Fig. 5
From: Dynamic constriction and fission of endoplasmic reticulum membranes by reticulon
Constriction-by-friction mechanism of Rtnl1-driven membrane fission. a Simultaneous increase of the pulling force and the curvature (both normalized to their initial static values) during the elongation of control (cyan, n = 3 tubes) and Rtnl1-containing (black, n = 3 tubes) nanotubes at constant speed vt. Error bars represent S.E.M. The image sequence shows the nanotube region used for the curvature calculations (Rh-DOPE fluorescence, scale bar 5 μm). The cartoon illustrates the dynamic force balance (fp = ft + ff), where the pulling force is equal to the sum of the tensile and friction forces. b Frame sequence (100 ms/frame) showing scission (arrow) of Rtnl1-nanotube. Scale bar 2 μm. The histogram displays the increased membrane curvature near the membrane reservoir on the pulling pipette (green arrow, n = 7 tubes) as compared to the curvature far from the reservoir (blue arrow, n = 20 tubes). Boxplots show IQR, whiskers indicate minimum and maximum of the dataset. c The increase of the axial force during elongation of Rtnl1-nanotubes at 0.1 μm/s (gray) and 8 μm/s (black) speeds caused fission seen as an abrupt decrease of the force to zero (arrows). d Dependence of the axial force measured at the moment of fission of Rtnl1-containing tubes (fRtnl1, black, n = 10 (vt = 0.1 μm/s), n = 5 (vt = 8 μm/s) n = 12 (vt = 106 μm/s)) or maximal force measured during 10 μm elongation of lipid tubes (flip, cyan, n = 9 (vt = 0.1 μm/s), n = 34 (vt = 8 μm/s) n = 24 (vt = 106 μm/s)) on the elongation speed vt. The inset shows the force difference Δfmax = (fRtnl1 − flip) dependence on vt, where the red line is the logarithmic fit (Supplementary Eq. (27)). The error bars show SD, three independent Rtnl1 preparations were used. e Radial constriction of the membrane nanotube measured by fluorescence microscopy (red, the error bars show SD, n = 25 tubes for both static and dynamic constriction) and recalculated from the force increase (shown in (d)) using either kl (black) or kp (gray). f Effect of the addition of conical lipids on the pulling force at the moment of fission and fission probability. The PE amounts indicated correspond to 20 mol% DOPE (n = 17 tubes, fission in 4 out of 17 cases) 40 mol% DOPE (n = 28 tubes, fission in 13 out of 28 cases) and 40 mol% DOPE + 20 mol% plasma (n = 8 tubes, fission in 5 out of 8 cases). Statistical significance: one-way ANOVA with multiple comparison. Source data are provided as a Source Data file.
