CaMKII autophosphorylation can occur between holoenzymes without subunit exchange

Area of Science:

• Molecular and Cellular Neuroscience
• Biochemistry
• Protein Kinase Signaling

Background:

• Calcium/calmodulin-dependent protein kinase II (CaMKII) is crucial for synaptic plasticity and memory.
• Its dodecameric structure and autophosphorylation enable sustained neuronal function.
• The prevailing model for CaMKII activity spread involved subunit exchange.

Purpose of the Study:

• To investigate the mechanism by which CaMKII activity spreads within neurons.
• To challenge the existing subunit exchange hypothesis for CaMKII activation propagation.
• To identify the precise molecular events governing CaMKII phosphorylation spread.

Main Methods:

• Utilized mass photometry, crosslinking mass spectrometry, and single-molecule TIRF microscopy.
• Performed biochemical assays to analyze CaMKII holoenzyme interactions.
• Investigated the role of subunit mobility and inter-holoenzyme phosphorylation.

Main Results:

• Found minimal evidence for subunit exchange upon CaMKII activation.
• Demonstrated that restraining subunits to parent holoenzymes had no effect on activity spread.
• Identified inter-holoenzyme phosphorylation (IHP) as the primary mechanism for spreading CaMKII phosphorylation.
• Observed transient, activity-dependent clustering of CaMKII holoenzymes.

Conclusions:

• Inter-holoenzyme phosphorylation (IHP) is the dominant mechanism for CaMKII activity propagation.
• The formation of CaMKII holoenzyme clusters facilitates rapid, activity-dependent signaling.
• These findings redefine our understanding of CaMKII activation and its role in neuronal plasticity.