Structural inhibition of dynamin-mediated membrane fission by endophilin

Abstract

Dynamin, which mediates membrane fission during endocytosis, binds endophilin and other members of the Bin-Amphiphysin-Rvs (BAR) protein family. How endophilin influences endocytic membrane fission is still unclear. Here, we show that dynamin-mediated membrane fission is potently inhibited in vitro when an excess of endophilin co-assembles with dynamin around membrane tubules. We further show by electron microscopy that endophilin intercalates between turns of the dynamin helix and impairs fission by preventing trans interactions between dynamin rungs that are thought to play critical roles in membrane constriction. In living cells, overexpression of endophilin delayed both fission and transferrin uptake. Together, our observations suggest that while endophilin helps shape endocytic tubules and recruit dynamin to endocytic sites, it can also block membrane fission when present in excess by inhibiting inter-dynamin interactions. The sequence of recruitment and the relative stoichiometry of the two proteins may be critical to regulated endocytic fission.

Quantitative and Statistical Study of the Dynamics of Clathrin-Dependent and -Independent Endocytosis Reveal a Differential Role of EndophilinA2

“…The advantage of genome editing is to avoid complications due to overexpression, since it preserves the native expression, stoichiometry, and localization of the protein. In contrast, EndoA2 overexpression induces massive tubulation, protein mislocalization, and defective vesicle scission, and it might unbalance protein recruitment to endocytosis sites (Figures S1D and S1E) (Hohendahl et al, 2017).…”

“…Interestingly, a recent in vitro study indicates that an excessive endophilin concentration inhibits dynamin-mediated fission of reconstituted lipid nanotube (Hohendahl et al, 2017). Thus, studying EndoA2 dynamics in cells through classical gene overexpression might disrupt its native localization but also impact its function.…”

Dynamic clustering of dynamin-amphiphysin helices regulates membrane constriction and fission coupled with GTP hydrolysis

“…Besides amphiphysin, other BAR domain proteins, endophilin and syndapin, are also implicated in synaptic vesicle recycling ( Dittman and Ryan, 2009 ; Koch et al, 2011 ; Milosevic et al, 2011 ). Interestingly, recent study showed that endophilin potently inhibits the dynamin-mediated membrane fission by intercalating dynamin rungs and preventing their trans-interactions required for membrane fission ( Hohendahl et al, 2017 ). Although the dynamin-mediated membrane fission is also inhibited when an excess of amphiphysin co-assembles with dynamin ( Figure 1—figure supplement 2C ), it is rather stimulatory when the molar ratio of dynamin to amphiphsyin is around 1:1 ( Yoshida et al, 2004 ).…”

Protein Amphipathic Helix Insertion: A Mechanism to Induce Membrane Fission

“…In a dynamindependent manner, endophilins A2 and A1 mediate clathrinindependent internalization route named FEME (Boucrot et al, 2015). Endophilin A1 inhibits dynamin-mediated membrane fission via intercalation between turns of the dynamin helix (Hohendahl et al, 2017). Endophilin B1 and dynamin 2 cooperatively induce the fission of Golgi membranes during autophagy (Takahashi et al, 2016), while the dynamic clustering of dynamin-amphiphysin 1 rings was suggested to regulate fission of large unilamellar vesicles (Takeda et al, 2018).…”