Densin-180 forms a ternary complex with the (alpha)-subunit of Ca2+/calmodulin-dependent protein kinase II and (alpha)-actinin

J Neurosci. 2001 Jan 15;21(2):423-33. doi: 10.1523/JNEUROSCI.21-02-00423.2001.

Abstract

Densin-180 is a transmembrane protein that is tightly associated with the postsynaptic density in CNS neurons and is postulated to function as a synaptic adhesion molecule. Here we report the identification of the alpha-subunit of Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) and alpha-actinin-4 as potential binding partners for the densin-180 intracellular segment. We demonstrate by yeast two-hybrid and biochemical assays that the intracellular portion of densin-180, the alpha-subunit of CaMKII (CaMKIIalpha), and alpha-actinin interact with each other at distinct binding sites and can form a ternary complex stabilized by multiple interactions. Densin-180 binds specifically to the association domain of CaMKIIalpha and does not bind with high affinity to holoenzymes of CaMKII that contain beta-subunit. The PDZ (PSD-95, DIg, Z0-1) domain of densin contributes to its binding to alpha-actinin. A distinct domain of alpha-actinin interacts with the kinase domains of both alpha- and beta-subunits of CaMKII. Autophosphorylation of CaMKII increases its affinity for densin-180 from an EC(50) of >1 micrometer to an EC(50) of <75-150 nM. In contrast, phosphorylation of densin-180 by CaMKII at serine-1397 only slightly decreases its affinity for CaMKII. The specific interaction of densin-180 with holoenzymes of CaMKII containing only alpha-subunit and the increased affinity of CaMKII for densin-180 after autophosphorylation suggest that densin-180 may be involved in localization of activated CaMKII synthesized in dendrites.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Actinin / genetics
  • Actinin / metabolism*
  • Animals
  • Binding Sites / drug effects
  • Calcium-Calmodulin-Dependent Protein Kinase Type 2
  • Calcium-Calmodulin-Dependent Protein Kinases / genetics
  • Calcium-Calmodulin-Dependent Protein Kinases / metabolism*
  • Humans
  • Macromolecular Substances
  • Microfilament Proteins*
  • Phosphorylation
  • Precipitin Tests
  • Prosencephalon / chemistry
  • Protein Binding
  • Protein Structure, Tertiary / physiology
  • Protein Subunits*
  • Rats
  • Recombinant Fusion Proteins / chemistry
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Sequence Analysis, Protein
  • Sialoglycoproteins / genetics
  • Sialoglycoproteins / metabolism*
  • Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
  • Synaptosomes / chemistry
  • Synaptosomes / metabolism
  • Two-Hybrid System Techniques

Substances

  • ACTN4 protein, human
  • Actn4 protein, rat
  • LRRC7 protein, human
  • Lrrc7 protein, rat
  • Macromolecular Substances
  • Microfilament Proteins
  • Protein Subunits
  • Recombinant Fusion Proteins
  • Sialoglycoproteins
  • Actinin
  • Calcium-Calmodulin-Dependent Protein Kinase Type 2
  • Calcium-Calmodulin-Dependent Protein Kinases