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

Time-lapse imaging of live cells revealed that the tubules formed by Amph-FL and N-BAR-NfM CTD were more transient in comparison to isolated N-BAR ( Fig. 4F and Movie S5). Specifically, the tubules in cells expressing N-BAR had an average lifetime of approximately 75±5 s s.e.m., whereas tubule lifetime was significantly shorter in cells expressing Amph-FL and N-BAR-NfM CTD, approximately 29±3 and 35±4 s s.e.m., respectively (Fig. 4F ). The tubules formed by N-BAR also had greater fluorescence intensity in the protein channel relative to the local background in comparison to Amph-FL and N-BAR-NfM CTD (Fig. S4B ,C). This finding indicates that the disordered domains of Amph-FL and N-BAR-NfM CTD did not promote tubule fission by enhancing protein binding to the membrane surface. Collectively, results from experiments in live cells indicate that bulky disordered domains are capable of disrupting the formation of stable tubules scaffolded by BAR domains, similar to observations in vitro ( Fig. 1C ,D). The disordered domains may have inhibited tubule formation by driving membrane fission, though future work is needed to test the role of disordered domains in physiological fission events.