In‐depth protein profiling of the postsynaptic density from mouse hippocampus using data‐independent acquisition proteomics

Abstract

Located at neuronal terminals, the postsynaptic density (PSD) is a highly complex network of cytoskeletal scaffolding and signaling proteins responsible for the transduction and modulation of glutamatergic signaling between neurons. Using ion-mobility enhanced data-independent label-free LC-MS/MS, we established a reference proteome of crude synaptosomes, synaptic junctions, and PSD derived from mouse hippocampus including TOP3-based absolute quantification values for identified proteins. The final dataset across all fractions comprised 49 491 peptides corresponding to 4558 protein groups. Of these, 2102 protein groups were identified in highly purified PSD in at least two biological replicates. Identified proteins play pivotal roles in neurological and synaptic processes providing a rich resource for studies on hippocampal PSD function as well as on the pathogenesis of neuropsychiatric disorders. All MS data have been deposited in the ProteomeXchange with identifier PXD000590 (http://proteomecentral.proteomexchange.org/dataset/PXD000590). Keywords:Brain / Data-independent acquisition / Hippocampus / Postsynaptic density Additional supporting information may be found in the online version of this article at the publisher's web-siteThe hippocampal formation has gained special attention due to its importance for learning and memory [1]. With its welldescribed trisynaptic circuit and a uniform neuronal population, the hippocampus continuously serves as a model brain region for the analysis of synaptic transmission. The majority Correspondence: Dr. Stefan Tenzer, Institute for Immunology, University Medical Center of the Johannes-Gutenberg University, Langenbeckstr. 1, 55131 Mainz, Germany E-mail: tenzer@uni-mainz.de Fax: +49-0-6131-17-6202 Abbreviations: DIA, data-independent acquisition; PSD, postsynaptic density of synaptic connections within the hippocampus are excitatory and glutamatergic and each of these contacts is mainly characterized by an electron-dense structure at the postsynaptic membrane called postsynaptic density (PSD) [2]. This spatially organized and dynamically regulated macromolecular complex is essential for the stability of postsynaptic architecture [3]. Importantly, molecular alterations of the PSD are key features of neuropsychiatric disorders such as autism,

Architecture of the Mouse Brain Synaptome

“…Synapses are also highly complex at the molecular level, with >1,000 genes encoding postsynaptic proteins in excitatory synapses (Bayés et al., 2011, Bayés et al., 2012, Bayés et al., 2017, Collins et al., 2006, Distler et al., 2014, Emes et al., 2008, Husi et al., 2000, Peng et al., 2004, Roy et al., 2018, Trinidad et al., 2008, Uezu et al., 2016, Yoshimura et al., 2004). The differential expression of these proteins raises the possibility that there is high synapse diversity within the brain (Grant, 2007, Emes et al., 2008, O’Rourke et al., 2012).…”

A mass spectrometry-based proteomic analysis of Homer2-interacting proteins in the mouse brain

“…Only three (i.e. RhoGEF33, PITRM1, and ZC2HC1A) of the 18 Homer2-interacting proteins we verified were not in the “high-confidence PSD dataset” reported by Distler et al [99]. Additionally, six (RhoGEF33, PITRM1, and ZC2HC1A as well as Homer3, LRRC7, and PITRM1) of the 18 Homer2-interacting proteins were not found in the data acquired by Föcking et al [100].…”

Optimization of calmodulin-affinity chromatography for brain and organelles

“…Several extracellular and integral to peripheral membrane proteins were also identified as unique for the synaptosomal fraction. Many of those proteins are localized to the synaptic vesicles during synaptic vesicle recycling and this is consistent with the enrichment of synapses and active zones within the synaptosomes [68]. Current information describes four different pathways of synaptic vesicle protein internalization from the plasma membrane [69].…”