Vesicular proteins exocytosed and subsequently retrieved by compensatory endocytosis are nonidentical
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The document discusses the non-identical nature of proteins exocytosed and retrieved during synaptic vesicle cycling, highlighting that most recycling synaptic vesicles lose their protein complement during exocytosis. It indicates that endocytosed vesicles are primarily composed of previously stranded molecules and questions the mechanisms behind temporal coupling and the involvement of new proteins and calcium. Future directions include exploring the implications for neurodegenerative diseases.
Vesicular proteins exocytosed and subsequently retrieved by compensatory endocytosis are nonidentical
- 1. Department of Neuroscience & Pharmachology Jessica Shipley & Rune Rasmussen Department of Neuroscience & Pharmachology Vesicular proteins exocytosed and subsequently retrived by compensatory endocytosis are nonidentical Wienish & Klingauf (2006)
- 2. Vesicle cycling: (Victoria et al. 2011) Department of Neuroscience & Pharmachology
- 3. Vesicle cycling: Kiss and Run (Molecular identity) Department of Neuroscience & Pharmachology
- 4. Kiss and Run Preassembled pool (Non-identity) Department of Neuroscience & Pharmachology Vesicle cycling:
- 5. Question: Are the the same molecules that are exocytosed retrieved during compensatory endocytosis? Department of Neuroscience & Pharmachology
- 6. g Measuring endocytic activity: Department of Neuroscience & Pharmachology spH: Synaptobrevin + pHluorin
- 7. g Department of Neuroscience & Pharmachology Measuring endocytic activity: spH: Synaptobrevin + pHluorin
- 8. g Department of Neuroscience & Pharmachology Measuring endocytic activity: spH: Synaptobrevin + pHluorin
- 9. spH: Synaptobrevin + pHluorin (pH sensitive GFP) (Burrone et al. 2007) Department of Neuroscience & Pharmachology Measuring endocytic activity:
- 10. spH-TEV: Synaptobrevin + protease cleavage site (TEV) + pHluorin
- 11. Tracking the fate of proteins after fusion: Presynaptic boutons Department of Neuroscience & Pharmachology
- 12. 33.1 % stranded in plasma membrane decrease pH increase pH Department of Neuroscience & Pharmachology Tracking the fate of proteins after fusion:
- 13. Silencing the stranded proteins with protease Department of Neuroscience & Pharmachology Tracking the fate of proteins after fusion:
- 14. Vesicular proteins are lost after fusion: 3 functional boutons Incomplete recovery Department of Neuroscience & Pharmachology
- 15. Proteins exo- and endocytosed are non-identical: Complete recovery (100 AP) - Identity? Department of Neuroscience & Pharmachology
- 16. Incomplete recovery (100 AP) - Silenced proteins taken up Department of Neuroscience & Pharmachology Proteins exo- and endocytosed are non-identical: TEV protease
- 17. Complete recovery (800 AP) - Protein pools mixed Department of Neuroscience & Pharmachology Proteins exo- and endocytosed are non-identical:
- 18. Protease treatment and pHluorin? Photobleaching: silencing the membrane proteins (90 s @ 488 nm) Department of Neuroscience & Pharmachology
- 19. Did not affect synaptobrevin recycling Department of Neuroscience & Pharmachology Protease treatment and pHluorin?
- 20. Stranded pool size is under modulatory control: 30 % vs. 10 % SpH over-expressed SpH + Synaptophysin over- expressed Department of Neuroscience & Pharmachology
- 21. Stranded pool size under native conditions: Pool size = 10 % Stranded synaptotagmin-1 All synaptotagmin-1 Department of Neuroscience & Pharmachology
- 22. A readily retrievable pool of vesicles? Stranded vesicle proteins are initially preferentially recycled 40 AP = Stranded proteins Department of Neuroscience & Pharmachology
- 23. Conclusion: 1. Most recycling synaptic vesicles lose their proteins complement during exocytosis (non-identical proteins) - Not kiss-and-run model 2. Endocytosed vesicles were initially largely composed of previously stranded molecules Department of Neuroscience & Pharmachology
- 24. Future directions: Readily releaseable pool : Readily retrievable pool Mechanisms for tight temporal coupling?; - Arrival of new proteins? - Intracellular calcium? Neurodegenerative diseases? Department of Neuroscience & Pharmachology