Recruitment of Endophilin to Clathrin-Coated Pit Necks Is Required for Efficient Vesicle Uncoating after Fission

Abstract

Endophilin is a membrane binding protein with curvature-generating/sensing properties that participates in clathrin-dependent endocytosis of synaptic vesicle membranes. Endophilin also binds the GTPase dynamin and the phosphoinositide phosphatase synaptojanin, and is thought to coordinate constriction of coated pits with membrane fission (via dynamin) and subsequent uncoating (via synaptojanin). We show that although synaptojanin is recruited by endophilin at bud necks before fission, the knockout of all three mouse endophilins results in the accumulation of clathrin-coated vesicles but not of clathrin-coated pits at synapses. The absence of endophilin impairs, but does not abolish synaptic transmission and results in perinatal lethality, while partial endophilin absence causes severe neurological defects, including epilepsy, and neurodegeneration. Our data supports a model in which endophilin recruitment to coated pit necks, due to its curvature-sensing properties, primes vesicle buds for subsequent uncoating after membrane fission, without being critically required for the fission reaction itself.

Endocytic Accessory Factors and Regulation of Clathrin-Mediated Endocytosis

“…This was later confirmed by the generation of endophilin knockout mice by the de Camilli lab (Milosevic et al 2011). Here, knockout of all three endophilin isoforms led to the accumulation of CCVs, but not CCPs, in mouse synapses (Milosevic et al 2011).…”

“…It was previously established that the BAR domain protein endophilin targets synaptojanin to newly scissioned vesicles in Drosophila (Verstreken et al 2002). This was later confirmed by the generation of endophilin knockout mice by the de Camilli lab (Milosevic et al 2011). Here, knockout of all three endophilin isoforms led to the accumulation of CCVs, but not CCPs, in mouse synapses (Milosevic et al 2011).…”

“…However, many questions remain concerning the mechanism of dynamin function and the roles of the numerous binding partners of dynamin including membrane curvature sensing/inducing BAR domain proteins such as amphiphysin and endophilin (Milosevic et al 2011;Meinecke et al 2013;Neumann and Schmid 2013) and proteins involved in actin polymerization/depolymerization (reviewed in Menon and Schafer 2013). Moreover, dynamin can also affect the earlier stages of CCP growth and invagination, although the precise mechanism by which it does so remains obscure (Loerke et al 2009;Mettlen et al 2009).…”

Cellular processes associated with LRRK2 function and dysfunction

“…The G2019S mutation impeded synaptic endocytosis 116 that was restored by pharmacological inhibition of LRRK2 kinase activity in G2019S overexpressing flies. Knockout of EndoA led to neurodegeneration 73, thus linking LRRK2‐associated defects to PD. These results were recently further validated in mammalian cells in which LRRK2 was described as being able to phosphorylate neuronal‐specific EndoA1 124.…”

Endophilin A1 regulates dendritic spine morphogenesis and stability through interaction with p140Cap

“…Both endophilin A2 and A3 interact npg www.cell-research.com | Cell Research with Arc/Arg3.1 to regulate endocytic trafficking of AM-PA-type glutamate receptors and synaptic plasticity [11,12]. It was reported recently that although single knockout mice appear normal and fertile, double knockout of endophilin A1 and A2 or knockout of all three endophilins in mouse causes accumulation of clathrin-coated vesicles at the presynaptic site, indicating that they play redundant functions in vesicle uncoating after membrane fission [13]. Although a critical role of endophilin A1 in regulating synaptic vesicle endocytosis at the presynaptic terminal has been well established [14][15][16][17][18][19][20][21], its function at the postsynaptic site remains largely unexplored.…”