Cloning and expression of the Ca2+ channel α(1C) and β(2a) subunits from guinea pig heart

Shan Ding, Sachiko Kuroki, Asako Kameyama, Akihiko Yoshimura, Masaki Kameyama

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11 Citations (Scopus)

Abstract

Complimentary DNA clones encoding the α(1C) and β(2a) subunits of guinea-pig cardiac L-type Ca2+ channels were isolated using the PCR method. The open reading frame encoded 2169 amino acids for the α(1C) and 597 amino acids for the β(2a) subunit. The proteins showed 94.2 and 94.8%, respectively, identity to the respective subunit of the rabbit protein. The message size of the guinea pig α(1C) and β(2a) subunits was 8.0 and 3.5/4.0 kb, respectively. RT-PCR analysis revealed that the α(1C) subunit is expressed exclusively in the heart, while the β(2a) subunit is expressed in the heart, cerebellum, whole brain, and stomach. The α(1C) and β(2a) subunits are transiently expressed in BHK (baby hamster kidney) cells, and the channel currents were studied using the whole-cell patch clamp technique in medium containing 30 mM Ba2+. In cells expressing α(1C) alone, the Ba2+ current was activated at -30 mV and more positive potentials and peaked at about 10 mV. The co-expression of β(2a) with α(1C) did not affect the voltage-dependence of the current, but increased the peak current and accelerated current decay. In cells transfected with guinea pig α(1C) and rabbit β1 + α2/δ, a Ba2+ current comparable to those in native myocytes was observed. The Ba2+ current can be blocked completely by nifedipine and is enhanced S-fold by Bay K 8644. On the other hand, neither forskolin nor okadaic acid affects the Ba2+ current, suggesting that cAMP-mediated modulation is not easily reproduced in transfected cells, unlike that seen in native cardiac myocytes.

Original languageEnglish
Pages (from-to)750-759
Number of pages10
JournalJournal of biochemistry
Volume125
Issue number4
DOIs
Publication statusPublished - 1999 Jan 1
Externally publishedYes

Keywords

  • Ca channel
  • Cloning
  • Guinea pig heart
  • α(1C) sununit
  • β subunit

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology

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