Sheehan, D; (2015) Membrane Dynamics of Neuroligin 2 at the Inhibitory Synapse. Doctoral thesis , UCL (University College London).

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Abstract

Recent innovations in live-cell imaging have demonstrated that the synapse undergoes constant remodelling and reorganisation. One well characterised aspect of this process is the lateral mobility of neurotransmitter gated receptors, which enables their dynamic exchange between synaptic and extrasynaptic populations. Regulation of this process, primarily via transient receptor-scaffold interactions, determines receptor number at the synapse and thus directly shapes the strength of synaptic neurotransmission. Neuroligins (NLs) are trans-synaptic proteins that project across the synaptic cleft and bind to partners positioned on the presynaptic side, physically fusing the two synapses in close apposition. Further to this function, the various NL isoforms (NL1-4) are essential regulators of the composition of the postsynaptic density. NL2 is primarily localised to inhibitory synapses, where it influences synaptic activity through interactions with, among others, gephyrin and collybistin. In contrast to neurotransmitter receptors, the membrane dynamics of the NL proteins are poorly understood. Thus, the aim of this thesis is to uncover the mechanisms underlying NL2 mobility on the membrane surface. Novel features of NL2 expression in non-neuronal cells were harnessed to create innovative systems in which the role of specific synaptic mechanisms was studied in isolation. Single particle tracking of NL2 with quantum dots in hippocampal neurons confirmed that endogenous NL2 exhibits transient confinement at inhibitory synapses, in line with previous findings on receptors. To uncover the underlying mechanisms, a number of important NL2 mutants were designed. Collectively, these experiments suggested that NL2 transient confinement primarily depends on intracellular interactions, specifically via phosphorylation and the PDZ and gephyrin binding domains, rather than trans-synaptic signalling mechanisms. Thus, the work described in this thesis contributes to the concept of the dynamic synapse and attributes a non-static membrane profile to NL2, which may ultimately influence synaptic remodelling and plasticity events.

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