Regulation of Neuronal Proapoptotic Potassium Currents by the Hepatitis C Virus Nonstructural Protein 5A

Abstract

Apoptosis-enabling neuronal potassium efflux is mediated by an enhancement of K+ currents. In cortical neurons, increased currents are triggered by dual phosphorylation of Kv2.1 by Src and p38 at channel residues Y124 and S800. It was recently shown that a K+ current surge is also present in hepatocytes undergoing apoptosis, and that the hepatitis C virus (HCV) nonstructural protein 5A (NS5A) could inhibit Kv2.1-mediated currents and block cell death. Here, we show that NS5A1b (from HCV genotype 1b) expression in rat neurons depresses delayed rectifier potassium currents, limits the magnitude of the K+ current surge following exposure to activated microglia, and is neuroprotective. In a non-neuronal recombinant expression system, cells expressing Kv2.1 mutated at residue Y124, but not S800 mutants, are insensitive to NS5A1b-mediated current inhibition. Accordingly, NS5A1b co-expression prevents phosphorylation of wild-type Kv2.1 by Src at Y124, but is unable to inhibit p38 phosphorylation of the channel at S800. The actions of the viral protein are genotype-selective, as NS5A1a does not depress neuronal potassium currents nor inhibit Src phosphorylation of Kv2.1. Our results indicate that NS5A1b limits K+ currents following injury, leading to increased neuronal viability. NS5A1b may thus serve as a model for a new generation of neuroprotective agents.

Viral dependence on cellular ion channels – an emerging anti-viral target?

“…It was also revealed that NS5A inhibits stress-mediated neuronal cell death through the inhibition of K v 2.1 activation via Src-mediated phosphorylation [17]. It is likely that HCV thus perturbs K v 2.1 activation to bypass reactive oxygen species-mediated apoptosis to allow virus persistence and survival [18]. In addition to regulating K v 2.1 currents, HCV was found to increase intracellular hepatic Cl À influx, which can be inhibited by selective Cl À channel blockers [19].…”

Voltage-Gated Potassium Channels at the Crossroads of Neuronal Function, Ischemic Tolerance, and Neurodegeneration

“…Zn 2+ release) and late pro-apoptotic processes are elicited by such a diverse range of toxic stimuli, converging on Kv2.1-mediated K + current enhancement, strongly suggests that this step represents a key mechanism in neuronal apoptosis that could be therapeutically targeted. In this vein, the hepatitis C virus nonstructural protein 5A (NS5A) was recently discovered to attenuate pro-apoptotic Kv2.1 K + current enhancement in hepatocytes and cortical neurons [125, 136, 137]. This K + current blockade has been suggested to occur through NS5A-mediated inhibition of mixed lineage kinase 3 (MLK3), a MAP kinase kinase kinase which promotes the activation of p38 kinase [125].…”

“…However, in another study, NS5A was shown to block Src kinase-facilitated phosphorylation of the Y124 residue, without affecting channel phosphorylation of S800 by p38 kinase. In fact, pseudo-phosphorylation of Kv2.1 channels at S800 does not eliminate NS5A-induced inhibition of K + currents, whereas Kv2.1 channels expressing a phospho-mimetic substitution at Y124F are no longer susceptible to K + current attenuation by NS5A, strongly indicating that NS5A exerts its inhibition of Kv2.1 currents and neuroprotective effects through preventing Src kinase-mediated Y124 phosphorylation rather than by blocking p38 kinase-induced S800 phosphorylation [137]. This mechanism warrants further exploration, as NS5A could serve as a model for new neuroprotective agents specifically targeting pro-apoptotic Kv2.1-mediated K + currents.…”

Disruption of K V 2.1 somato-dendritic clusters prevents the apoptogenic increase of potassium currents

“…The delayed rectifier current, predominantly mediated by K V 2.1 channels (Murakoshi & Trimmer, 1999; Malin & Nerbonne, 2002), has been demonstrated to perform a critical role in apoptogenic K + efflux in cortical, nigral, and hippocampal neurons (Pal et al , 2003; Redman et al , 2006; Shen et al , 2009; Shepherd et al , 2012). Suppressing delayed rectifier current-mediated K + efflux decreases cellular susceptibility to apoptotic stimuli, including oxidative stress (Yu et al , 1997; Aizenman et al , 2000; McLaughlin et al , 2001; Wei et al , 2004; Pal et al , 2006; Redman et al , 2007; Redman et al , 2009; Norris et al , 2012; Shepherd et al , 2012). …”