Abstract
L-type Ca(2+) channels in native tissues have been found to contain
a pore-forming alpha(1) subunit that is often truncated at the C
terminus. However, the C terminus contains many important domains
that regulate channel function. To test the hypothesis that C-terminal
fragments may associate with and regulate C-terminal-truncated alpha(1C)
(Ca(V)1.2) subunits, we performed electrophysiological and biochemical
experiments. In tsA201 cells expressing either wild type or C-terminal-truncated
alpha(1C) subunits in combination with a beta(2a) subunit, truncation
of the alpha(1C) subunit by as little as 147 amino acids led to a
10-15-fold increase in currents compared with those obtained from
control, full-length alpha(1C) subunits. Dialysis of cells expressing
the truncated alpha(1C) subunits with C-terminal fragments applied
through the patch pipette reconstituted the inhibition of the channels
seen with full-length alpha(1C) subunits. In addition, C-terminal
deletion mutants containing a tethered C terminus also exhibited
the C-terminal-induced inhibition. Immunoprecipitation assays demonstrated
the association of the C-terminal fragments with truncated alpha(1C)
subunits. In addition, glutathione S-transferase pull-down assays
demonstrated that the C-terminal inhibitory fragment could associate
with at least two domains within the C terminus. The results support
the hypothesis the C- terminal fragments of the alpha(1C) subunit
can associate with C-terminal-truncated alpha(1C) subunits and inhibit
the currents through L-type Ca(2+) channels.
Users
Please
log in to take part in the discussion (add own reviews or comments).