Proteomics of the Synapse--A Quantitative Approach to Neuronal Plasticity

Abstract

The advances in mass spectrometry based proteomics in the past 15 years have contributed to a deeper appreciation of protein networks and the composition of functional synaptic protein complexes. However, research on protein dynamics underlying core mechanisms of synaptic plasticity in brain lag far behind. In this review, we provide a synopsis on proteomic research addressing various aspects of synaptic function. We discuss the major topics in the study of protein dynamics of the chemical synapse and the limitations of current methodology. We highlight recent developments and the future importance of multidimensional proteomics and metabolic labeling. Finally, emphasis is given on the conceptual framework of modern proteomics and its current shortcomings in the quest to gain a deeper understanding of synaptic plasticity.

Fig. 1.

Electron micrograph of rat cortex showing multiple pre- and postsynaptic structures, as well as astrocytic endfeet (*) in close contact with synapses. Note the presence of numerous synaptic vesicles in the presynaptic boutons. CAZ, cytomatrix at the active zone; PSD, postsynaptic density; SV, synaptic vesicles. Scalebar: 100 nm.

Fig. 2.

The tetrapartite synapse of principal neurons in the forebrain, consisting of the pre- and postsynaptic compartment, astrocytic endfeet, and the extracellular matrix has a tightly regulated protein composition. A microsceretory system is present in synapses and dendrites that allows for translation of mRNA, local synthesis of, processing and insertion of transmembrane proteins. Hence the turnover of the synaptic protein machinery is controlled by local and somatic de novo protein synthesis, protein degradation by the ubiquitin proteasome system, lysosomes and autophagosomes. In addition, the association of proteins with pre- and postsynaptic compartments is highly dynamic. Molecular machineries and organelles for proteostasis are shared between synapses in dendritic segments. Proteins are transported in and out of the synapse as well as by diffusion of transmembrane proteins. These processes govern the activity-dependent assembly of the pre- and postsynaptic scaffold and the synaptic surface expression of receptors, calcium channels and cell adhesion molecules. Abbreviations: CAM, cell adhesion molecules; CAZ, cytomatrix at the active zone; ECM, extracellular matrix; ER, endoplasmatic reticulum; ERGIC, endoplasmatic reticulum Golgi intermediate compartment; MT, microtubules; PSD, postsynaptic density; RE, recycling endosomes; Lys, lysomes; SV, synaptic vesicle.

Fig. 3.

Workflow for common brain and synapse proteomics approaches tackling proteins, glycans, lipids, and phosphorylation sites of synaptic proteins, as well as receptor complexes. Synaptic fractions such as synaptosomes, CAZ, and the detergent extracted PSD are prepared using sucrose or Percoll density centrifugations followed by subsequent specific MS analyses. Dashed lining indicates further necessary processing of fractions to analyze specific subproteomes such as the lipidome, the phosphoproteome, or the different glycoproteomes. In case of global brain lipidome analysis or interactome analyses of receptor complexes organic solvents or mild detergents, respectively, are used for extraction from homogenates prior to further processing and MS analysis. IP, immunoprecipitation; AC, affinity chromatography; CAZ, cytomatrix of the active zone; PSD, postsynaptic density.