Molecular mechanisms of presynaptic membrane recycling, turnover, and transport

• Applicant: Professor Volker Haucke, Ph.D.
• Subject Area: Biochemistry ; Molecular Biology and Physiology of Neurons and Glial Cells
• Term: from 2017 to 2022
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 327545797

• Final Report Year: 2022

Final Report Abstract

Neuronal communication depends on the regulated release of neurotransmitters from synaptic vesicles (SVs) by calcium-triggered exocytic fusion at specialized release sites within active zones(1-3). Exocytic fusion has to be balanced by endocytic membrane retrieval and synaptic vesicle (SV) reformation to replenish the SV pool, clear release sites from exocytosed material, and maintain presynaptic integrity. In spite of decades of research the mechanism of SV recycling remains controversial(4-6). Genetic, morphological(7), and biochemical evidence indicates an important function for clathrin and clathrin adaptors (e.g. AP-2) in presynaptic function and has led to the proposal that SVs are recycled directly from the neuronal surface by clathrin-mediated endocytosis (CME) (7, 8). In contrast, evidence from us(9) and others(10-12) suggests that SV membrane retrieval and the reformation of functional SVs are distinct, separable processes that may allow synapses to rapidly restore membrane surface area over a wide range of stimulations. Apart from the discrepant views regarding the mechanism of SV membrane retrieval other key questions regarding presynaptic membrane cycling are unsolved. It is unknown how synapses maintain membrane balance, e.g. for each SV exocytosed one SV membrane equivalent is internalized. Two non-exclusive mechanisms that can be envisioned are (i) restoration of membrane tension postexocytosis via so-far unknown tension sensors, and/ or (ii) sensing of surface-stranded SV proteins that may drive membrane internalization, e.g. via rapid changes in membrane phosphoinositides(13). Finally, it is unknown how the presynaptic compartment(14) ensures quality control to prevent the accumulation of non-functional proteins or organelles such as mitochondria or the endoplasmic reticulum that serve as calcium buffers. The main goals of the proposed research project were to (1) unravel the mechanism of CIE of SV membranes, (2) analyze the physiological function and molecular mechanisms that underlie the dual function of AP-2 in autophagy and SV reformation, and, finally, (3) to uncover how neuronal autophagosome formation and transport are regulated, determine its major substrates and analyze how autophagy preserves neuronal function and prevents neurodegeneration.

Publications

• Retrograde transport of TrkB-containing autophagosomes via the endocytic adaptor AP-2 mediates neuronal complexity and prevents neurodegeneration. Nature Communications 8,14819
  Kononenko, N.L., Claßen, G.A., Kuijpers, M., Puchkov, D., Maritzen, T., Tempes, A., Malik, A.R., Skalecka, A., Bera, S., Jaworski, J., Haucke, V.
  
• Synaptic vesicle endocytosis occurs on multiple timescales and is mediated by formin-dependent actin assembly. Neuron 93, 854-866
  Soykan, T., Kaempf, N., Sakaba, T., Vollweiter, D., Goerdeler, F., Puchkov, D., Kononenko, N.L., Haucke, V.
  
• EuroEPINOMICS-RES Consortium; GRIN Consortium.A recurrent missense variant in AP2M1 impairs clathrin-mediated endocytosis and causes developmental and epileptic encephalopathy. Am J Hum Genet. 104, 1060-1072
  Helbig, I., Lopez-Hernandez, T., Shor, O. et al., Benninger, F., Katherine L. Helbig, K.L., Haucke, V., Weber, Y.G.
  
• A presynaptic perspective on transport and assembly mechanisms for synapse formation. Neuron 109, 27-41
  Rizalar, F. S., Roosen, D., Haucke, V.
  
• Mechanism of synaptic protein turnover and its regulation by neuronal activity. Curr Op Neurobiol 69, 76-83
  Soykan T., Haucke V., Kuijpers M.
  
• Neuronal autophagy controls the axonal endoplasmic reticulum to regulate neurotransmission in healthy neurons. Autophagy 17, 1049-1051
  Kuijpers M, Haucke V.
  
• Neuronal autophagy regulates presynaptic neurotransmission by controlling the axonal endoplasmic reticulum. Neuron 109, 299-313
  Kuijpers, M., Kochlamazashvili, G., Stumpf, A., Puchkov, D., Swaminathan, A., Lucht, M.T., Krause, E., Schmitz, D., Haucke, V.
  
• The axonal endolysosomal and autophagic systems. J Neurochem. 158,589–602
  Kuijpers M., Azarnia Tehran D., Haucke V., Soykan T.
  
• Clathrin-independent endocytic retrieval of SV proteins mediated by the clathrin adaptor AP-2 at mammalian central synapses. eLife, 11:e71198
  Lopez-Hernandez, T., Takenaka, K., Mori, Y., Kongpracha, P., Nagamori, S., Haucke, V., Takamori, S.
  
• Endosomal phosphatidylinositol 3- phosphate levels control presynaptic vesicle cycling and neurotransmission. EMBO J
  Liu, G.T., Kochlamazashvili, G., Puchkov, D., Müller, R., Schultz, C., Mackintosh, A.I., Vollweiter, D., Haucke, V., Soykan, T.