Composition and regulation of connexin 36-containing gap junctions

• Applicant: Dr. Stephan Tetenborg
• Subject Area: Molecular Biology and Physiology of Neurons and Glial Cells ; Cell Biology
• Term: from 2021 to 2023
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 458153629

Neurons communicate via cellular junctions, called synapses, which can be divided into two functional types: chemical and electrical synapses. Chemical synapses require specific substances, so-called neurotransmitters, to convey information from a presynaptic to a postsynaptic neuron. Electrical synapses, on the contrary, rely on intercellular channels that transfer signals in form of ionic currents to adjacent neurons. Decades of intense research led to a profound understanding of the molecular machinery regulating chemical synaptic transmission. It has been shown that electrical synapses underlie similar regulatory principles in terms of plasticity. However, only a few key regulators have been identified in recent years and we are still lacking a detailed understanding of protein-protein interactions at electrical synapses. Gap junctions represent the morphological substrates of electrical synapses. They consist of intercellular channels that form a conductive link for ionic currents. The molecular components of these junctions are connexins, a family of transmembrane proteins that can assemble into oligomers. Although different connexins are expressed in the nervous system, connexin 36 (Cx36) is the most abundant one to form electrical synapses, which is why it is often referred to as the major neuronal connexin. As part of this project, I intend to gain a deeper understanding of the molecular mechanisms that regulate electrical synapses. My work will focus on three different projects that address several aspects of Cx36.No 1: As part of a first project, I aim to characterize the interactome of electrical synapses in the mouse retina using BioID-based proteomics. This project relies on a recently developed Cx36-BioID construct that can be used to biotinylate interacting proteins in the vicinity of Cx36. My aim is to deliver this construct into retinal neurons by means of an intravitreal virus injection and to search for proteins that interact with Cx36 in vivo.No 2: Several studies have shown that Ca2+/Calmodulin-dependent protein kinase II (CaMKII) is an essential regulator of Cx36-containing gap junctions. CaMKII phosphorylates Cx36 in an activity-dependent manner and increases electrical coupling. As part of a second project in this proposal, I will investigate the role of scaffolding mechanisms in targeting CaMKII to neuronal gap junctions. My goal is to understand how the kinase can be structurally connected to gap junctions.No 3: The C-terminal four amino acids of Cx36 constitute a PDZ binding motif (or PDZ ligand), a short sequence that mediates interactions with PDZ domains. The PDZ ligand appears to be essential for synapse formation, suggesting that this process is mediated by a specific PDZ domain containing protein. However, it remains unknown, how exactly the PDZ ligand of Cx36 coordinates the delivery of the protein to the gap junction. In this project, I plan to understand, how this domain regulates the transport of Cx36.

• DFG Programme: WBP Fellowship
• International Connection: USA
• Host: Professor John O' Brien, Ph.D.