Biology subjects

Sumino, A.

Publications and source records attributed to Sumino, A..

5 recordsLinked to original sources

bioRxiv2026

CaMKIIα holoenzymes self-organize into worm-like mesoscale clusters

Ca2+/calmodulin-dependent protein kinase II (CaMKII) is highly enriched in dendritic spines at concentrations comparable to those of cytoskeletal proteins and plays a central role in synaptic plasticity. During long-term potentiation (LTP), CaMKII further accumulates in spines. However, the mechanisms governing its higher-order organization remain poorly understood. Here, we use high-speed atomic force microscopy to visualize inter-holoenzyme interaction of CaMKII at mesoscopic scales (5-500 nm). Under freely diffusible conditions, CaMKII holoenzymes do not form stable clusters. In contrast, when spatially confined, they assemble into worm-like chain clusters mediated by kinase-domain interactions. These clusters expand upon activation, concomitant with the dissociation of the regulatory segment. Notably, the CaMKII P212L mutant associated with neurodevelopmental disorders, forms extensive clusters even in the basal state. Together, our findings demonstrate that CaMKII-CaMKII interactions drive mesoscale cluster formation and that precise regulation of cluster size and activation-dependent growth might be critical for synaptic signaling.

neuroscience↗

bioRxiv2025

Structural dynamics of mixed-subunit CaMKIIα/β heterododecamers filmed by high-speed AFM

CaMKII predominantly assembles into a 12-meric ring assembly, primarily consisting of CaMKII and CaMKII{beta} variants in the brain. Previous biochemical studies have reported varying ratios of these CaMKII variants across different brain regions and developmental stages. However, direct evidence for the formation of CaMKII/{beta} heterooligomers within a 12-meric ring assembly has been lacking at the single-molecule level. Here, we employed high-speed atomic force microscopy to visualize the conformational dynamics of forebrain-mimicked CaMKII/{beta} at a 3:1 ratio. Our findings revealed that the CaMKII and CaMKII{beta} subunits are intermixed within the 12-meric ring assembly, with more than 83% probability that CaMKII{beta} subunits adjacent to one another. Furthermore, in the activated state, CaMKII/{beta} heterooligomers form a stable kinase domain complex via interactions between adjacent CaMKII{beta} subunits, resulting in a long-lasting structure with an exposed target binding site. Collectively, our observations provide insights into the structural role of CaMKII{beta} subunits within the CaMKII/{beta} heterododecamer.

neuroscience↗

bioRxiv2023

High-speed AFM reveals fluctuations and dimer splitting of the N-terminal domain of GluA2-γ2

AMPA glutamate receptors (AMPARs) enable rapid excitatory synaptic transmission by localizing to the postsynaptic density of glutamatergic spines. AMPARs possess large extracellular N-terminal domains (NTDs), which participate in AMPAR clustering at synapses. Nevertheless, the dynamics of NTDs and the molecular mechanism governing their synaptic clustering remain elusive. Here, we employed high-speed atomic force microscopy (HS-AFM) to directly visualize the conformational dynamics of NTDs in the GluA2 subunit with TARP {gamma}2 in lipid environments. HS-AFM videos of GluA2-{gamma}2 in the resting and open states revealed fluctuations in NTD dimers. Conversely, in the desensitized state, the two NTD dimers adopted a separated conformation with less fluctuation. Notably, we visualized individual NTD dimers transitioning into monomers. Furthermore, this NTD-dimer splitting resulted in intersubunit exchange between NTD dimers and an increased number of binding sites with the synaptic protein neuronal pentraxin 1. Therefore, our findings illuminate the significance of NTD dynamics in the synaptic clustering of AMPARs.

neuroscience↗

bioRxiv2023

Antithetic effects of agonists and antagonists on the structural fluctuations of TRPV1 channel

Transient receptor potential vanilloid member 1 (TRPV1) is a heat and capsaicin receptor that allows cations to permeate and cause pain. As the molecular basis for temperature sensing, the heat capacity ({Delta}Cp) model (D. E. Clapham, C. Miller, Proc. Natl. Acad. Sci. U. S. A. 108, 19492-19497 (2011).) has been proposed and experimentally supported. Theoretically, heat capacity is proportional to a variance in enthalpy, presumably related to structural fluctuation; however, the fluctuation of TRPV1 has not been directly visualized. In this study, we directly visualized single-molecule structural fluctuations of the TRPV1 channels in a lipid bilayer with the ligands resiniferatoxin (RTX: agonist, 1000 times hotter than capsaicin) and capsazepine (CPZ: antagonist) by high-speed atomic force microscopy (HS-AFM). We observed the structural fluctuations of TRPV1 in an apo state and found that RTX binding enhances structural fluctuations, while CPZ binding suppresses fluctuations. These ligand-dependent differences in structural fluctuation would play a key role in the gating of TRPV1.

physiology↗

bioRxiv2023

Evolutionarily acquired activity-dependent transformation of the CaMKII holoenzyme

Ca2+/calmodulin-dependent protein kinase II (CaMKII) has long been central in synaptic plasticity research. CaMKII is a dodecameric serine/threonine kinase that has been essentially conserved across metazoans for over a million years. While the mechanisms of CaMKII activation are well studied, its "behavior" at the molecular level has remained unobserved. Here, high-speed atomic force microscopy was used to visualize the activity-dependent structural dynamics of rat/hydra/C. elegans CaMKII in various states at nanometer resolution. Among the species, rat CaMKII underwent internal kinase domain aggregation in an activity-dependent manner and showed a higher tolerance to dephosphorylation by phosphatase. Our findings suggest that mammalian CaMKII has evolutionarily acquired a new structural form and a tolerance to phosphatase to maintain robust CaMKII activity for proper neuronal function. One-Sentence SummaryHigh-speed atomic force microscopy reveals the activity-dependent structural dynamics of rat/hydra/C. elegans CaMKII

neuroscience↗