The turnover of endoplasmic reticulum (ER) ensures the correct biological activity of its distinct domains. In mammalian cells, the ER is degraded via a selective autophagy pathway (ER-phagy), mediated by two specific receptors: FAM134B, responsible for the turnover of ER sheets and SEC62 that regulates ER recovery following stress. Here, we identified reticulon 3 (RTN3) as a specific receptor for the degradation of ER tubules. Oligomerization of the long isoform of RTN3 is sufficient to trigger fragmentation of ER tubules. The long N-terminal region of RTN3 contains several newly identified LC3-interacting regions (LIR). Binding to LC3s/GABARAPs is essential for the fragmentation of ER tubules and their delivery to lysosomes. RTN3-mediated ER-phagy requires conventional autophagy components, but is independent of FAM134B. None of the other reticulon family members have the ability to induce fragmentation of ER tubules during starvation. Therefore, we assign a unique function to RTN3 during autophagy.

The GTPase dynamin has been clearly implicated in clathrin-mediated endocytosis of synaptic vesicle membranes at the presynaptic nerve terminal. Here we describe a novel 52-kDa protein in rat brain that binds the proline-rich C terminus of dynamin. Syndapin I (synaptic, dynamin-associated protein I) is highly enriched in brain where it exists in a high molecular weight complex. Syndapin I can be involved in multiple protein-protein interactions via a src homology 3 (SH3) domain at the C terminus and two predicted coiled-coil stretches. Coprecipitation studies and blot overlay analyses revealed that syndapin I binds the brain-specific proteins dynamin I, synaptojanin, and synapsin I via an SH3 domain-specific interaction. Coimmunoprecipitation of dynamin I with antibodies recognizing syndapin I and colocalization of syndapin I with dynamin I at vesicular structures in primary neurons indicate that syndapin I associates with dynamin I in vivo and may play a role in synaptic vesicle endocytosis. Furthermore, syndapin I associates with the neural Wiskott-Aldrich syndrome protein, an actin-depolymerizing protein that regulates cytoskeletal rearrangement. These characteristics of syndapin I suggest a molecular link between cytoskeletal dynamics and synaptic vesicle recycling in the nerve terminal. INTRODUCTIONNeurotransmitter release requires that synaptic vesicles fuse with the plasma membrane when intraterminal calcium rises, after which the synaptic vesicle membrane is rapidly recycled and refilled with neurotransmitter. Recovery of plasma membrane after stimulated exocytosis is commonly referred to as compensatory endocytosis. Compensatory endocytosis of synaptic vesicle membrane proteins from the plasma membrane was originally attributed to only two cytoplasmic proteins, clathrin and the heterotetrameric adaptor complex, adaptor protein 2 (AP2) 1 (for review, see Schmid, 1997). A more complex model of endocytosis became necessary when the Drosophila shibire mutant that could not recycle synaptic vesicle membranes was shown to be defective in a GTPase, dynamin (Kosaka and Ikeda, 1983;. Dynamin is now known to form tightly wound helical structures that participate in pinching off the constricted neck of a clathrincoated pit (Hinshaw and Schmid, 1995;Takei et al., 1995Takei et al., , 1998Sweitzer and Hinshaw, 1998). It is likely that dynamin is normally part of a much larger molecular machine that is responsible for compensatory endocytosis after exocytosis. Dynamin binds with high affinity via a proline-rich domain (PRD) to four brain-specific proteins: amphiphysin I , amphiphysin II (Leprince et al., 1997;Ramjaun et al., 1997;Wigge et al., 1997a) , 1997;Ringstad et al., 1997), and an undescribed protein of ϳ52 kDa (Roos and Kelly, 1998). Amphiphysin I, amphiphysin II, and endophilin bind to dynamin and other PRD-containing proteins such as synapsin I (De Camilli et al., 1983) and synaptojanin via their src homology 3 (SH3) domains.Interaction of dynamin via its PRD with SH3 domain-containing proteins is essential...

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Syndapin I (SdpI) interacts with proteins involved in endocytosis and actin dynamics and was therefore proposed to be a molecular link between the machineries for synaptic vesicle recycling and cytoskeletal organization. We here report the identification and characterization of SdpII, a ubiquitously expressed isoform of the brain-specific SdpI. Certain splice variants of rat SdpII in other species were named FAP52 and PACSIN 2. SdpII binds dynamin I, synaptojanin, synapsin I, and the neural Wiskott-Aldrich syndrome protein (N-WASP), a stimulator of Arp2/3 induced actin filament nucleation. In neuroendocrine cells, SdpII colocalizes with dynamin, consistent with a role for syndapin in dynamin-mediated endocytic processes. The src homology 3 (SH3) domain of SdpI and -II inhibited receptor-mediated internalization of transferrin, demonstrating syndapin involvement in endocytosis in vivo. Overexpression of full-length syndapins, but not the NH2-terminal part or the SH3 domains alone, had a strong effect on cortical actin organization and induced filopodia. This syndapin overexpression phenotype appears to be mediated by the Arp2/3 complex at the cell periphery because it was completely suppressed by coexpression of a cytosolic COOH-terminal fragment of N-WASP. Consistent with a role in actin dynamics, syndapins localized to sites of high actin turnover, such as filopodia tips and lamellipodia. Our results strongly suggest that syndapins link endocytosis and actin dynamics.