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ABSTRACT: Voltage-gated ion channels play a central role in the generation of action potentials in the nervous system. They are selective for one type of ion - sodium, calcium, or potassium. Voltage-gated ion channels are composed of a central pore that allows ions to pass through the membrane and four peripheral voltage sensing domains that respond to changes in the membrane potential. Upon depolarization, voltage sensors in voltage-gated potassium channels (Kv) undergo conformational changes driven by positive charges in the S4 segment and aided by pairwise electrostatic interactions with the surrounding voltage sensor. Structure-function relations of Kv channels have been investigated in detail, and the resulting models on the movement of the voltage sensors now converge to a consensus; the S4 segment undergoes a combined movement of rotation, tilt, and vertical displacement in order to bring 3-4e(+) each through the electric field focused in this region. Nevertheless, the mechanism by which the voltage sensor movement leads to pore opening, the electromechanical coupling, is still not fully understood. Thus, recently, electromechanical coupling in different Kv channels has been investigated with a multitude of techniques including electrophysiology, 3D crystal structures, fluorescence spectroscopy, and molecular dynamics simulations. Evidently, the S4-S5 linker, the covalent link between the voltage sensor and pore, plays a crucial role. The linker transfers the energy from the voltage sensor movement to the pore domain via an interaction with the S6 C-termini, which are pulled open during gating. In addition, other contact regions have been proposed. This review aims to provide (i) an in-depth comparison of the molecular mechanisms of electromechanical coupling in different Kv channels; (ii) insight as to how the voltage sensor and pore domain influence one another; and (iii) theoretical predictions on the movement of the cytosolic face of the Kv channels during gating.
Frontiers in pharmacology. 01/2012; 3:166.
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ABSTRACT: Voltage-gated ion channels are controlled by the membrane potential, which is sensed by peripheral, positively charged voltage sensors. The movement of the charged residues in the voltage sensor may be detected as gating currents. In Shaker K(+) channels, the gating currents are asymmetric; although the on-gating currents are fast, the off-gating currents contain a slow component. This slow component is caused by a stabilization of the activated state of the voltage sensor and has been suggested to be linked to ion permeation or C-type inactivation. The molecular determinants responsible for the stabilization, however, remain unknown. Here, we identified an interaction between Arg-394, Glu-395, and Leu-398 on the C termini of the S4-S5 linker and Tyr-485 on the S6 of the neighboring subunit, which is responsible for the development of the slow off-gating component. Mutation of residues involved in this intersubunit interaction modulated the strength of the associated interaction. Impairment of the interaction still led to pore opening but did not exhibit slow gating kinetics. Development of this interaction occurs under physiological ion conduction and is correlated with pore opening. We, thus, suggest that the above residues stabilize the channel in the open state.
Journal of Biological Chemistry 03/2010; 285(18):14005-19. · 4.77 Impact Factor
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ABSTRACT: High threshold for stress-induced activation of the heat shock transcription factor, Hsf1, may contribute to vulnerability of motor neurons to disease and limit efficacy of agents promoting expression of neuroprotective heat shock proteins (Hsps) through this transcription factor. Plasmid encoding a constitutively active form of Hsf1, Hsf1act, and chemicals shown to activate Hsf1 in other cells were investigated in a primary culture model of familial amyotrophic lateral sclerosis. Hsf1act and the Hsp90 inhibitor, geldanamycin, induced high expression of multiple Hsps in cultured motor neurons and conferred dramatic neuroprotection against SOD1G93A in comparison to Hsp70 or Hsp25 alone. Two other Hsp90 inhibitors, 17-allylamino-17-demethoxygeldanamycin (17-AAG) and radicicol, and pyrrolidine dithiocarbamate induced robust expression of Hsp70 and Hsp40 in motor neurons, but at cytotoxic concentrations. 17-AAG, which penetrates the blood-brain barrier, has exhibited a higher therapeutic index than geldanamycin, but this may not be the case when activation of Hsf1 in neurons is targeted.
Neurobiology of Disease 12/2006; 24(2):213-25. · 5.40 Impact Factor
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ABSTRACT: Nonsteroidal anti-inflammatory drugs (NSAIDs) have been shown to amplify the heat shock response in cell lines by increasing the binding of heat shock transcription factor-1 to heat shock elements within heat shock gene promoters. Because overexpression of the inducible heat shock protein 70 (Hsp70) was neuroprotective in a culture model of motor neuron disease, this study investigated whether NSAIDs induce Hsp70 and confer cytoprotection in motor neurons of dissociated spinal cord cultures exposed to various stresses. Two NSAIDs, sodium salicylate and niflumic acid, lowered the temperature threshold for induction of Hsp70 in glia but failed to do so in motor neurons. At concentrations that increased Hsp70 in heat shocked glial cells, sodium salicylate failed to delay death of motor neurons exposed to hyperthermia, paraquat-mediated oxidative stress, and glutamate excitotoxicity. Neither sodium salicylate nor the cyclooxygenase-2 inhibitor, niflumic acid, protected motor neurons from the toxicity of mutated Cu/Zn-superoxide dismutase (SOD-1) linked to a familial form of the motor neuron disease, amyotrophic lateral sclerosis. Thus, treatment with 2 types of NSAIDs failed to overcome the high threshold for the activation of heat shock response in motor neurons.
