Asher Peretz

A recessive C-terminal Jervell and Lange-Nielsen mutation of the KCNQ1 channel impairs subunit assembly

The LQT1 locus (KCNQ1) has been correlated with the most common form of inherited long QT (LQT) syndrome. LQT patients suffer from syncopal episodes and high risk of sudden death. The KCNQ1 gene encodes KvLQT1 α‐subunits, which together with auxiliary IsK (KCNE1, minK) subunits form IKs K+ channels. Mutant KvLQT1 subunits may be associated either with an autosomal dominant form of inherited LQT, Romano–Ward syndrome, or an autosomal recessive form, Jervell and Lange‐Nielsen syndrome (JLNS). We have identified a small domain between residues 589 and 620 in the KvLQT1 C‐terminus, which may function as an assembly domain for KvLQT1 subunits. KvLQT1 C‐termini do not assemble and KvLQT1 subunits do not express functional K+ channels without this domain. We showed that a JLN deletion–insertion mutation at KvLQT1 residue 544 eliminates important parts of the C‐terminal assembly domain. Therefore, JLN mutants may be defective in KvLQT1 subunit assembly. The results provide a molecular basis for the clinical observation that heterozygous JLN carriers show slight cardiac dysfunctions and that the severe JLNS phenotype is characterized by the absence of KvLQT1 channel.

Hypomyelination and increased activity of voltage‐gated K + channels in mice lacking protein tyrosine phosphatase ϵ

Protein tyrosine phosphatase epsilon (PTPϵ) is strongly expressed in the nervous system; however, little is known about its physiological role. We report that mice lacking PTPϵ exhibit hypomyelination of sciatic nerve axons at an early post‐natal age. This occurs together with increased activity of delayed‐ rectifier, voltage‐gated potassium (Kv) channels and with hyperphosphorylation of Kv1.5 and Kv2.1 Kv channel α‐subunits in sciatic nerve tissue and in primary Schwann cells. PTPϵ markedly reduces Kv1.5 or Kv2.1 current amplitudes in Xenopus oocytes. Kv2.1 associates with a substrate‐trapping mutant of PTPϵ, and PTPϵ profoundly reduces Src‐ or Fyn‐stimulated Kv2.1 currents and tyrosine phosphorylation in transfected HEK 293 cells. In all, PTPϵ antagonizes activation of Kv channels by tyrosine kinases in vivo, and affects Schwann cell function during a critical period of Schwann cell growth and myelination.

Constitutive activation of delayed‐rectifier potassium channels by a Src family tyrosine kinase in Schwann cells

In the nervous system, Src family tyrosine kinases are thought to be involved in cell growth, migration, differentiation, apoptosis, as well as in myelination and synaptic plasticity. Emerging evidence indicates that K+ channels are crucial targets of Src tyrosine kinases. However, most of the data accumulated so far refer to heterologous expression, and native K+‐channel substrates of Src or Fyn in neurons and glia remain to be elucidated. The present study shows that a Src family tyrosine kinase constitutively activates delayed‐rectifier K+ channels (IK) in mouse Schwann cells (SCs). IK currents are markedly downregulated upon exposure of cells to the tyrosine kinase inhibitors herbimycin A and genistein, while a potent upregulation of IK is observed when recombinant Fyn kinase is introduced through the patch pipette. The Kv1.5 and Kv2.1 K+‐channel α subunits are constitutively tyrosine phosphorylated and physically associate with Fyn both in cultured SCs and in the sciatic nerve in vivo. Kv2.1‐ channel subunits are found to interact with the Fyn SH2 domain. Inhibition of Schwann cell proliferation by herbimycin A and by K+‐channel blockers suggests that the functional linkage between Src tyrosine kinases and IK channels could be important for Schwann cell proliferation and the onset of myelination.

