FIGURE 2.

Halofuginone (HF) reversibly increases whole-cell K⁺ currents in HEK cells transiently transfected with the human KCNA5 gene. (a) Representative outward K⁺ currents, elicited by depolarizing a cell from a holding potential of −70 mV to a series of test potentials ranging from −80 to +80 mV in increment of 20 mV, before (control), during (HF) and after (washout) extracellular application of 1-μM halofuginone. The halofuginone -sensitive K⁺ currents through KCNA5 channels (I_KCNA5) shown in inset are obtained by subtracting the currents recorded before halofuginone application (control) from the currents recorded during HF application (HF). (b) Averaged current–voltage (I–V) relationship curves (means ± SE, n = 8) for I_KCNA5 recorded before (control), during (HF) and after (washout) halofuginone application. The halofuginone curve is significantly different from control and washout curves. (c) Amplitude of I_KCNA5 at +80 mV under control, HF and washout conditions. Data shown are means ± SEM; n = 8 cells; for two of the 8 cells we were unable to record current after application of HF. *P < 0.05, significantly different from control and washout. (d) Averaged halofuginone -sensitive I_KCNA5 (left panel) and its I–V curve (right panel). Inset: enlarged portion of the I–V curve showing the halofuginone -sensitive I_KCNA5 at negative potentials (−80 to −40 mV). (e) Time course of halofuginone -induced effect on I_KCNA5. Normalized I_KCNA5 at +80 mV before, during and after extracellular application of 1-μM halofuginone. (f) Western bot analysis on KCNA5 in HEK cells transfected with an empty vector (control, n = 6 cells) or the human KCNA5 gene (KCNA5, n = 6 cells)