Fission and Uncoating of Synaptic Clathrin-Coated Vesicles Are Perturbed by Disruption of Interactions with the SH3 Domain of Endophilin

Gad, Helge; Ringstad, Niels; Lőw, Péter; Kjærulff, Ole; Gustafsson, Jenny; Wenk, Markus R.; Paolo, Gilbert Di; Nemoto, Yasuo; Crum, John; Ellisman, Mark H.; Camilli, Pietro De; Shupliakov, Oleg; Brodin, Lennart

doi:10.1016/s0896-6273(00)00038-6

Cited by 278 publications

(321 citation statements)

References 51 publications

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“…Other structures that could recruit endophilin include caveolae (34) or micropinocytic vesicles. Although dynamin was recruited to CCPs independent of endophilin expression, the accumulation of SJ1-145 to CCPs strongly depended on the expression of endophilin, consistent with an important role of endophilin in the targeting of SJ1 (20,21,23 tial association by its BAR domain, with the curved membrane of the vesicle stalk (5,18,25,26); (ii) interaction by its SH3 domain with dynamin (15); and (iii) binding, again by its SH3 domain, with cargo proteins present in the nascent vesicle (35)(36)(37). Because endophilin exists as a dimer (38), it may simultaneously bind two different proteins, e.g., synaptojanin and either vesicle cargo or dynamin, by the two SH3 domains of the dimer.…”

Section: Discussionsupporting

confidence: 53%

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Two synaptojanin 1 isoforms are recruited to clathrin-coated pits at different stages

Perera¹,

Zoncu

Lucast

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

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155

187

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Phosphoinositides are thought to play an important role in clathrincoated pit (CCP) dynamics. Biochemical and structural studies have shown a direct interaction of phosphatidylinositol (4,5)-bisphosphate [PI(4,5)P 2] with endocytic clathrin adaptors, whereas functional studies using cell-free systems or intact cells have demonstrated the importance of PI(4,5)P 2 synthesis and dephosphorylation in clathrin coating and uncoating, respectively. Furthermore, genetic manipulations of kinases and phosphatases involved in PI(4,5)P 2 metabolism result in major defects in synaptic vesicle recycling and other forms of clathrin-dependent endocytosis. However, live imaging studies of these enzymes at CCPs have not been conducted. We have used multicolor total internal reflection fluorescence microscopy (TIRFM) to visualize the spatialtemporal recruitment of synaptojanin 1 (SJ1), a polyphosphoinositide phosphatase, and its binding partner endophilin to CCPs. Strikingly, we observed differential temporal recruitment of the two major SJ1 splice variants to CCPs. The 145-kDa isoform, the predominant isoform expressed in the brain, was rapidly recruited as a ''burst,'' together with endophilin, at a late stage of CCP formation. In contrast, the nonneuronal ubiquitously expressed 170-kDa isoform of SJ1 was present at all stages of CCP formation. These results raise the possibility that dynamic phosphoinositide metabolism may occur throughout the lifetime of a CCP.dynamin ͉ endocytosis ͉ endophilin ͉ phosphoinositides ͉ phosphatidylinositol (4,5)-bisphosphate

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Section: Discussionsupporting

confidence: 53%

“…Endophilin is thought to be a particularly important interactor of SJ1 and to play a major role in its recruitment to sites of endocytosis (2,15,19,20). In both flies and worms, mutations of endophilin and synaptojanin have similar phenotypes, and loss of endophilin results in destabilization and mislocalization of synaptojanin (21)(22)(23)(24).…”

mentioning

confidence: 99%

Two synaptojanin 1 isoforms are recruited to clathrin-coated pits at different stages

Perera¹,

Zoncu

Lucast

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

155

187

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“…The Dyn peptide ( figure 1A) was synthesized by Elim Biopharmaceuticals (Hayward, CA) after a previously-published sequence [17] and the PP-19 peptide ( figure 1A) was synthesised by Sigma Genosys (The Woodlands, TX, USA) after a previously published sequence [18]. Cells were transfected by using the Chariot (Active Motif, Carlsbad, CA) transfection method.…”

