activity of transcription factors in the nucleus. Enzymes for sumoylation and desumoylation were shown to reside at the plasma membrane and now has been seen in all mammalian cells studied and the number of ion channels recognized to be modulated by the pathway is rapidly increasing, Figure 5.

Illustrative citations: (Rajan, Plant et al. 2005), (Plant, Dementieva et al. 2010), (Plant, Dowdell et al. 2011).

(4) Development of new genetic and high throughput methods for ion channels. A major focus is de novo development of peptide neurotoxins for "orphan" receptors. Among the most powerful tools in the arsenal for studies of the nervous system, these potent reagents are not available for the vast majority of membrane receptors critical to normal physiology and disease, see (Takacs, Toups et al. 2009), Figure 6.

Other ongoing work is allowing studies of single ion channel complexes in living cells by total internal reflection microscopy and fluorescence energy transfer, see (Plant, Dowdell et al. 2011); reveal the mechanism of operation of Killer RNA viruses that impact agriculture, commercial fermentation and fungal infections in immuno-compromised patients (a coupled toxin-immunity system acting via fungal two P domain channels), see, (Sesti, Shih et al. 2001); and use random mutation and selective pressure to clone or investigate potassium channel structure and function, see, (Sesti, Rajan et al. 2003).

figure 6
Figure 6. Moka1 (GQ153941), a de novo toxin. (A) KTX phage bind to KcsA-1.3 but not wildtype KcsA channels in ELISA (Left). (B) Combinatorial library construction and sorting. Thirty-one scorpion toxins with the KTx scaffold aligned to define 30, 22, and 17 unique domains and combine for a library diversity of 11,220. Moka1 is composed of residues in three natural toxins Ce3, AgTx2, and CTX (Takacs, Toups et al., 2009).

(5) Mechanism, diagnosis and treatment for ion channel disease. We study disorders of the heart, skeletal muscle, and the nervous system that are inherited and acquired as well as sudden infant death syndrome (SIDS) to understand cause, provide diagnostic tools, develop therapeutic strategies and avoid untoward effects of medications. Thus, rare inherited mutations of MiRP1 are associated with the cardiac arrhythmia long QT syndrome (LQTS) and sudden death while a common polymorphism present in 1.6% of the general population predisposes to a prevalent and equally dangerous disorder drug-induced LQTS (Figure 7).
Illustrative citations: (Sesti, Abbott et al. 2000), (Plant, Bowers et al. 2006), (Silva and Goldstein 2012).

figure 7

Figure 7. A SNP in SCN5A for a variant cardiac sodium channel linked to SIDS. (a) dHPLC waveform and direct sequencing of wild type and S1103Y variant. (b) Topology of sodium channel encoded by SCN5A indicating the cytoplasmic location of the S1103Y missense change (red), the four homologous membrane domains (DI-DIV), the pore-forming (P) loops and voltage sensing segments (+).

Illustrative Citations

Abbott, G.W., M.H. Butler, S. Bendahhou, M.C. Dalakas, L.J. Ptacek and S.A. Goldstein (2001). "MiRP2 forms potassium channels in skeletal muscle with Kv3.4 and is associated with periodic paralysis." Cell 104(2): 217-231.

Abbott, G.W. and S.A.N. Goldstein (1998). "A superfamily of small potassium channel subunits: form and function of the MinK-related peptides (MiRPs)." Quarterly Rev Biophys 31: 357-398.

Bockenhauer, D., N. Zilberberg and S.A.N. Goldstein (2001). "KCNK2: reversible conversion of a hippocampal potassium leak into a voltage-dependent channel." Nature Neuroscience 4: 486-491.

Goldstein, S.A.N., L.A. Price, D.N. Rosenthal and M.H. Pausch (1996). "ORK1, a potassium-selective leak channel with two pore domains cloned from Drosophila melanogaster by expression in Saccharomyces cerevisiae." Proc Natl Acad Sci 93: 13256-13261.

Ketchum, K.A., W.J. Joiner, A.J. Sellers, L.K. Kaczmarek and S.A.N. Goldstein (1995). "A new family of outwardly-rectifying potassium channel proteins with two pore domains in tandem." Nature 376: 690-695.

