Targeted tandem affinity purification of PSD-95 recovers core
postsynaptic complexes and schizophrenia susceptibility proteins
Esperanza Fernandez, Mark O Collins, Rachel T Uren, Maksym V Kopanitsa, Noboru H Komiyama,
Mike DR Croning, Lysimachos Zografos, J Douglas Armstrong, Jyoti S Choudhary and Seth GN Grant
The synapse is the connection between nerve cells (neurons). The function of the synapse is to transmit the electrical activity from one neuron to the next thus passing information through the nervous system. Inside synapses there are over a thousand proteins and they are the components from which the neurotransmission machinery is built. The organisation of these hundreds of proteins is an important issue since proteins bind and interact to form multiprotein complexes, which are molecular machines. Understanding the composition of these multiprotein complexes may shed light on diseases as well as fundamental biology of learning and other forms of behaviour.
Isolating protein complexes from cells and tissues is technically demanding and particularly from the brain, which is a very complex organ. New methods are required for complex tissues, and in this study we have further developed a system employed in yeast. Our objective was to isolate the core components of the synapse, particularly those involving proteins bound to PSD-95, which is a key molecule required for learning and memory.
We combined proteomic and mouse genetic engineering methods to isolate multiprotein complexes from the mouse brain. A Tandem Affinity Purification tag was integrated into the genomic locus of PSD-95 so that PSD-95 could be readily isolated with other proteins attached to it. This method combined with mass spectrometry analysis describes a core complex of 118 proteins that comprise containing key proteins involved in the neurotransmission.
Protein interaction network of PSD-95 interacting proteins.
PSD-95/Dlg4 is shown in red, it's primary interactors are shown in
blue and secondary interactors are shown in yellow.
Click on circles to view genetic and genomic information in G2Cdb.
The medical importance of this complex is revealed since 49 proteins are involved with a range of important common brain diseases including epilepsy, depression, schizophrenia, bipolar disease, mental retardation and neurodegenerative diseases including Alzheimer's and Huntington's diseases.
Details of the mice generation and protein purification conditions are
described in Fernandez et. al. The principal datasets for the synapses
complex(es) are found in Table 1 and Table 2. These datasets are also integrated in
Supplementary Table 1 with
links to G2Cdb
resources as well as the comparison with other proteomic datasets. G2Cdb
access enables one to identify the functional roles for individual proteins/genes in knockout mice and human genetics.
UnMASCing diseases of the brain provides discussion of the papers principal findings and their relevance to brain disease.
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