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Architectural Dynamics of CaMKII-Actin Networks.
- Source :
-
Biophysical journal [Biophys J] 2019 Jan 08; Vol. 116 (1), pp. 104-119. Date of Electronic Publication: 2018 Nov 10. - Publication Year :
- 2019
-
Abstract
- Calcium-calmodulin-dependent kinase II (CaMKII) has an important role in dendritic spine remodeling upon synaptic stimulation. Using fluorescence video microscopy and image analysis, we investigated the architectural dynamics of rhodamine-phalloidin stabilized filamentous actin (F-actin) networks cross-linked by CaMKII. We used automated image analysis to identify F-actin bundles and crossover junctions and developed a dimensionless metric to characterize network architecture. Similar networks were formed by three different CaMKII species with a 10-fold length difference in the linker region between the kinase domain and holoenzyme hub, implying linker length is not a primary determinant of F-actin cross-linking. Electron micrographs showed that at physiological molar ratios, single CaMKII holoenzymes cross-linked multiple F-actin filaments at random, whereas at higher CaMKII/F-actin ratios, filaments bundled. Light microscopy established that the random network architecture resisted macromolecular crowding with polyethylene glycol and blocked ATP-powered compaction by myosin-II miniature filaments. Importantly, the networks disassembled after the addition of calcium-calmodulin and were then spaced within 3 min into compacted foci by myosin motors or more slowly (30 min) aggregated by crowding. Single-molecule total internal reflection fluorescence microscopy showed CaMKII dissociation from surface-immobilized globular actin exhibited a monoexponential dwell-time distribution, whereas CaMKII bound to F-actin networks had a long-lived fraction, trapped at crossover junctions. Release of CaMKII from F-actin, triggered by calcium-calmodulin, was too rapid to measure with flow-cell exchange (<20 s). The residual bound fraction was reduced substantially upon addition of an N-methyl-D-aspartate receptor peptide analog but not ATP. These results provide mechanistic insights to CaMKII-actin interactions at the collective network and single-molecule level. Our findings argue that CaMKII-actin networks in dendritic spines maintain spine size against physical stress. Upon synaptic stimulation, CaMKII is disengaged by calcium-calmodulin, triggering network disassembly, expansion, and subsequent compaction by myosin motors with kinetics compatible with the times recorded for the poststimulus changes in spine volume.<br /> (Copyright © 2018 Biophysical Society. Published by Elsevier Inc. All rights reserved.)
- Subjects :
- Actin Cytoskeleton chemistry
Actin Cytoskeleton metabolism
Actins chemistry
Adenosine Triphosphate metabolism
Animals
COS Cells
Caenorhabditis elegans
Calcium metabolism
Calcium-Calmodulin-Dependent Protein Kinase Type 2 chemistry
Chlorocebus aethiops
Humans
Models, Theoretical
Myosins metabolism
Protein Domains
Rats
Actin Cytoskeleton ultrastructure
Actins metabolism
Calcium-Calmodulin-Dependent Protein Kinase Type 2 metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 1542-0086
- Volume :
- 116
- Issue :
- 1
- Database :
- MEDLINE
- Journal :
- Biophysical journal
- Publication Type :
- Academic Journal
- Accession number :
- 30527447
- Full Text :
- https://doi.org/10.1016/j.bpj.2018.11.006