Enigma homolog 1 scaffolds protein kinase D1 to regulate the activity of the cardiac L-type voltage-gated calcium channel
Autor: | Masahiko Hoshijima, Sébastien Wälchli, Miki Iwata, Yasuhiro Ikeda, Andrés D. Maturana, Stephan Ryser, Shun'ichi Kuroda, Werner Schlegel, Johannes Van Lint, Katsuyuki Tanizawa |
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Rok vydání: | 2008 |
Předmět: |
Patch-Clamp Techniques
Calcium Channels L-Type Physiology Genetic Vectors chemistry.chemical_element Calcium Biology urologic and male genital diseases Article Adenoviridae Membrane Potentials Phenylephrine Two-Hybrid System Techniques Physiology (medical) Animals Humans Immunoprecipitation Myocytes Cardiac Calcium Signaling Patch clamp RNA Small Interfering Protein kinase A Cells Cultured Protein Kinase C Protein kinase C Adaptor Proteins Signal Transducing Calcium signaling Voltage-dependent calcium channel Calcium channel LIM Domain Proteins Molecular biology female genital diseases and pregnancy complications Protein Structure Tertiary Rats Cell biology Animals Newborn chemistry Mutation RNA Interference Rabbits Protein kinase D1 Cardiology and Cardiovascular Medicine Adrenergic alpha-Agonists Protein Kinases HeLa Cells Protein Binding |
Zdroj: | Cardiovascular Research. 78:458-465 |
ISSN: | 1755-3245 0008-6363 |
Popis: | Aims In cardiomyocytes, protein kinase D1 (PKD1) plays a central role in the response to stress signals. From a yeast two-hybrid assay, we have identified Enigma Homolog 1 (ENH1) as a new binding partner of PKD1. Since in neurons, ENH1, associated with protein kinase Cepsilon, was shown to modulate the activity of N-type calcium channels, and the pore-forming subunit of the cardiac L-type voltage-gated calcium channel, alpha1C, possesses a potential phosphorylation site for PKD1, we studied here a possible role of ENH1 and PKD1 in the regulation of the cardiac L-type voltage-gated calcium channel. Methods and results PKD1-interacting proteins were searched by yeast two-hybrid screening. In vivo protein interactions in cardiomyocytes isolated from heart ventricles of newborn rats were tested by co-immunoprecipitation. Small interfering RNA and a dominant negative mutant of PKD1 were delivered into cardiomyocytes by use of an adenovirus. Calcium currents were measured by the patch-clamp technique. Both ENH1 and PKD1 interact with alpha1C in cardiomyocytes. This interaction is increased upon stimulation. Silencing of ENH1 prevented the binding of PKD1 to alpha1C. Moreover, a dominant negative mutant of PKD1 or the silencing of ENH1 inhibited the alpha-adrenergic-induced increase of L-type calcium currents. Conclusion We found a new binding partner, ENH1, and a new target, alpha1C, for PKD1 in neonatal rat cardiomyocytes. We propose a model where ENH1 scaffolds PKD1 to alpha1C in order to form a signalling complex that regulates the activity of cardiac L-type voltage-gated Ca(2+) channels. |
Databáze: | OpenAIRE |
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