Isolation and characterization of kidney-specific CLC-K2 chloride channel gene promoter.
CLC-K1 and CLC-K2 are highly homologous kidney-specific chloride channels, but they are expressed in the different nephron segments. To understand the molecular mechanisms of kidney-specific and nephron-segment-specific expression of CLC-K channel genes, the rat ClC-K2 gene promoter was cloned and compared with that of CLC-K1. In the 1.5-kb pair 5'-flanking region of the CLC-K2 gene, no TATA box was identified around the transcriptional start site, and the proximal region (-32 to -68) was characterized by a GA-rich motif that had a significant sequence similarity to that of the previously isolated CLC-K1 gene promoter. In contrast, the distal portion did not have significant sequence similarity to that of CLC-K1. Reporter gene assay and gel-retardation analysis revealed that the GA-rich motif and the binding of a specific protein(s) to this element were indispensable for the basal promoter activity of the CLC-K2 gene. These results suggest that the GA-rich element may have an important role in the promoter activities of the kidney-specific CLC-K1 and -K2 genes, but that the GA-element alone is not sufficient for the strict regulation of nephron-segment-specific expression of CLC-K1 and CLC-K2 genes.
Rai
T
T
Second Department of Internal Medicine, Tokyo Medical and Dental University, School of Medicine, 1-5-45 Yushima, Bunkyo-ku, Tokyo, 113-8519, Japan.
Uchida
S
S
Sasaki
S
S
Marumo
F
F
D000818:Animals; D027321:Anion Transport Proteins; D001483:Base Sequence; D018118:Chloride Channels / Q000235:genetics*; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D007668:Kidney / Q000378:metabolism*; D008565:Membrane Proteins; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D051381:Rats; D012689:Sequence Homology, Nucleic Acid
D027321:Anion Transport Proteins; C489932:CLCNKA protein, human; D018118:Chloride Channels; C082118:Clcnka protein, rat; C486572:Clcnkb protein, rat; D018076:DNA, Complementary; D008565:Membrane Proteins