Splice variants of a ClC-2 chloride channel with differing functional characteristics.
We identified two ClC-2 clones in a guinea pig intestinal epithelial cDNA library, one of which carries a 30-bp deletion in the NH(2) terminus. PCR using primers encompassing the deletion gave two products that furthermore were amplified with specific primers confirming their authenticity. The corresponding genomic DNA sequence gave a structure of three exons and two introns. An internal donor site occurring within one of the exons accounts for the deletion, consistent with alternative splicing. Expression of the variants gpClC-2 and gpClC-2Delta77-86 in HEK-293 cells generated inwardly rectifying chloride currents with similar activation characteristics. Deactivation, however, occurred with faster kinetics in gpClC-2Delta77-86. Site-directed mutagenesis suggests that a protein kinase C-mediated phosphorylation consensus site lost in gpClC-2Delta77-86 is not responsible for the observed change. The deletion-carrying variant is found in most tissues examined, and it appears more abundant in proximal colon, kidney, and testis. The presence of a splice variant of ClC-2 modified in its NH(2)-terminal domain could have functional consequences in tissues where their relative expression levels are different.
Cid
L P
LP
Instituto de Ciencias Biomédicas, Facultad de Medicina, Universidad de Chile, Santiago-7, Chile. pcid@cecs.cl
Niemeyer
M I
MI
Ramírez
A
A
Sepúlveda
F V
FV
D017398:Alternative Splicing / Q000235:genetics*; D000818:Animals; D001483:Base Sequence; D000090007:CLC-2 Chloride Channels; D002460:Cell Line; D018118:Chloride Channels / Q000235:genetics* / Q000378:metabolism*; D005091:Exons; D015870:Gene Expression; D006168:Guinea Pigs; D006801:Humans; D007413:Intestinal Mucosa / Q000378:metabolism*; D007438:Introns; D015640:Ion Channel Gating / Q000235:genetics; D007668:Kidney / Q000166:cytology / Q000378:metabolism; D008297:Male; D008564:Membrane Potentials; D008969:Molecular Sequence Data; D009928:Organ Specificity; D018408:Patch-Clamp Techniques; D017434:Protein Structure, Tertiary / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D012689:Sequence Homology, Nucleic Acid; D014162:Transfection
D000090007:CLC-2 Chloride Channels; D018118:Chloride Channels