Neurobiology; Ion Channel Disorder Mechanisms; Ion Channel Molecular Biology. Gene expression, biosynthesis, trafficking, regulation, and function of membrane proteins (e.g. channels, receptors, pumps, transporters): Membrane proteins play critical roles in all cells. ~25% of mammalian genomes code for these proteins which are expressed in the plasma membrane and organelle membranes of all cells. Nervous systems rely on ion channels for simple reflexes to complex interactions involved in learning/memory. Ion channels set the resting membrane potential, affect excitability, and generate and modify action potentials in excitable cells- they also play other important roles in all cells. Our overall goal is to elucidate underlying principles/mechanisms governing membrane protein biology in the normal and disease state, particularly for ion channels, as well as to identify new roles for membrane proteins in cell/animal biology.

 

We are: (a) Identifying amino acid determinants on ion channels (e.g. potassium channels and glutamate receptor channels) that play positive and negative roles in their intracellular trafficking from the ER to the Golgi and from the Golgi to the cell surface and elucidating which determinant is dominant when heteromeric channels are expressed. These determinants affect the channel's cell surface level. (b) Determining the effects of glycosylation and phosphorylation in ion channel biology. Glycosylation, particularly sialidation in the Golgi, does affect channel function and it also affects cell surface levels of some channels. ER-dependent phosphorylation of some channels may also affect their cell surface levels. (c) Investigating the role of abnormal trafficking to the cell surface of mutant ion channels in human disorders (e.g. Episodic Ataxia I, SpinoCerebellar Ataxia 13). (d) Examining the role ofchannel mRNA untranslated regions in modifying their protein expression levels. (e) Cloning/characterizing new channels.

 

Modifying cell surface levels or function of ion channels by posttranslational mechanisms could increase or fine tune the functional response of a cell by altering its signaling characteristics. Understanding the role of posttranslational modifications and different trafficking programs of ion channels to the cell surface also has clinical relevance because a number of human disorders are caused by alterations or mutations that affect these parameters (e.g. ER retention disorders such as a form of episodic ataxia, epilepsy, long QT syndrome, and cystic fibrosis).

Zhu J, Yan J, & Thornhill WB (2012). N-Glycosylation promotes the cell surface expression of Kv1.3 potassium channels. FEBS J., In press, epub online.

 

Zhu J, Alsaber R, Zhao J, Ribeiro-Hurley E, & Thornhill WB (2012). Characterization of the Kv1.1 I262T and S342I mutations associated with episodic ataxia 1 with distinct phenotypes. Arch. Biochem. Biophys. In press, epub online

 

Von Bergen Granell AE, Palter KB, Akan I, Aich U, Yarema KJ, Betenbaugh MJ, Thornhill WB, Recio-Pinto E (2011). DmSAS is required for sialic acid biosynthesis in cultured Drosophila third instar larvae CNS neurons. ACS Chemical Biology 6:1287-1295.

 

Zhu J, Recio-Pinto E, Hartwig T, Sellers W, Yan J, & Thornhill WB (2009). The Kv1.2 potassium channel: the position of an N-glycan on the extracellular linkers affects its protein expression and function. Brain Res. 1251:16-29.

 

Zhu J, Gomez B, Watanabe I, & Thornhill WB (2007). Kv1 potassium channel C-terminus constant HRETE region: arginine substitution affects surface protein level and conductance level of subfamily members differentially. Mol. Memb. Biol. (England) 24:194-205.

 

Watanabe I, Zhu J, Sutachan JJ, Gottschalk A, Recio-Pinto E, & Thornhill WB (2007). The glycosylation state of Kv1.2 potassium channels affects trafficking, gating, and simulated action potentials. Brain Res. 1144:1-18.

 

Zhu J, Gomez B, Watanabe I, & Thornhill WB (2005). Amino acids in the pore region of Kv1 potassium channels dictate cell surface protein levels: a possible trafficking code in the Kv1 subfamily. Biochem. J. (England) 388:355-362.

 

Sutachan JJ, Watanabe I, Zhu J, Gottschalk A, Recio-Pinto E, & Thornhill WB (2005). Effects of Kv1.1 channel glycosylation on C-type inactivation and simulated action potentials. Brain Res. 1058:30-43.

 

Watanabe I, Zhu J, Recio-Pinto E, & Thornhill WB (2004). Glycosylation affects the protein stability and cell surface expression of Kv1.4 but not Kv1.1 potassium channels: a pore region determinant dictates the effect of glycosylation on trafficking. J. Biol. Chem. 279:8879-8885.

 

Zhu J, Watanabe I, Gomez B, & Thornhill WB (2003). Heteromeric Kv1 potassium channel expression: amino acid determinants involved in processing and trafficking to the cell surface. J. Biol. Chem. 278:25558-25567.

 

Zhu J, Watanabe I, Gomez B, & Thornhill WB (2003). Trafficking of Kv1.4 potassium channels: interdependence of a pore region determinant and a cytoplasmic C-terminal VXXSL determinant in regulating cell surface trafficking. Biochem. J. (England) 375:761-768.

 

Zhu J, Watanabe I, Poholek A, Koss M, Gomez B, Yan C, Recio-Pinto E, & Thornhill WB (2003). Allowed N-glycosylation sites on the Kv1.2 potassium channel S1-S2 linker: implications for linker secondary structure and the glycosylation effect on function. Biochem. J. (England) 375:769-775.

