title: Multiple spatial and kinetic subpopulations of CaMKII in spines and dendrites as resolved by single-molecule tracking PALM.
abstract: Calcium/calmodulin-dependent protein kinase II (CaMKII) is essential for synaptic plasticity underlying memory formation. Some functions of CaMKII are mediated by interactions with synaptic proteins, and activity-triggered translocation of CaMKII to synapses has been heavily studied. However, CaMKII actions away from the postsynaptic density (PSD) remain poorly understood, in part because of the difficulty in discerning where CaMKII binds in live cells. We used photoactivated localization microscopy (PALM) in rat hippocampal neurons to track single molecules of CaMKIIα, mapping its spatial and kinetic heterogeneity at high resolution. We found that CaMKIIα exhibits at least three kinetic subpopulations, even within individual spines. Latrunculin application or coexpression of CaMKIIβ carrying its actin-binding domain strongly modulated CaMKII diffusion, indicating that a major subpopulation is regulated by the actin cytoskeleton. CaMKII in spines was typically more slowly mobile than in dendrites, consistent with presence of a higher density of binding partners or obstacles. Importantly, NMDA receptor stimulation that triggered CaMKII activation prompted the immobilization and presumed binding of CaMKII in spines not only at PSDs but also at other points up to several hundred nanometers away, suggesting that activated kinase does not target only the PSD. Consistent with this, single endogenous activated CaMKII molecules detected via STORM immunocytochemistry were concentrated in spines both at the PSD and at points quite distant from the synapse. Together, these results indicate that CaMKII mobility within spines is determined by association with multiple interacting proteins, even outside the PSD, suggesting diverse mechanisms by which CaMKII may regulate synaptic transmission.
authors.affiliation: Department of Physiology, Program in Molecular Medicine, and.
authors.forename: Hsiangmin E
authors.initials: HE
authors.lastname: Lu
authors.affiliation: Department of Physiology, Program in Neuroscience, University of Maryland School of Medicine, Baltimore, Maryland 21201, and.
authors.forename: Harold D
authors.initials: HD
authors.lastname: MacGillavry
authors.affiliation: Department of Physiology, Program in Neuroscience, University of Maryland School of Medicine, Baltimore, Maryland 21201, and Department of Neurology, University of California San Francisco, San Francisco, California 94143.
authors.forename: Nicholas A
authors.initials: NA
authors.lastname: Frost
authors.affiliation: Department of Physiology, Program in Molecular Medicine, and Program in Neuroscience, University of Maryland School of Medicine, Baltimore, Maryland 21201, and tblanpied@som.umaryland.edu.
authors.forename: Thomas A
authors.initials: TA
authors.lastname: Blanpied
mesh_terms: D000818:Animals; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2 / Q000032:analysis / Q000502:physiology*; D002478:Cells, Cultured; D003712:Dendrites / Q000737:chemistry / Q000201:enzymology; D049229:Dendritic Spines / Q000737:chemistry* / Q000201:enzymology*; D019706:Excitatory Postsynaptic Potentials / Q000502:physiology; D005260:Female; D006624:Hippocampus / Q000737:chemistry / Q000166:cytology / Q000201:enzymology; D008297:Male; D018613:Microscopy, Confocal / Q000379:methods; D051381:Rats
chemical_list: D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2
keywords: NMDA receptors; kinase; long-term synaptic plasticity; postsynaptic density; super-resolution imaging; synapse