Fig. 1.

Identification of PKD2L1 hydrophobic gate in the S6 helix. a Left panel, membrane topology of polycystin channels. The TOP domain between S1 and S2 and the P-loop between S5 and S6 are indicated. Right panel, amino acid (aa) sequence alignment of S6 helix of human PKD2, PKD2L1 and PKD2L2. Each single underlined aa of PKD2L1 was mutated to asparagines (N). b Representative current traces obtained from oocytes injected with WT or a mutant (L557N or A558N) PKD2L1 cRNA, or water (Ctrl). Oocytes were voltage clamped at -50 mV in the presence of the extracellular solution containing (in mM) 100 N-methyl-d-glucamine (NMDG)-Cl, 2 KCl, 1 MgCl₂, 10 HEPES at pH 7.5 (“NMDG”), or Na-containing solution (equimolar Na⁺ substituting NMDG) (“Na”). Dashed lines are the baselines from which plateau current values were determined. c Averaged currents as the difference between NMDG- and Na-containing solutions obtained at −50 mV, as in b, in oocytes expressing the single mutants, as indicated. For each mutant, currents were averaged from 10 to 17 oocytes from three batches. Data are presented as mean ± SEM. d Representative I–V curves from Ctrl (water-injected oocytes), PKD2L1 WT, L557N or A558N expressing oocytes obtained at current points indicated by the vertical bars in the traces in b, using the indicated voltage ramp protocol. e Western blot of surface biotinylated and total PKD2L1 WT or indicated mutants. f Whole-mount immunofluorescence showing the oocyte surface expression of PKD2L1 WT or mutants from e. Scale bar, 50 μm