title: Enigma homolog 1 scaffolds protein kinase D1 to regulate the activity of the cardiac L-type voltage-gated calcium channel.
abstract: OBJECTIVE
  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.
  
  
  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.
  
  
  CONCLUSIONS
  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.
authors.affiliation: Department of Structural Molecular Biology, Institute of Scientific and Industrial Research, Osaka University, Osaka 567-0047, Japan. amaturana@bio.titech.ac.jp
authors.forename: Andrés D
authors.initials: AD
authors.lastname: Maturana
authors.forename: Sébastien
authors.initials: S
authors.lastname: Wälchli
authors.forename: Miki
authors.initials: M
authors.lastname: Iwata
authors.forename: Stephan
authors.initials: S
authors.lastname: Ryser
authors.forename: Johannes
authors.initials: J
authors.lastname: Van Lint
authors.forename: Masahiko
authors.initials: M
authors.lastname: Hoshijima
authors.forename: Werner
authors.initials: W
authors.lastname: Schlegel
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Ikeda
authors.forename: Katsuyuki
authors.initials: K
authors.lastname: Tanizawa
authors.forename: Shun'ichi
authors.initials: S
authors.lastname: Kuroda
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism*; D000256:Adenoviridae / Q000235:genetics; D000316:Adrenergic alpha-Agonists / Q000494:pharmacology; D000818:Animals; D000831:Animals, Newborn; D020746:Calcium Channels, L-Type / Q000235:genetics / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects; D002478:Cells, Cultured; D005822:Genetic Vectors; D006367:HeLa Cells; D006801:Humans; D047468:Immunoprecipitation; D060588:LIM Domain Proteins; D008564:Membrane Potentials; D009154:Mutation; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000201:enzymology*; D018408:Patch-Clamp Techniques; D010656:Phenylephrine / Q000494:pharmacology; D011485:Protein Binding; D011493:Protein Kinase C; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D011817:Rabbits; D051381:Rats; D020798:Two-Hybrid System Techniques
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D000316:Adrenergic alpha-Agonists; C504728:Cacna1c protein, rat; D020746:Calcium Channels, L-Type; D060588:LIM Domain Proteins; C102578:Pdlim5 protein, rat; C099008:Pdlim7 protein, rat; D034741:RNA, Small Interfering; D010656:Phenylephrine; D011494:Protein Kinases; C089212:protein kinase D; D011493:Protein Kinase C