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KMID : 0613820180280121432
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Journal of Life Science
2018 Volume.28 No. 12 p.1432 ~ p.1437
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Repression of CCSP Expression by KLF4
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Kwak In-Seok
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Abstract
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Clara cell secretory protein (CCSP) plays an important role in protecting the lungs from inflammation. This research focuses on identifying the cis-element for binding the repressor of CCSP gene expression. A DNase I footprinting experiment revealed three protected regions between -812 and -768 bp (45 bp) of the mCCSP promoter. One motif (D3: GCCTGGGAA) was 100% conserved across rat, hamster, and human. The addition of excess amounts of the D3 motif exhibited high competition within that 45 bp range in an electrophoretic mobility shift assay. However, when mutated D3 (GAATGTTAA) was used, the competition was significantly reduced. This demonstrates that the D3 motif within that 45 bp region of the mCCSP promoter is an important site for the protein-DNA interaction. Transient transfection assays with -756 Luc resulted in highly decreased expression of CCSP than those with -812 Luc, suggesting that the 45 bp could function as a binding site for the repressor. Co-transfection of KLF4 exhibited significant repression of the -812 Luc but not the -768 Luc which clearly shows that KLF4 might function as a repressor for the CCSP gene and also suggests that the D3 motif is strongly involved in the binding of KLF4. In addition, when anti-KLF4 antibody was added, super-shifted bands were observed. This result demonstrates that KLF4 could function as a repressor by binding to this 45 bp region of the CCSP promoter and that the D3 motif might be involved in the specific binding of KLF4.
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KEYWORD
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Clara cells, CCSP, KLF4, lung, repression
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