Pharmacological Approaches for the Modulation of the Potassium Channel KV4.x and KChIPs

Figures

• 

  Figure 1. Alignment of human KV4. Alignment with Clustal within Jalview. Residues that are identical in the four sequences are boxed [16].

• 

  Figure 2. Alignment of human KChIPs. Alignment with ClustalO within Jalview. Residues that are identical in the four sequences are boxed [16].

• 

  Figure 3. Three-dimensional structures of free KChIPs. The four EF-hand motifs are shown, as representative examples: (A) KChIP3 (PDB ID 2JUL) [51] and (B) KChIP4a (PDB ID 3DD4) [52]. Different from KChIP1-3, KChIP4a C-terminal α-helix H10 (red) swings outward at ~45° from the hydrophobic pocket. Ca2+ ions are shown as yellow spheres.

• 

  Figure 4. Three-dimensional structures of KChIPs–KV4 complexes. (A) KChIP1(34–216)–KV4.2N30 complex structure (PDB ID 1S6C) showing the four EF-hand motifs (helices H2–H9) and the central hydrophobic pocket occupied by H10 (red) and KV4.2N30 (light blue) helices [53]. (B) Left panel: Overall structure of the KChIP1–KV4.3 T1 complex as seen from the cytosolic side (PDB ID 2I2R) [55]. KChIP1s are shown in dark blue and KV4.3 members are shown in white. Ca2+ ions are shown as yellow spheres and Zn2+ ions are shown in orange. Boxes represent interaction site 1 and interaction site 2. Top right panel: Interaction site 1 showing KChIP1 in dark blue and KV4.3 T1N helix, T1N linker, and T1 domain in white. Ca2+ ions are shown as yellow spheres. Bottom right panel: Interaction site 2 showing the main interface KChIP1(34–216)-KV4.2N30 residues. KChIP1 in dark blue and KV4.2N30 residues in white.

• 

  Figure 5. Original KV4.3 (A) and KV4.3+KChIP3 (B) records generated after applying the pulse protocol shown in the upper part of the Figure. Note that KV4.3+KChIP3 channels exhibit a slower inactivation rate than KV4.3 ones.

• 

  Figure 6. Cartoon and surface representation of (A) KChIP1 (PDB ID 2I2R) residues suggested by molecular modeling to be involved in CL-888 binding [59]. (B) KChIP3 (PDB ID 2JUL) residues suggested by molecular modeling to participate in 8-anilino-1-naphthalene sulfonate (1,8- ANS) binding [62].

• 

  Figure 7. Binding site of IQM-PC330 and IQM-PC332 [69]. (A) Cartoon representation of the molecular docking complex of KChIP3 bound to IQM-PC330 (right) and IQM-PC332 (left). (B) Direct binding of IQM-PC330 and IQM-PC332 in surface plasmon resonance (SPR) assays to immobilized wtKChIP3 compared with Tyr118Ala and Tyr130Ala KChIP3 mutants. (C) Electrophysiological effects of IQM-PC330 and IQM-PC332 on KV4.3/KChIP3 wt and KV4.3/KChIP3 mutant channels. Data are shown as mean ± SEM. * p < 0.05 vs. block produced in KV4.3/DREAM wt channels; ** p < 0.01 vs. block produced in KV4.3/DREAM wt channels; *** p < 0.001 vs. block produced in KV4.3/DREAM wt channels.

• 

  Figure 8. Electrophysiological effects of IQM-PC330 on KV4.3 and KV4.3/KChIP3 channels [69]. (A) Bar chart comparing the inhibition of KV4.3/KChIP3 currents produced by IQM-PC330, measured at the maximum peak current and at the charge. Inset shows original current recordings. (B) Original normalized current records: control (black line) and with IQM-PC330 (blue line). (C) Left panel shows current recordings elicited by the recovery protocol shown in the top. Right panel shows the effects of IQM-PC330 on the recovery process. Dashed lines represent the recovery process of KV4.3 current without KChIP3. Note that IQM-PC330 slows the recovery process, reverting KChIP3 effects. Data are shown as mean ± SEM. * p < 0.05; ** p < 0.01.

• 

  Figure 9. Concentration–dependence of inhibition and/or increase of the current or the charge through KV4.3/KChIP3 channels produced by IQM-266 [70]. (A) Concentration–response curve of the effects of IQM-266 (continuous line) on KV4.3/KChIP3 channels. The dashed line represents the fit of the data to a Hill equation with nH = 1 (n = 34). (B) Effects on the current induced by IQM-266 in KV4.3/KChIP3 channels at the peak current and in the charge (measured as the area of the current after applying 250 ms pulses to +60 mV; n = 34). (C) Current records of KV4.3/KChIP3 to +60 mV in the absence and in the presence of IQM-266 (3 μM). (D) Current records of KV4.3/KChIP3 to +60 mV in the absence and in the presence of IQM-266 (10 μM). * p < 0.05 when comparing the effect of IQM-266 on the peak current and on the charge through KV4.3/KChIP3 channels.

• 

  Figure 10. IQM-266 increases the closed-state inactivation of the KV4.3/KChIP3 channels [70]. (A) Current records generated by KV4.3/KChIP3 channels in the absence and in the presence of IQM-266 after applying the pulse protocols shown in the upper part of the figure. (B) Time course of closed-state inactivation of KV4.3/KChIP3 channels at a membrane potential (−40 mV) at which this current inactivates, but does not conduct. Note how IQM-266 increases the closed-state inactivation.