Neuronal synapse interaction reconstituted between live cells and supported lipid bilayers

Abstract

In the nervous system, homophilic and heterophilic adhesion molecules participate in the induction and differentiation of presynaptic transmitter release sites. We focus on the heterophilic interaction between postsynaptic neuroligin-1 (Nlg) and presynaptic β-neurexin (Nrx). Nlg has previously been shown to trigger presynaptic differentiation in a Nrx-expressing axon even when presented on a non-neuronal cell or on beads coated with lipid bilayers. We have now developed a new method to measure single molecule and ensemble distribution of Nrx and Nlg at the contact site between a non-neuronal Nrx-expressing cell and a flat supported glycosylphosphoinositol–neuroligin-1 (GPI-Nlg) lipid bilayer and relate them to adhesion as measured by cell migration and gravity dissociation. We find that within minutes after cell-bilayer contact, Nrx accumulates at the contact site and the contact area is expanded. The strength of cell-bilayer adhesion depends on the morphology of Nrx accumulation, with the focal concentration strengthening adhesion. The results suggest that Nlg-Nrx interaction rapidly establishes a weak, but specific, adhesion between dynamic pre- and postsynaptic processes, which may ultimately require additional molecules for synapse stabilization.

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Figure 1: Reconstitution of GPI-Nlg in a fluid-supported lipid bilayer.
Figure 2: Distribution of GFP-Nrx in a HEK293 cell in contact with GPI-Nlg–supported lipid bilayer.
Figure 3: Cell immobilization associated with formation of Nrx-Nlg puncta.
Figure 4: Correlation between distribution of GFP-Nrx in cells and cell immobilization.
Figure 5: Inversion assay: relation between Nrx-Nlg distribution and adhesion strength.

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Acknowledgements

We thank C. Dean and P. Scheiffele for the GPI-Nlg construct, C. Dean for help with initial protein preparation, A. Pralle for help with the TIRF microscopy, H. Aaron for help with the confocal microscopy, and C. Dean, P. Scheiffele, E. Douglas, M. Forstner and S. Rozovsky for fruitful discussions. This work was supported by the Chemical Sciences, Geosciences and Biosciences Division, Office of Basic Energy Sciences, US Department of Energy (contract No. DE-AC03-76SF00098).

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Correspondence to Ehud Y Isacoff or Jay T Groves.

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Pautot, S., Lee, H., Isacoff, E. et al. Neuronal synapse interaction reconstituted between live cells and supported lipid bilayers. Nat Chem Biol 1, 283–289 (2005). https://doi.org/10.1038/nchembio737

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