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Molecular characterization of the postsynaptic density

Citation

Apperson, Michelle Louise (1996) Molecular characterization of the postsynaptic density. Dissertation (Ph.D.), California Institute of Technology. http://resolver.caltech.edu/CaltechETD:etd-01052007-083947

Abstract

The postsynaptic density (PSD) is an electron dense structure just beneath the postsynaptic membrane. Several functions have been proposed for the PSD including regulating receptor number and clustering, anchoring signal transduction molecules at the synapse and mediating adhesion between the presynaptic and postsynaptic membranes. However, little was known about the proteins that make up the PSD until the biochemical purification of a PSD fraction from brain was established in 1974. Since then, several interesting proteins have been localized to the PSD fraction. The most abundant PSD protein is the [alpha] subunit of the type II calcium/calmodulin dependent protein kinase ([alpha]CaMKII). This protein is likely to play a role in the calcium-mediated signal transduction at the synapse that mediates certain forms of synaptic plasticity. Another major PSD protein is PSD-95, a member of the guanylate kinase family (GUK) of proteins.

Here, I describe the purification and identification of three additional PSD proteins that comigrate at a molecular weight of 180 kDa on SDS-polyacrylamide gels. First, PSDgp180 is identified as the 2B subunit of the N-methyl-D-aspartate receptor (NR2B). NR2B is a major component of the PSD fraction and binds to PSD-95 in vitro. This interaction may anchor NMDA receptors at the synapse.

Next, I report the cloning and characterization of densin-180, a 180 kDa PSD protein with a novel adhesion molecule-like sequence. Densin-180 is a brain-specific sialomucin that is enriched in the PSD fraction and localized to the synapse by immunocytochemistry.

In order to study the assembly of PSD proteins at the synapse, I use antibodies against [alpha]CaMKII, PSD-95 and densin-180 for double labeling cultured hippocampal neurons. In these cultures, densin-180 protein is the first marker to be expressed and this early densin-180 expression is in a diffuse membrane pattern along dendrites. When synapse formation begins at about 5 days after plating, the densin-180 protein is clustered at synapses and PSD-95 expression is induced. PSD-95 colocalizes with densin-180 clusters. The [alpha]CaMKII protein is expressed later in synapse formation (7 to 9 days in vitro) and may be a marker of mature excitatory neurons.

In the brain, densin-180 is localized to the neuropil regions in a punctate pattern likely to represent synaptic staining. In addition, anti-densin-180 is localized to a specific set of cells and that may represent undifferentiated neurons and small processes that may represent dendritic filopodia.

The third 180 kDa PSD protein is citron, a recently identified Rho/Rac binding protein. The citron sequence contains numerous motifs found in signal transduction proteins and a myosin-like coiled coil domain. Citron may be a target for Rho/Racdependent signal transduction at the synapse and may mediate physical stabilization of the postsynaptic density.

Item Type:Thesis (Dissertation (Ph.D.))
Degree Grantor:California Institute of Technology
Division:Biology
Major Option:Molecular Biology
Thesis Availability:Restricted to Caltech community only
Research Advisor(s):
  • Kennedy, Mary B.
Thesis Committee:
  • Kennedy, Mary B. (chair)
  • Schuman, Erin Margaret
  • Lester, Henry A.
  • Zinn, Kai George
  • Dunphy, William G.
Defense Date:21 May 1996
Record Number:CaltechETD:etd-01052007-083947
Persistent URL:http://resolver.caltech.edu/CaltechETD:etd-01052007-083947
Default Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:35
Collection:CaltechTHESIS
Deposited By: Imported from ETD-db
Deposited On:05 Jan 2007
Last Modified:26 Dec 2012 02:26

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