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Calmodulin mediates Ca2+-dependent modulation of M-type K+ channels
Nikita Gamper1, Mark S Shapiro
1Department of Physiology, MS 7756, University of Texas Health Science Center at San Antonio, 7703 Floyd Curl Drive, San Antonio, TX 78229, USA.
The Journal of General Physiology
|June 18, 2003
Summary
Calmodulin (CaM) acts as a calcium sensor for M-type currents, significantly modulating their activity. This study demonstrates CaM
Area of Science:
- Neuroscience
- Molecular Biology
- Ion Channel Physiology
Background:
- M-type currents, mediated by KCNQ2/3 channels, play crucial roles in neuronal excitability.
- The precise mechanisms regulating M-type current sensitivity to intracellular calcium ([Ca2+]i) remain incompletely understood.
- Calmodulin (CaM) is a ubiquitous calcium-binding protein implicated in various cellular signaling pathways.
Purpose of the Study:
- To quantify the modulation of KCNQ2/3 currents by intracellular calcium ([Ca2+]i).
- To determine if calmodulin (CaM) mediates the calcium sensitivity of KCNQ2/3 channels.
- To investigate the role of CaM in the regulation of endogenous M-currents in superior cervical ganglion (SCG) neurons.
Main Methods:
- Simultaneous whole-cell recording and Ca2+ imaging in CHO cells expressing KCNQ2/3 channels with or without wild-type (wt) or dominant-negative (DN) CaM.
- Manipulation of intracellular calcium ([Ca2+]i) levels using ionomycin and buffered solutions.
- Coimmunoprecipitation and gel-shift assays to assess CaM-KCNQ channel interactions.
- Exogenous expression of wt or DN CaM in SCG neurons via pseudovirions or gene gun to study bradykinin modulation of M-current.
Main Results:
- Coexpression of wt CaM rendered KCNQ2/3 currents highly sensitive to [Ca2+]i (IC50 ~70 nM), while DN CaM rendered them insensitive.
- Endogenous M-currents in SCG neurons exhibited [Ca2+]i sensitivity patterns similar to KCNQ2/3 channels coexpressed with wt CaM.
- CaM directly binds to KCNQ2-5 subunits in a Ca2+-dependent manner, likely via an IQ-like motif.
- Bradykinin-induced inhibition of M-current in SCG neurons was significantly reduced by wt or DN CaM expression, suggesting CaM's involvement.
Conclusions:
- M-type currents are highly sensitive to intracellular calcium ([Ca2+]i).
- Calmodulin (CaM) acts as the primary intracellular calcium sensor for M-type currents.
- CaM plays a critical role in mediating the modulation of M-currents by external stimuli like bradykinin.