Sub-synaptic, multiplexed analysis of proteins reveals Fragile X related protein 2 is mislocalized in Fmr1 KO synapses

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Abstract

The distribution of proteins within sub-synaptic compartments is an essential aspect of their neurological function. Current methodology such as electron microscopy (EM) and super-resolution imaging techniques can provide precise localization of proteins, but are often limited to a small number of one-time observations with narrow spatial and molecular coverage. The diversity of synaptic proteins and synapse types demands synapse analysis on a scale that is prohibitive with current methods. Here, we demonstrate SubSynMAP, a fast, multiplexed sub-synaptic protein analysis method using wide-field data from deconvolution array tomography (ATD). SubSynMAP generates probability distributions of proteins that reveal their functional range within the averaged synapse of a particular class. This enables the differentiation of closely juxtaposed proteins. Using this method, we analyzed 15 synaptic proteins in normal and Fragile X mental retardation syndrome (FXS) model mouse cortex, and revealed disease specific modifications of sub-synaptic protein distributions across synapse classes and cortical layers.

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Gordon X Wang

Department of Psychiatry and Behavioral Sciences, Stanford University School of Medicine, Stanford, United States

For correspondence
drwonder@stanford.edu

Competing interests
The authors declare that no competing interests exist.

"This ORCID iD identifies the author of this article:" 0000-0002-2707-7118
Stephen J Smith

Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, United States

Competing interests
The authors declare that no competing interests exist.
Philippe Mourrain

Department of Psychiatry and Behavioral Sciences, Stanford University School of Medicine, Stanford, United States

Competing interests
The authors declare that no competing interests exist.

Funding

National Institute of Mental Health (1R01MH099647)

Stephen J Smith

John Merck Fund

Gordon X Wang
Philippe Mourrain

FRAXA Research Foundation

Gordon X Wang

National Institute of Neurological Disorders and Stroke (1R01NS062798)

Philippe Mourrain

National Institute of Neurological Disorders and Stroke (1R01NS075252)

Philippe Mourrain

National Institute of Neurological Disorders and Stroke (1R21NS063210)

Philippe Mourrain

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Ethics

Animal experimentation: Animals were studied in accordance with animal use guide lines issued by the National Institutes of Health. All animals were handled with care in accordance with IACUC protocols at Stanford University. A minimum number of animal was used as necessitated by the experiments, and all animals were anesthetized using isofluorane to minimize suffering.

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This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.