Figure 6

Diminished effects of ADPR in the K908N variant of nvTRPM2. (a) Upper. Cell surface expression, assessed with biotinylation assays, of wild-type nvTRPM2 and the variant K908N (as indicated), each containing a C-terminally attached 3xHA tag (only for Western blot analysis). Western blots on the NeutrAvidin-bound fractions (right) and on total HEK-293 cell lysates (left) were probed with anti-HA antibody. Full-length blots are presented in Supplementary Figure S2. Lower. Sketch indicating the approximate location of the point mutation K908N within the nvTRPM2 structure. (b) On nvTRPM2-K908N, ADPR acts with diminished potency and efficacy. Plotted are peak current densities (mean ± S.E.M; n = 4–9) of wild-type and mutant, in response to various concentrations of ADPR in the pipette solution (as indicated). (c) Current amplitude as well as lag time of current development depend on the ADPR concentrations, as shown in representative traces with various concentrations of ADPR in the pipette solution (as indicated). (d) Summary of experiments as derived from panel c. Note the logarithmic ordinate. Significant differences are indicated with asterisks (*** P < 0.001; Student’s t-test, n = 5–7). Error bars are s.e.