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Molecular Characterization of Two Members of the T‐Type Calcium Channel Family
Author(s) -
PEREZREYES EDWARD,
LEE JUNGHA,
CRIBBS LEANNE L.
Publication year - 1999
Publication title -
annals of the new york academy of sciences
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.712
H-Index - 248
eISSN - 1749-6632
pISSN - 0077-8923
DOI - 10.1111/j.1749-6632.1999.tb11283.x
Subject(s) - xenopus , molecular cloning , cloning (programming) , biology , calcium activated potassium channel , genetics , amino acid , membrane potential , gene , microbiology and biotechnology , peptide sequence , chemistry , biophysics , computer science , programming language
ABSTRACT: In this chapter we review our recent studies on the cloning of two novel cDNAs (α1G and α1H), and present electrophysiological evidence that they encode low voltage‐activated, T‐type calcium channels (Ca v T.1 and Ca v T.2, respectively). The nucleotide sequences of these T channels are very different from high voltage‐activated Ca 2+ channels, which explains why they were not cloned earlier using homology‐based strategies. We used a bioinformatic approach, cloning the first fragment in silico . We then used this fragment to screen human heart and rat brain λgt10 libraries, leading to the cloning of two full‐length cDNAs derived from distinct genes ( CACNA1G and CACNA1H ). The deduced amino acid sequences of the T channels (α1G and α1H) are also very different from previously cloned Ca 2+ and Na + channels; however, there are regions of structural similarity. For example, the T channels also contain four repeats, and within each repeat there are six putative membrane‐spanning regions and a pore loop. Expression of these cloned channels in either Xenopus oocytes or HEK‐293 cells leads to the formation of typical T‐type currents. As observed for native T currents, these channels activate at potentials near the resting membrane potential, inactivate rapidly, deactivate slowly, and have a tiny single‐channel conductance. The currents generated by α1G and α1H are nearly identical in terms of their voltage dependence and kinetics. We present preliminary evidence that nickel may serve as a valuable tool in discriminating between these subtypes.