Dataset Information

Stable incorporation versus dynamic exchange of ? subunits in a native Ca2+ channel complex.

ABSTRACT: Voltage-gated Ca(2+) channels are multi-subunit membrane proteins that transduce depolarization into cellular functions such as excitation-contraction coupling in muscle or neurotransmitter release in neurons. The auxiliary ? subunits function in membrane targeting of the channel and modulation of its gating properties. However, whether ? subunits can reversibly interact with, and thus differentially modulate, channels in the membrane is still unresolved. In the present study we applied fluorescence recovery after photobleaching (FRAP) of GFP-tagged ?1 and ? subunits expressed in dysgenic myotubes to study the relative dynamics of these Ca(2+) channel subunits for the first time in a native functional signaling complex. Identical fluorescence recovery rates of both subunits indicate stable interactions, distinct recovery rates indicate dynamic interactions. Whereas the skeletal muscle ?1a isoform formed stable complexes with CaV1.1 and CaV1.2, the non-skeletal muscle ?2a and ?4b isoforms dynamically interacted with both ?1 subunits. Neither replacing the I-II loop of CaV1.1 with that of CaV2.1, nor deletions in the proximal I-II loop, known to change the orientation of ? relative to the ?1 subunit, altered the specific dynamic properties of the ? subunits. In contrast, a single residue substitution in the ? interaction pocket of ?1aM293A increased the FRAP rate threefold. Taken together, these findings indicate that in skeletal muscle triads the homologous ?1a subunit forms a stable complex, whereas the heterologous ?2a and ?4b subunits form dynamic complexes with the Ca(2+) channel. The distinct binding properties are not determined by differences in the I-II loop sequences of the ?1 subunits, but are intrinsic properties of the ? subunit isoforms.

SUBMITTER: Campiglio M

PROVIDER: S-EPMC4148589 | biostudies-literature | 2013 May

REPOSITORIES: biostudies-literature

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Stable incorporation versus dynamic exchange of β subunits in a native Ca2+ channel complex.

Campiglio Marta M Di Biase Valentina V Tuluc Petronel P Flucher Bernhard E BE

Journal of cell science 20130227 Pt 9

Voltage-gated Ca(2+) channels are multi-subunit membrane proteins that transduce depolarization into cellular functions such as excitation-contraction coupling in muscle or neurotransmitter release in neurons. The auxiliary β subunits function in membrane targeting of the channel and modulation of its gating properties. However, whether β subunits can reversibly interact with, and thus differentially modulate, channels in the membrane is still unresolved. In the present study we applied fluoresc ...[more]

PMID: 23447673

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Project description:During the acquisition of memories, influx of Ca2+ into the postsynaptic spine through the pores of activated N-methyl-D-aspartate-type glutamate receptors triggers processes that change the strength of excitatory synapses. The pattern of Ca2+influx during the first few seconds of activity is interpreted within the Ca2+-dependent signaling network such that synaptic strength is eventually either potentiated or depressed. Many of the critical signaling enzymes that control synaptic plasticity,including Ca2+/calmodulin-dependent protein kinase II (CaMKII), are regulated by calmodulin, a small protein that can bindup to 4 Ca2+ ions. As a first step toward clarifying how the Ca2+-signaling network decides between potentiation or depression, we have created a kinetic model of the interactions of Ca2+, calmodulin, and CaMKII that represents our best understanding of the dynamics of these interactions under conditions that resemble those in a postsynaptic spine. We constrained parameters of the model from data in the literature, or from our own measurements, and then predicted time courses of activation and autophosphorylation of CaMKII under a variety of conditions. Simulations showed that species of calmodulin with fewer than four bound Ca2+ play a significant role in activation of CaMKII in the physiological regime,supporting the notion that processing of Ca2+ signals in a spine involves competition among target enzymes for binding to unsaturated species of CaM in an environment in which the concentration of Ca2+ is fluctuating rapidly. Indeed, we showed that dependence of activation on the frequency of Ca2+ transients arises from the kinetics of interaction of fluctuating Ca2+with calmodulin/CaMKII complexes. We used parameter sensitivity analysis to identify which parameters will be most beneficial to measure more carefully to improve the accuracy of predictions. This model provides a quantitative base from which to build more complex dynamic models of postsynaptic signal transduction during learning.

Dataset Information

Stable incorporation versus dynamic exchange of ? subunits in a native Ca2+ channel complex.

Publications

Stable incorporation versus dynamic exchange of β subunits in a native Ca2+ channel complex.

Similar Datasets

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