A novel particulate form of Ca(2+)/calmodulin-dependent [correction of Ca(2+)/CaMKII-dependent] protein kinase II in neurons.
Cytoskeletal and postsynaptic density (PSD) fractions from forebrain contain discrete spherical structures that are immunopositive for Ca(2+)/calmodulin-dependent protein kinase II (CaMKII). Spherical structures viewed by rotary shadow electron microscopy have an average diameter of approximately 100 nm and, in distinction to postsynaptic densities, do not immunolabel for PSD-95. These structures were purified to near homogeneity by extraction with the detergent N-lauryl sarcosinate. Biochemical analysis revealed that CaMKII accounts for virtually all of the protein in the purified preparation, suggesting that spherical structures are clusters of self-associated CaMKII. Exposure of cultured hippocampal neurons to a mitochondrial uncoupler in glucose-free medium promotes the formation of numerous CaMKII-immunopositive structures identical in size and shape to the CaMKII clusters observed in subcellular fractions. Clustering of CaMKII would reduce its kinase function by preventing its access to fixed substrates. On the other hand, clustering would not affect the ability of the large cellular pool of CaMKII to act as a calmodulin sink, as demonstrated by the Ca(2+)-dependent binding of gold-conjugated calmodulin to CaMKII clusters. We propose that the observed clustering of CaMKII into spherical structures is a protective mechanism preventing excessive protein phosphorylation upon loss of Ca(2+) homeostasis, without compromising calmodulin regulation.
Dosemeci
A
A
Laboratory of Neurobiology, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland 20892, USA. dosemeci@marinebiombl.edu
Reese
T S
TS
Petersen
J
J
Tao-Cheng
J H
JH
D000818:Animals; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000032:analysis* / Q000187:drug effects; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone / Q000494:pharmacology; D002478:Cells, Cultured; D003599:Cytoskeleton / Q000737:chemistry*; D000076122:Disks Large Homolog 4 Protein; D006624:Hippocampus / Q000737:chemistry / Q000187:drug effects; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins / Q000032:analysis* / Q000187:drug effects; D009474:Neurons / Q000737:chemistry / Q000187:drug effects; D051381:Rats; D017207:Rats, Sprague-Dawley; D013570:Synaptic Membranes / Q000737:chemistry* / Q000187:drug effects; D014475:Uncoupling Agents / Q000494:pharmacology
D000076122:Disks Large Homolog 4 Protein; C501449:Dlg4 protein, rat; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D014475:Uncoupling Agents; C033652:postsynaptic density proteins; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone; D017871:Calcium-Calmodulin-Dependent Protein Kinases