Role of regulatory C‐terminal motifs in synaptic confinement of LRRTM2

• Published in: Biology of the cell Vol. 113; no. 12; pp. 492 - 506
• Main Authors: Liouta, Konstantina, Chabbert, Julia, Benquet, Sebastien, Tessier, Béatrice, Studer, Vincent, Sainlos, Matthieu, De Wit, Joris, Thoumine, Olivier, Chamma, Ingrid
• Format: Journal Article
• Language: English
• Published: England 01.12.2021
• Subjects: cell adhesion; Cell Adhesion Molecules, Neuronal - metabolism; Cells, Cultured; Hippocampus - metabolism; light microscopy; membrane protein; Membrane Proteins - metabolism; Nerve Tissue Proteins - genetics; Nerve Tissue Proteins - metabolism; Neural Cell Adhesion Molecules - metabolism; synapse; Synapses; membrane protein; synapse; light microscopy; cell adhesion
• ISSN: 0248-4900; 1768-322X; 1768-322X
• DOI: 10.1111/boc.202100026

Leucine Rich Repeat Transmembrane proteins (LRRTMs) are neuronal cell adhesion molecules involved in synapse development and plasticity. LRRTM2 is the most synaptogenic isoform of the family, and its expression is strongly restricted to excitatory synapses in mature neurons. However, the mechanisms by which LRRTM2 is trafficked and stabilized at synapses remain unknown. Here, we examine the role of LRRTM2 intracellular domain on its membrane expression and stabilization at excitatory synapses, using a knock‐down strategy combined to single molecule tracking and super‐resolution dSTORM microscopy. We show that LRRTM2 operates an important shift in mobility after synaptogenesis in hippocampal neurons. Knock‐down of LRRTM2 during synapse formation reduced excitatory synapse density in mature neurons. Deletion of LRRTM2 C‐terminal domain abolished the compartmentalization of LRRTM2 in dendrites and disrupted its synaptic enrichment. Furtheremore, we show that LRRTM2 diffusion is increased in the absence of its intracellular domain, and that the protein is more dispersed at synapses. Surprisingly, LRRTM2 confinement at synapses was strongly dependent on a YxxC motif in the C‐terminal domain, but was independent of the PDZ‐like binding motif ECEV. Finally, the nanoscale organization of LRRTM2 at excitatory synapses depended on its C‐terminal domain, with involvement of both the PDZ‐binding and YxxC motifs. Altogether, these results demonstrate that LRRTM2 trafficking and enrichment at excitatory synapses are dependent on its intracellular domain. The Leucine Rich Repeat Transmembrane protein LRRTM2 is a cell adhesion molecule exclusively localized at excitatory post‐synapses, where it forms compact nanoclusters. In this study, we find that LRRTM2 expression and compartmentalization are developmentally regulated, and that synaptic confinement depends on a critical YxxC motif in the C‐terminal domain. Thus, this study identifies critical motifs for LRRTM2 diffusion, and highlights novel regulatory mechanisms of LRRTM2 membrane expression.