In vivo proximity proteomics of nascent synapses reveals a novel regulator of cytoskeleton-mediated synaptic maturation

• Published in: Nature communications Vol. 10; no. 1; pp. 386 - 16
• Main Authors: Spence, Erin F., Dube, Shataakshi, Uezu, Akiyoshi, Locke, Margaret, Soderblom, Erik J., Soderling, Scott H.
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 23.01.2019; Nature Publishing Group; Nature Portfolio
• Subjects: 14/19; 38/70; 631/378/340; 631/378/87; 64/60; 82/58; 82/80; 82/83; Actin; Actin Capping Proteins - genetics; Actin Capping Proteins - metabolism; Animals; Biotin; Brain; Capping; Clustered Regularly Interspaced Short Palindromic Repeats; CRISPR; CRISPR-Cas Systems; Cytoskeletal Proteins - metabolism; Cytoskeleton; Cytoskeleton - metabolism; Dendritic Spines - metabolism; Excitatory Postsynaptic Potentials - physiology; Gene Expression Regulation; GTPase-Activating Proteins; HEK293 Cells; Humanities and Social Sciences; Humans; Maturation; Mice, Inbred C57BL; Microfilament Proteins - genetics; Microfilament Proteins - metabolism; Microtubules - metabolism; Molecular modelling; multidisciplinary; Neurogenesis - genetics; Neurogenesis - physiology; Neurons - metabolism; Organic chemistry; Postsynapse; Proteins; Proteome - genetics; Proteome - metabolism; Proteomics; Recruitment; Science; Science (multidisciplinary); Spine; Structure-function relationships; Synapses; Synapses - genetics; Synapses - metabolism; Synaptogenesis; α-Amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptors
• ISSN: 2041-1723; 2041-1723
• DOI: 10.1038/s41467-019-08288-w

Excitatory synapse formation during development involves the complex orchestration of both structural and functional alterations at the postsynapse. However, the molecular mechanisms that underlie excitatory synaptogenesis are only partially resolved, in part because the internal machinery of developing synapses is largely unknown. To address this, we apply a chemicogenetic approach, in vivo biotin identification (iBioID), to discover aspects of the proteome of nascent synapses. This approach uncovered sixty proteins, including a previously uncharacterized protein, CARMIL3, which interacts in vivo with the synaptic cytoskeletal regulator proteins SrGAP3 (or WRP) and actin capping protein. Using new CRISPR-based approaches, we validate that endogenous CARMIL3 is localized to developing synapses where it facilitates the recruitment of capping protein and is required for spine structural maturation and AMPAR recruitment associated with synapse unsilencing. Together these proteomic and functional studies reveal a previously unknown mechanism important for excitatory synapse development in the developing perinatal brain. The internal molecular mechanisms that underlie excitatory synaptogenesis remain poorly characterized. This study utilizes a chemogenetic approach, in vivo biotin identification (iBioID), to discover previously uncharacterized proteins at nascent synapses. CARMIL3 is identified as a cytoskeletal protein that facilitates spine maturation and AMPAR recruitment.