Increased expression of alpha 1A Ca2+ channel currents arising from expanded trinucleotide repeats in spinocerebellar ataxia type 6.
The expansion of polyglutamine tracts encoded by CAG trinucleotide repeats is a common mutational mechanism in inherited neurodegenerative diseases. Spinocerebellar ataxia type 6 (SCA6), an autosomal dominant, progressive disease, arises from trinucleotide repeat expansions present in the coding region of CACNA1A (chromosome 19p13). This gene encodes alpha(1A), the principal subunit of P/Q-type Ca(2+) channels, which are abundant in the CNS, particularly in cerebellar Purkinje and granule neurons. We assayed ion channel function by introduction of human alpha(1A) cDNAs in human embryonic kidney 293 cells that stably coexpressed beta(1) and alpha(2)delta subunits. Immunocytochemical analysis showed a rise in intracellular and surface expression of alpha(1A) protein when CAG repeat lengths reached or exceeded the pathogenic range for SCA6. This gain at the protein level was not a consequence of changes in RNA stability, as indicated by Northern blot analysis. The electrophysiological behavior of alpha(1A) subunits containing expanded (EXP) numbers of CAG repeats (23, 27, and 72) was compared against that of wild-type subunits (WT) (4 and 11 repeats) using standard whole-cell patch-clamp recording conditions. The EXP alpha(1A) subunits yielded functional ion channels that supported inward Ca(2+) channel currents, with a sharp increase in P/Q Ca(2+) channel current density relative to WT. Our results showed that Ca(2+) channels from SCA6 patients display near-normal biophysical properties but increased current density attributable to elevated protein expression at the cell surface.
Piedras-Renteria
E S
ES
Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, California 94305, USA.
Watase
K
K
Harata
N
N
Zhuchenko
O
O
Zoghbi
H Y
HY
Lee
C C
CC
Tsien
R W
RW
D015152:Blotting, Northern; D002118:Calcium / Q000378:metabolism; D015220:Calcium Channels / Q000235:genetics* / Q000378:metabolism*; D020865:Calcium Channels, P-Type / Q000235:genetics / Q000378:metabolism; D020907:Calcium Channels, Q-Type / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D002888:Chromosomes, Human, Pair 19 / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D005799:Genes, Dominant; D006801:Humans; D007150:Immunohistochemistry; D017136:Ion Transport / Q000235:genetics; D007668:Kidney / Q000166:cytology / Q000378:metabolism; D018408:Patch-Clamp Techniques; D021122:Protein Subunits; D012333:RNA, Messenger / Q000378:metabolism; D020754:Spinocerebellar Ataxias / Q000209:etiology* / Q000378:metabolism; D014162:Transfection; D019680:Trinucleotide Repeat Expansion / Q000235:genetics*; D020864:Calcium Channels, N-Type
D002118:Calcium; D015220:Calcium Channels; D020865:Calcium Channels, P-Type; D020907:Calcium Channels, Q-Type; D018076:DNA, Complementary; D021122:Protein Subunits; D012333:RNA, Messenger; C105656:CACNA1A protein, human; D020864:Calcium Channels, N-Type