Section: Peptide Transfectionmentioning

confidence: 99%

“…We also took advantage of this peptide transfection technique to disrupt a key component in clathrin-mediated endocytosis. The well-described 19 amino acid peptide 'PP-19' acts to disrupt normal association between synaptojanin and endophilin and blocks the formation of clathrin-coated pits [18]. Previous work in chromaffin cells has shown PP-19 to effectively block endocytosis evoked by 15 Hz APe under similar experimental conditions [19].…”

Section: Membrane Trafficking Exhibits Differential Activity-dependenmentioning

confidence: 99%

Dynamin I plays dual roles in the activity-dependent shift in exocytic mode in mouse adrenal chromaffin cells

Fulop

Doreian

Smith

2008

Archives of Biochemistry and Biophysics

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Under low stimulation, adrenal chromaffin cells release freely-soluble catecholamines through a restricted granule fusion pore while retaining the large neuropeptide-containing proteinacious granule core. Elevated activity causes dilation of the pore and release of all granule contents. Thus, physiological differential transmitter release is achieved through regulation of fusion pore dilation. We examined the mechanism for pore dilation utilizing a combined approach of peptide transfection, electrophysiology, electrochemistry and quantitative imaging techniques. We report that disruption of dynamin I function alters both fusion modes. Under low stimulation, interference with dynamin I does not affect granule fusion but blocks its re-internalization. In full collapse mode, disruption of dynamin I limits fusion pore dilation, but does not block membrane re-internalization. These data suggest that dynamin I is involved in both modes of exocytosis by regulating contraction or dilation of the fusion pore and thus contributes to activity-dependent differential transmitter release from the adrenal medulla.

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“…The Nterminus mainly participates at the base of the membrane invagination in clathrin-coated endocytosis [14][15][16] . In brain, the SH3 domain interacts selectively with proteins containing a proline-rich domain (PRD), such as synaptojanin, amphiphysin and GTPase dynamin [17][18][19][20][21][22] .…”

Section: Introductionmentioning

confidence: 99%

Endophilin isoforms have distinct characteristics in interactions with N-type Ca2+ channels and dynamin I

Tian¹,

Zhang²,

Fan

et al. 2012

Neurosci. Bull.

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Objective Formation of the endophilin II-Ca 2+ channel complex is Ca 2+ -dependent in clathrin-mediated endocytosis. However, little is known about whether the other two endophilin isoforms have the same features. The present study aimed to investigate the characteristics of the interactions of all three isoforms with Ca 2+ channels and dynamin I. Methods N-type Ca 2+ channel C-terminal fragments (NCFs) synthesized with a 3 H-leucine-labeled kit, were incubated with endophilin-GST fusion proteins, followed by pull-down assay. Results were counted on a scintillation counter. In addition, the different endophilin isoforms were each co-transfected with dynamin I into 293T cells, followed by flow cytometry and co-immunoprecipitation assay. Immunostaining was performed and an image analysis program was used to evaluate the overlap coefficient of cells expressing endophilin and dynamin I. Results All three isoforms interacted with NCF. Endophilins I and II demonstrated clear Ca 2+ -dependent interactions with NCF, whereas endophilin III did not. Co-immunoprecipitation showed that, compared to endophilin I/II, the interaction between endophilin III and dynamin I was significantly increased. Similar results were obtained from flow cytometry. Furthermore, endophilin III had a higher overlap coefficient with dynamin I in co-transfected 293T cells. Conclusion Endophilin isoforms have distinct characteristics in interactions with NCF and dynamin I. Endophilin III binding to NCF is Ca 2+ -independent, implying that it plays a different role in clathrin-mediated endocytosis.

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Fission and Uncoating of Synaptic Clathrin-Coated Vesicles Are Perturbed by Disruption of Interactions with the SH3 Domain of Endophilin

Cited by 278 publications

References 51 publications

Two synaptojanin 1 isoforms are recruited to clathrin-coated pits at different stages

Two synaptojanin 1 isoforms are recruited to clathrin-coated pits at different stages

Dynamin I plays dual roles in the activity-dependent shift in exocytic mode in mouse adrenal chromaffin cells

Endophilin isoforms have distinct characteristics in interactions with N-type Ca2+ channels and dynamin I

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