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Related Experiment Video

Updated: Sep 23, 2025

Inhibitory Synapse Formation in a Co-culture Model Incorporating GABAergic Medium Spiny Neurons and HEK293 Cells Stably Expressing GABAA Receptors
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Identification of Synaptic DGKθ Interactors That Stimulate DGKθ Activity.

Casey N Barber1,2, Hana L Goldschmidt2, Qianqian Ma1

  • 1Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore, MD, United States.

Frontiers in Synaptic Neuroscience
|May 16, 2022
PubMed
Summary

Diacylglycerol kinase θ (DGKθ) regulates synaptic vesicle recycling. This study identified synaptotagmin-1 (Syt1) as a key protein that enhances DGKθ activity, crucial for efficient endocytosis.

Keywords:
APEX2Syt1lipidsneurotransmissionproximity labelingsynapsesynaptic vesicles

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Quantifying Synapses: an Immunocytochemistry-based Assay to Quantify Synapse Number
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Area of Science:

  • Neuroscience
  • Cell Biology
  • Biochemistry

Background:

  • Lipid metabolism and membrane curvature are vital for synaptic vesicle recycling.
  • Diacylglycerol kinase θ (DGKθ) produces phosphatidic acid (PtdOH), influencing membrane curvature and endocytosis.
  • DGKθ's kinase activity is essential for its role in synaptic regulation, but its protein interactors remain unknown.

Purpose of the Study:

  • To identify endogenous protein interactors of DGKθ in neurons.
  • To investigate how these interactors modulate DGKθ's kinase activity.
  • To elucidate the role of DGKθ interactors in synaptic vesicle recycling.

Main Methods:

  • APEX2 proximity labeling in neurons to identify DGKθ interactors.
  • Mass spectrometry to analyze protein complexes.
  • In vitro kinase assays to measure DGKθ activity modulation.

Main Results:

  • Seven endogenous DGKθ interactors were identified in neurons.
  • Synaptotagmin-1 (Syt1) was found to significantly increase DGKθ kinase activity (10-fold).
  • This is the first validation of endogenous DGKθ interactors at the mammalian synapse.

Conclusions:

  • Synaptotagmin-1 (Syt1) is a novel regulator of DGKθ kinase activity.
  • A coordinated function between DGKθ-produced PtdOH and Syt1 is suggested for synaptic vesicle recycling.
  • These findings provide new insights into the molecular mechanisms governing synaptic plasticity and function.