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Takeshi Uemura, Tomoko Shiroshima, Asami Maeda, Misato Yasumura, Takashi Shimada, Yuko Fukata, Masaki Fukata, Tomoyuki Yoshida, In situ screening for postsynaptic cell adhesion molecules during synapse formation, The Journal of Biochemistry, Volume 162, Issue 4, October 2017, Pages 295–302, https://doi.org/10.1093/jb/mvx030
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Abstract
Neuronal synapse formation is regulated by pre- and postsynaptic cell adhesion molecules. Presynaptic neurexins (NRXNs) and receptor protein tyrosine phosphatases (RPTPs; PTPδ, PTPσ and LAR in mammals) can induce postsynaptic differentiation through the interaction with various postsynaptic cell adhesion molecules. Here, we developed a novel in situ screening method to identify postsynaptic membranous proteins involved in synaptogenesis. Magnetic beads coated with the extracellular domains of NRXN1β(–S4) and PTPδ-A6 variants preferentially induced excitatory postsynaptic differentiation on the beads’ surface when co-cultured with cortical neurons. After inducing postsynaptic sites on these beads, protein complexes including NRXN1β(–S4)/PTPδ-A6 and their ligands on the neuronal membrane were chemically cross-linked and purified using a magnetic separator. Liquid chromatography-tandem mass spectrometry analysis of the complexes revealed two types of postsynaptic ligands for NRXN1β(–S4) and PTPδ-A6, one has an activity to induce presynaptic differentiation in a trans manner, whereas the other has no such activity. These results suggest that synapse formation is regulated by the interplay between presynaptic NRXN/PTPδ and their postsynaptic ligands with functionally different impacts on pre- and postsynaptic differentiation. Thus, our in situ screening method for identifying synapse-organizing complexes will help to understand the molecular basis for elaborate neuronal networks.