Disordered domains disrupt N-BAR mediated membrane tubulation in live cells. (A) Schematic of mCherry (PDB 2H5Q) fusion constructs expressed in cells. (B) Confocal images of RPE cells expressing N-BAR (top), Amph-FL (middle), and N-BAR-NfM CTD (bottom). Yellow dashed boxes indicate zoomed regions to the right. White arrows indicate tubules. All cells are within the same range of protein expression level used for quantification in D and E. (C) Number of tubes per cell as a function of protein expression level, quantified as the background-subtracted protein intensity at the plasma membrane (see Materials and methods). Lines indicate linear regression with y-intercept set to 0. Shaded regions indicate 99% CI. Line color matches the respective marker color. n > 90 cells per condition from two independent transfections. (D) Number of tubes per cell within the expression level range of 200–400 brightness units. Bars indicate mean ± SEM; n > 20 cells per condition. (E) Length of tubes in cells within the expression level range of 200–400 brightness units. Points indicate data; black lines indicate means. n > 80 tubes per condition. (F) Lifetime of tubes in cells measured from time-lapse total internal reflection fluorescence (TIRF) microscopy videos (see Materials and methods). Points indicate data; black lines indicate means. n > 40 tubes per condition. All P values: two-tailed, unpaired Student’s t tests. (B) Bars, 10 µm; insets, 5 µm. See also Fig. S4 and Video 6.