 

Watanabe I, Wang HG, Sutachan JJ, Zhu J, Recio-Pinto E, & Thornhill WB (2003). Glycosylation affects Kv1.1 potassium channel gating a combined surface potential and a cooperative subunit interaction mechanism. J. Physiol. (England) 550:51-66.

 

Thornhill WB, Watanabe I, Sutachan JJ, Wu MB, Wu X, Zhu J, & Recio-Pinto E (2003). Molecular cloning and functional expression of a Kv1.1-like potassium channel from the electric organ of Electrophorus electricus. J. Memb. Biol. 196:1-8.

 

Castillo C, Thornhill WB, Zhu J, & Recio-Pinto E (2003). The permeation and activation properties of brain sodium channels changes during development. Develop. Brain Res. 144:99-106.

 

Zhu J, Watanabe I, Gomez B, & Thornhill WB (2001). Determinants involved in Kv1 potassium channel folding in the endoplasmic reticulum, glycosylation in the Golgi, and cell surface expression. J. Biol. Chem. 276:39419-39427.

 

Zhu L, Wu X, Wu MB, Chan KW, Logothetis DE, & Thornhill WB (2001). Cloning and characterization of G protein-gated inward rectifier K+ channel (GIRK1) isoforms from heart and brain. J. Mol. Neurosci 16:21-32.

 

Pabon A, Chan KW, Sui JL, Wu X, Logothetis DE, & Thornhill WB (2000). Glycosylation of GIRK1 at N119 and ROMK1 at N117 have different consequences in potassium channel function. J. Biol. Chem. 275:30677-30682.

 

Castaneda-Castellanos D, Cano M, Wang JKT, Corbett A, Benson D, Blanck TJJ, Thornhill WB, & Recio-Pinto E (2000). CNS voltage-dependent Na+ channel expression and distribution in an undifferentiated and differentiated CNS cell line. Brain Res. 866:281-285.

 

Jing J, Chikvashvili D, Singer-Laht D, Thornhill WB, Reuveny E, & Lotan I (1999). Fast inactivation of a brain K+ channel composed of Kv1.1 and Kv1.1 subunits modulated by G protein subunits. EMBO J 18:1245-1256.

 

Levi M, Jie J, Chikvashvili D, Thornhill WB, & Lotan I (1998). Activation of metabotropic glutamate receptor and protein kinase C reduce the extent of inactivation of the K+ channel Kv1.1/Kvbeta1.1 via dephosphorylation of Kv1.1. J. Biol. Chem. 273: 6495-6502.

 

Jing J, Peretz T, Singer-Lahat D, Chikavshvili D, Thornhill WB, & Lotan I (1997). Inactivation of a voltage-dependent K+ channel by beta-subunit: modulation by a phosphorylation-dependent interaction between the distal C-terminus of alpha-subunit and cytoskeleton. J. Biol. Chem. 272:14021-14024.

 

Ponce A, Vega-Saenz de Miera E, Kentros C, Moreno H, Thornhill WB, & Rudy B (1997). K+ channel subunit isoforms with divergent carboxy-terminal sequences carry distinct membrane targeting signals. J. Memb. Biol. 159:149-159.

 

Castillo C, Diaz ME, Balbi D, Thornhill WB, & Recio-Pinto E (1997). Changes in sodium channel function during postnatal brain development reflect increases in the level of channel sialidation. Develop. Brain Res. 104:119-130.

 

Peretz T, Levin G, Moran O, Thornhill WB, Chikavshvili D, & Lotan I (1996). Modulation by protein kinase C of rat brain delayed-rectifier K+ channel expressed in Xenopus oocytes. FEBS LET 381:71-76.

 

Ponce A, Bueno E, Kentros C, Vega-Saenz de Miera E, Chow L, Hillman D, Chen S, Wu MB, Wu X, Zhu L, Rudy B, & Thornhill WB (1996). G protein-gated inward rectifier K+ channel proteins (GIRK1) are present in the soma and dendrites as well as nerve terminals of specific neurons in the brain. J. Neurosci. 16: 1990-2001.

 

Thornhill WB, Wu MB, Jiang X, Wu X, Morgan P, & Margiotta JF (1996). Expression of Kv1.1 delayed-rectifier potassium channels in Lec mutant CHO cell lines reveals a role for sialidation in channel function. J. Biol. Chem. 271:19093-19098.

 

Levin G, Chikvashvili D, Singer-Lahat D, Peretz T, Thornhill WB, & Lotan I (1996). Phosphorylation of a K+ channel? alpha-subunit modulates the inactivation conferred by a beta-subunit; involvement of cytoskeleton. J. Biol. Chem. 271:29321-29328.

 

Weiser M, Bueno E, Sekirnjak C, Martone ME, Baker H, Hillman D, Thornhill WB, Ellisman M, & Rudy B (1995). The potassium channel subunit Kv3.1b is localized to somatic and axonal membranes of specific populations of CNS neurons. J. Neurosci. 15:4298-4314.

Levin G, Keren T, Peretz T, Chikavshvili D, Thornhill WB, & Lotan I (1995). Regulation of RCK1 currents with a cAMP analog via enchanced protein synthesis and direct channel phosphorylation. J. Biol. Chem. 270:14611-14618.

 

Moreno H, Kentros C, Bueno E, Weiser M, Herdandez A, Vega-Saenz de Miera E, Thornhill WB, & Rudy B (1995). Thalamocortical projections have a K+ channel that is phosphorylated and modulated by cAMP-dependent protein kinase. J. Neurosci. 15:5486-5501.