Open AccessTranscriptional Regulation of the CLC-K1 Promoter by myc-Associated Zinc Finger Protein and Kidney-Enriched Krüppel-Like Factor, a Novel Zinc Finger Repressor
Author(s) -
Shinichi Uchida,
Yujiro Tanaka,
Hiroshi Ito,
Fumiko Saitoh-Ohara,
Johji Inazawa,
Kazunari K. Yokoyama,
Sei Sasaki,
Fumiaki Marumo
Publication year - 2000
Publication title -
molecular and cellular biology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.14
H-Index - 327
eISSN - 1067-8824
pISSN - 0270-7306
DOI - 10.1128/mcb.20.19.7319-7331.2000
Subject(s) - zinc finger , biology , microbiology and biotechnology , transcription factor , promoter , zinc finger transcription factor , krüppel , sp1 transcription factor , repressor , reporter gene , zinc finger nuclease , gene , gene expression , biochemistry
The expression of CLC-K1 and CLC-K2, two kidney-specific CLC chloride channels, is transcriptionally regulated on a tissue-specific basis. Previous studies have shown that a GA element near their transcriptional start sites is important for basal and cell-specific activities of the CLC-K1 and CLC-K2 gene promoters. To identify the GA-binding proteins, the human kidney cDNA library was screened by a yeast one-hybrid system. A novel member of the Cys2-His2 zinc finger gene designated KKLF (for “kidney-enriched Krüppel-like factor”) and the previously isolated MAZ (for “myc-associated zinc finger protein”) were cloned. KKLF was found to be abundantly expressed in the liver, kidneys, heart, and skeletal muscle, and immunohistochemistry revealed the nuclear localization of KKLF protein in interstitial cells in heart and skeletal muscle, stellate cells, and fibroblasts in the liver. In the kidneys, KKLF protein was localized in interstitial cells, mesangial cells, and nephron segments, where CLC-K1 and CLC-K2 were not expressed. A gel mobility shift assay revealed sequence-specific binding of recombinant KKLF and MAZ proteins to the CLC-K1 GA element, and the fine-mutation assay clarified that the consensus sequence for the KKLF binding site was GGGGNGGNG. In a transient-transfection experiment, MAZ had a strong activating effect on transcription of the CLC-K1–luciferase reporter gene. On the other hand, KKLF coexpression with MAZ appeared to block the activating effect of MAZ. These results suggest that a novel set of zinc finger proteins may help regulate the strict tissue- and nephron segment-specific expression of the CLC-K1 and CLC-K2 channel genes through their GAcis element.