Cell Stress and Chaperones 02/2005; 10(3):185-96. · 3.01 Impact Factor
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ABSTRACT: Heat shock protein 70 (Hsp70) protects cultured motor neurons from the toxic effects of mutations in Cu/Zn-superoxide dismutase (SOD-1), which is responsible for a familial form of the disease, amyotrophic lateral sclerosis (ALS). Here, the endogenous heat shock response of motor neurons was investigated to determine whether a high threshold for activating this protective mechanism contributes to their vulnerability to stresses associated with ALS. When heat shocked, cultured motor neurons failed to express Hsp70 or transactivate a green fluorescent protein reporter gene driven by the Hsp70 promoter, although Hsp70 was induced in glial cells. No increase in Hsp70 occurred in motor neurons after exposure to excitotoxic glutamate or expression of mutant SOD-1 with a glycine--> alanine substitution at residue 93 (G93A), nor was Hsp70 increased in spinal cords of G93A SOD-1 transgenic mice or sporadic or familial ALS patients. In contrast, strong Hsp70 induction occurred in motor neurons with expression of a constitutively active form of heat shock transcription factor (HSF)-1 or when proteasome activity was sufficiently inhibited to induce accumulation of an alternative transcription factor HSF2. These results indicate that the high threshold for induction of the stress response in motor neurons stems from an impaired ability to activate the main heat shock-stress sensor, HSF1.
Journal of Neuroscience 08/2003; 23(13):5789-98. · 7.11 Impact Factor
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Zarah Batulan
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ABSTRACT: Heat shock proteins (HSPs) play important roles in the maintenance and preservation of cellular homeostasis. Since previous findings demonstrated a neuroprotective role of the stress-inducible Hsp70 in an in vitro model of the neurodegenerative disease, amyotrophic lateral sclerosis (ALS) (Bruening et al., 1999), the objectives of this research was to study the endogenous heat shock response of motor neurons exposed to various stresses and to determine if known pharmacological inducers of HSPs can enhance the stress response of these cells. Neither heat shock nor glutamate excitotoxicity induced Hsp70 in motor neurons. In primary spinal cord cultures, expression of Cu/Zn superoxide dismutase (SOD-1) with mutations responsible for familial ALS also failed to result in Hsp70 induction, nor was Hsp70 expressed in spinal motor neurons of mutant SOD-1 (mSOD) transgenic mice or patients with sporadic or familial ALS. The lack of a strong heat shock response was associated with an inability to activate the main stress-sensing transcription factor, heat shock transcription factor-1 (HSF1). Overexpression of an activated form of HSF1, but not wildtype HSF1, in cultured motor neurons resulted in significant protection from mutant SOD-1 toxicity, which was accompanied by increased expression of Hsp70, Hsp40, and to a lesser degree, Hsp25. The effects of two nonsteroidal anti-inflammatory drugs, sodium salicylate, shown in cell lines to increase HSF1 binding to promoters of HSP genes (Jurivich et al., 1992), and niflumic acid, a preferential COX-2 inhibitor, on the heat shock response of motor neurons were assessed. Both drugs lowered the temperature threshold for induction of Hsp70 in glia and non-neuronal cells, but not in motor neurons. Concomitantly, neither drug prevented mSOD-mediated motor neuron death. Pyrrolidine dithiocarbamate (PDTC), previously shown to upregulate Hsp70 in cell lines through increased DNA binding and activation of HSF1 (DeMeester et al., 1998; Stuhlmeier, 2000), induced robust Hsp70 expression in motor neurons, but only at cytotoxic concentrations. Geldanamycin, which binds to Hsp90, disrupting its associations with various proteins including HSF1 (Zou et al., 1998) and several kinases (reviewed in (Blagosklonny, 2002)), induced Hsp70 and Hsp40 expression and protected against the toxic effects of mutant SOD-1 in motor neurons. The results indicate that the high threshold for induction of the stress response in motor neurons stems from an impaired ability to activate the main heat shock/stress sensor, HSF1, a property of motor neurons that may contribute to their vulnerability to disease. Drugs which target the heat shock response at the level of HSF-1 activation may be considered as therapeutic prospects in ALS and other motor neuron diseases.
