Phosphorylation of synaptotagmin-1 controls a post-priming step in PKC-dependent presynaptic plasticity.
Presynaptic activation of the diacylglycerol (DAG)/protein kinase C (PKC) pathway is a central event in short-term synaptic plasticity. Two substrates, Munc13-1 and Munc18-1, are essential for DAG-induced potentiation of vesicle priming, but the role of most presynaptic PKC substrates is not understood. Here, we show that a mutation in synaptotagmin-1 (Syt1(T112A)), which prevents its PKC-dependent phosphorylation, abolishes DAG-induced potentiation of synaptic transmission in hippocampal neurons. This mutant also reduces potentiation of spontaneous release, but only if alternative Ca(2+) sensors, Doc2A/B proteins, are absent. However, unlike mutations in Munc13-1 or Munc18-1 that prevent DAG-induced potentiation, the synaptotagmin-1 mutation does not affect paired-pulse facilitation. Furthermore, experiments to probe vesicle priming (recovery after train stimulation and dual application of hypertonic solutions) also reveal no abnormalities. Expression of synaptotagmin-2, which lacks a seven amino acid sequence that contains the phosphorylation site in synaptotagmin-1, or a synaptotagmin-1 variant with these seven residues removed (Syt1(Δ109-116)), supports normal DAG-induced potentiation. These data suggest that this seven residue sequence in synaptotagmin-1 situated in the linker between the transmembrane and C2A domains is inhibitory in the unphosphorylated state and becomes permissive of potentiation upon phosphorylation. We conclude that synaptotagmin-1 phosphorylation is an essential step in PKC-dependent potentiation of synaptic transmission, acting downstream of the two other essential DAG/PKC substrates, Munc13-1 and Munc18-1.
Department of Functional Genomics, Center for Neurogenomics and Cognitive Research, Neuroscience Campus Amsterdam, VU University and VU Medical Center, Amsterdam 1081HV, The Netherlands;
Arthur P H
AP
de Jong
Department of Functional Genomics, Center for Neurogenomics and Cognitive Research, Neuroscience Campus Amsterdam, VU University and VU Medical Center, Amsterdam 1081HV, The Netherlands; Department of Clinical Genetics, Center for Neurogenomics and Cognitive Research, Neuroscience Campus Amsterdam, VU University and VU Medical Center, Amsterdam 1081HV, The Netherlands;
Marieke
M
Meijer
Department of Clinical Genetics, Center for Neurogenomics and Cognitive Research, Neuroscience Campus Amsterdam, VU University and VU Medical Center, Amsterdam 1081HV, The Netherlands;
Ingrid
I
Saarloos
Department of Clinical Genetics, Center for Neurogenomics and Cognitive Research, Neuroscience Campus Amsterdam, VU University and VU Medical Center, Amsterdam 1081HV, The Netherlands;
Lennart Niels
LN
Cornelisse
Department of Functional Genomics, Center for Neurogenomics and Cognitive Research, Neuroscience Campus Amsterdam, VU University and VU Medical Center, Amsterdam 1081HV, The Netherlands;
Ruud F G
RF
Toonen
Department of Neuroscience and Pharmacology, Faculty of Health Sciences, University of Copenhagen, Copenhagen DK-2200, Denmark; Lundbeck Foundation Center for Biomembranes in Nanomedicine, University of Copenhagen, Copenhagen DK-2200, Denmark.
Jakob B
JB
Sørensen
Department of Functional Genomics, Center for Neurogenomics and Cognitive Research, Neuroscience Campus Amsterdam, VU University and VU Medical Center, Amsterdam 1081HV, The Netherlands; Department of Clinical Genetics, Center for Neurogenomics and Cognitive Research, Neuroscience Campus Amsterdam, VU University and VU Medical Center, Amsterdam 1081HV, The Netherlands; matthijs@cncr.vu.nl.
Matthijs
M
Verhage
D000200:Action Potentials / Q000502:physiology*; D000818:Animals; D002478:Cells, Cultured; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D051938:Munc18 Proteins / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000378:metabolism; D009473:Neuronal Plasticity / Q000502:physiology*; D009474:Neurons / Q000502:physiology*; D010766:Phosphorylation / Q000502:physiology; D017729:Presynaptic Terminals / Q000502:physiology*; D011493:Protein Kinase C / Q000378:metabolism*; D050863:Synaptotagmin I / Q000378:metabolism*
D051938:Munc18 Proteins; D009419:Nerve Tissue Proteins; C497938:Stxbp1 protein, mouse; D050863:Synaptotagmin I; C494311:Syt1 protein, mouse; C503321:Unc13a protein, mouse; D011493:Protein Kinase C
Doc2; diacylglycerol; protein kinase C; short-term plasticity; synaptotagmin