BAR scaffolds drive membrane fission by crowding disordered domains

Wilton T. Snead; Wade F. Zeno; Grace Kago; Ryan W. Perkins; J Blair Richter; Chi Zhao; Eileen M. Lafer; Jeanne C. Stachowiak

The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. . https://doi.org/10.1101/276147 doi: bioRxiv preprint Figure S4 . Imaging tubules generated in live cells by N-BAR, Amph-FL, and N-BAR-NfM CTD. (A) Spinning disc confocal image of a live RPE cell overexpressing the N-BAR domain of amphiphysin tagged with mCherry and stained with CellMask Green plasma membrane stain. Yellow dashed box indicates the zoomed region below, where the individual channels are shown. White arrows indicate lipid tubes. The co-localization of the plasma membrane stain with N-BAR-coated tubes indicates that the tubes are derived from the plasma membrane. Scale bar in top image: 10 µm. Scale bar in zoomed region: 5 µm. (B) Representative tubes in RPE cells overexpressing the indicated proteins tagged with mCherry. Images were acquired in TIRF at 37 °C. Tubules are from cells with similar protein expression level, and all images are displayed with equal contrast settings. White arrows indicate tubes. N-BAR shows greater enrichment on the tube relative to the local background compared to Amph-FL and N-BAR-NfM CTD. Scale bars: 1 µm. (C) Protein intensity on membrane tubes in live cells, quantified as the ratio of the tube intensity to the local background intensity. Points indicate individual tubes, and black lines indicate means. Data All rights reserved. No reuse allowed without permission.