Kim, L.A., J. Furst, D. Gutierrez, M.H. Butler, S. Xu, S.A. Goldstein and N. Grigorieff (2004). "Three-dimensional structure of I(to); Kv4.2-KChIP2 ion channels by electron microscopy at 21 Angstrom resolution." Neuron 41(4): 513-519.

O'Kelly, I. and S.A.N. Goldstein (2008). "Forward transport of K2P3.1: mediation by 14-3-3 and COPI, modulation by p11." Traffic 9(1): 72-78.

Plant, L.D., P.N. Bowers, Q. Liu, T. Morgan, T.T. Zhang, M.W. State, W. Chen, R.A. Kittles and S.A.N. Goldstein (2006). "A common cardiac sodium channel variant associated with sudden infant death in African Americans, SCN5A S1103Y." J Clin Invest 116: 430-435.

Plant, L.D., I.S. Dementieva, A. Kollewe, S. Olikara, J.D. Marks and S.A. Goldstein (2010). "One SUMO is sufficient to silence the dimeric potassium channel K2P1." Proc Natl Acad Sci 107(23): 10743-10748.

Plant, L.D., E.J. Dowdell, I.S. Dementieva, J.D. Marks and S.A.N. Goldstein (2011). "SUMO modification of cell surface Kv2.1 potassium channels regulates the activity of rat hippocampal neurons." J Gen Physiol 137(5): 441-454.

Plant, L.D., L. Zuniga, D. Araki, J.D. Marks and S.A. Goldstein (2012). "SUMOylation silences heterodimeric TASK potassium channels containing K2P1 subunits in cerebellar granule neurons." Sci Signal 5(251): ra84.

Rajan, S., L.D. Plant, M.L. Rabin, M.H. Butler and S.A.N. Goldstein (2005). "Sumoylation silences the plasma membrane leak K+ channel K2P1." Cell 121: 37-47.

Ruscic, K. J., F. Miceli, C. A. Villalba-Galea, H. Dai, Y. Mishina, F. Bezanilla and S. A. Goldstein (2013). "IKs channels open slowly because KCNE1 accessory subunits slow the movement of S4 voltage sensors in KCNQ1 pore-forming subunits." Proc Natl Acad Sci U S A 110(7): E559-566.

Sesti, F., G.W. Abbott, J.Wei, K.T. Murray, S. Saksena, P.J. Schwartz, S.G. Priori, D.M. Roden, A.L. George, Jr. and S.A. Goldstein (2000). "A common polymorphism associated with antibiotic-induced cardiac arrhythmia." Proc Natl Acad Sci 97(19): 10613-10618.

Sesti, F., S. Rajan, R. Gonzalez-Colaso, N. Nikolaeva and S.A.N. Goldstein (2003). "Hyperpolarization moves S4 sensors inward to open MVP, a methanococcal voltage-gated potassium channel." Nature Neuroscience 6(4): 353-361.

Sesti, F., T.M. Shih, N. Nikolaeva and S.A. Goldstein (2001). "Immunity to K1 killer toxin: internal TOK1 blockade.' Cell 105(5): 637-644.

Silva, J., R. and S.A.N. Goldstein (2012). "Voltage Sensor Movements Describe Slow inactivation of Voltage-gated Sodium Channels. II L689I, a Mutation Causing Hyperkalemic Periodic Paralysis." J Gen Physiol 141(3): 323-334.

Tai, K.K. and S.A.N. Goldstein (1998). "The conduction pore of a cardiac potassium channel." Nature 391: 605-608.

Takacs, Z., M. Toups, A. Kollewe, E. Johnson, L.G. Cuello, G. Driessens, M. Biancalana, A. Koide, C.G. Ponte, E. Perozo, T.F. Gajewski, G. Suarez-Kurtz, S. Koide and S.A.N. Goldstein (2009). "A designer ligand specific for Kv1.3 channels from a scorpion neurotoxin-based library." Proceedings of the National Academy of Sciences 106: 22211-22216.

Thomas, D., L.D. Plant, C.M. Wilkens, Z.A. McCrossan and S.A.N. Goldstein (2008). "Alternative Translation Initiation in Rat Brain Yields K2P2.1 Potassium Channels Permeable to Sodium." Neuron 58(6): 859-870.

KEY WORDS: ion channels, potassium channels, sodium channels, background channels, heart, brain, skeletal muscle, disease, sudden death, SIDS, periodic paralysis, single molecule biophysics

Last review: August 8, 2013

 

 
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