Stilbenes and fenamates rescue the loss of I KS channel function induced by an LQT5 mutation and other IsK mutants

Genetic and physiological studies have established a link between potassium channel dysfunction and a number of neurological and muscular disorders. Many ‘channelopathies’ are accounted for by a dominant‐lethal suppression of potassium channel function. In the cardiac IKS channel complex comprising the α and β subunits, KvLQT1 and IsK, respectively, several mutations lead to a dominant‐negative loss of channel function. These defects are responsible for a human cardiovascular disease called long QT (LQT) syndrome. Here we show that binding of IKS channel activators, such as stilbenes and fenamates, to an extracellular domain flanking the human IsK transmembrane segment, restores normal IKS channel gating in otherwise inactive IsK C‐terminal mutants, including the naturally occurring LQT5 mutant, D76N. Our data support a model in which allosteric interactions exist between the extracellular and intracellular boundaries of the IsK transmembrane segment as well as between domains of the α and β subunits. Disruption of this allosteric interplay impedes slow activation gating, decreases current amplitude and restores channel inactivation. Owing to allosteric interactions, stilbene and fenamate compounds can rescue the dominant‐negative suppression of IKS produced by IsK mutations and thus, may have important therapeutic relevance for LQT syndrome.

Heteromultimeric Delayed-Rectifier K+ Channels in Schwann Cells: Developmental Expression and Role in Cell Proliferation

Schwann cells (SCs) are responsible for myelination of nerve fibers in the peripheral nervous system. Voltage-dependent K+ currents, including inactivating A-type (KA), delayed-rectifier (KD), and inward-rectifier (KIR) K+ channels, constitute the main conductances found in SCs. Physiological studies have shown that KD channels may play an important role in SC proliferation and that they are downregulated in the soma as proliferation ceases and myelination proceeds. Recent studies have begun to address the molecular identity of K+ channels in SCs. Here, we show that a large repertoire of K+ channel α subunits of theShaker (Kv1.1, Kv1.2, Kv1.4, and Kv1.5),Shab (Kv2.1), and Shaw (Kv3.1b and Kv3.2) families is expressed in mouse SCs and sciatic nerve. We characterized heteromultimeric channel complexes that consist of either Kv1.5 and Kv1.2 or Kv1.5 and Kv1.4. In postnatal day 4 (P4) sciatic nerve, most of the Kv1.2 channel subunits are involved in heteromultimeric association with Kv1.5. Despite the presence of Kv1.1 and Kv1.2 α subunits, the K+ currents were unaffected by dendrotoxin I (DTX), suggesting that DTX-sensitive channel complexes do not account substantially for SC KDcurrents. SC proliferation was found to be potently blocked by quinidine or 4-aminopyridine but not by DTX. Consistent with previous physiological studies, our data show that there is a marked downregulation of all KD channel α subunits from P1–P4 to P40 in the sciatic nerve. Our results suggest that KD currents are accounted for by a complex combinatorial activity of distinct K+channel complexes and confirm that KDchannels are involved in SC proliferation.

Tyrosine kinases modulate K+ channel gating in mouse Schwann cells

1 The whole‐cell configuration of the patch‐clamp technique and immunoprecipitation experiments were used to investigate the effects of tyrosine kinases on voltage‐dependent K+ channel gating in cultured mouse Schwann cells. 2 Genistein, a broad‐spectrum tyrosine kinase inhibitor, markedly reduced the amplitude of a slowly inactivating delayed‐rectifier current (IK) and, to a lesser extent, that of a transient K+ current (IA). Similar results were obtained on IK with another tyrosine kinase inhibitor, herbimycin A. Daidzein, the inactive analogue of genistein, was without effect. 3 Unlike herbimycin A, genistein produced additional effects on IA by profoundly affecting its gating properties. These changes consisted of slower activation kinetics with an increased time to peak, a positive shift in the voltage dependence of activation (by +30 mV), a decrease in the steepness of activation gating (9 mV per e‐fold change) and an acceleration of channel deactivation. 4 The steepness of the steady‐state inactivation was increased by genistein treatment, while the recovery from inactivation was not significantly altered. 5 The action of genistein on voltage‐dependent K+ (Kv) currents was accompanied by a decrease in tyrosine phosphorylation of Kv1.4 as well as Kv1.5 and Kv2.1 encoding transient and slowly inactivating delayed‐rectifier K+ channel α subunits, respectively. 6 In conclusion, the present study shows that tyrosine kinases markedly affect the amplitude of voltage‐dependent K+ currents in Schwann cells and finely tune the gating properties of the transient K+ current component IA. These modulations may be functionally relevant in the control of K+ channel activity during Schwann cell development and peripheral myelinogenesis.