PTP{sigma} Drives Excitatory Presynaptic Assembly via Various Extracellular and Intracellular Mechanisms

Leukocyte common antigen-receptor protein tyrosine phosphatases (LAR-RPTPs) are hub proteins that organize excitatory and inhibitory synapse development through binding to various extracellular ligands. Here, we report that knockdown (KD) of the LAR-RPTP family member PTP reduced excitatory synapse number and transmission in cultured rat hippocampal neurons, whereas KD of PTP produced comparable decreases at inhibitory synapses, in both cases without altering expression levels of interacting proteins. An extensive series of rescue experiments revealed that extracellular interactions of PTP with Slitrks are important for excitatory synapse development. These experiments further showed that the intracellular D2 domain of PTP is required for induction of heterologous synapse formation by Slitrk1 or TrkC, suggesting that interaction of LAR-RPTPs with distinct intracellular presynaptic proteins, drives presynaptic machinery assembly. Consistent with this, double-KD of liprin-α2 and -α3 or KD of PTP substrates (N-cadherin and p250RhoGAP) in neurons inhibited Slitrk6-induced, PTP-mediated heterologous synapse formation activity. We propose a synaptogenesis model in presynaptic neurons involving LAR-RPTP-organized retrograde signaling cascades, in which both extracellular and intracellular mechanisms are critical in orchestrating distinct synapse types.

SIGNIFICANCE STATEMENT In this study, we sought to test the unproven hypothesis that PTP and PTP are required for excitatory and inhibitory synapse formation/transmission, respectively, in cultured hippocampal neurons, using knockdown-based loss-of-function analyses. We further performed extensive structure–function analyses, focusing on PTP-mediated actions, to address the mechanisms of presynaptic assembly at excitatory synaptic sites. Using interdisciplinary approaches, we systematically applied a varied set of PTP deletion variants, point mutants, and splice variants to demonstrate that both extracellular and intracellular mechanisms are involved in organizing presynaptic assembly. Strikingly, extracellular interactions of PTP with heparan sulfates and Slitrks, intracellular interactions of PTP with liprin-α and its associated proteins through the D2 domain, as well as distinct substrates are all critical.