Identification of cis-acting sequences responsible for phorbol ester induction of human serum amyloid A gene expression via a nuclear factor kB-like transcription factor
The authors have analyzed the 5'-flanking region of one of the genes coding for the human acute-phase protein, serum amyloid A (SAA). They found that SAA mRNA could be increased fivefold in transfected cells by treatment with phorbol 12-myristate 13-acetate (PMA). To analyze this observation further, they placed a 265-base-pair 5' SAA fragment upstream of the reporter chloramphenicol acetyltransferase (CAT) gene and transfected this construct into HeLa cells. PMA treatment of these transient transfectants resulted in increased CAT expression. Nuclear proteins from PMA-treated HeLa cells bound to this DNA fragment, and methylation interference analysis showed that the binding was specific to the sequence GGGACTTTCC (between -82 and -91), a sequence previously described by others as the binding site for the nuclear factor NF/kappa/B. In a cotransfection competition experiment, they could abolish PMA-induced CAT activity by using cloned human immunodeficiency virus long-terminal-repeat DNA containing the NF/kappa/B-binding sequence. The same long-terminal-repeat DNA containing mutant NF/kappa/B-binding sequences did not affect CAT expression, which suggested that binding by an NF/kappa/B-like factor is required for increased SAA transcription.
- Research Organization:
- MRC Clinical Research Centre, Harrow, Middlesex HA1 3UJ (GB); AFRC Physiology and Genetics Research Station, Roslin, Midlothian EH25 9PS (GB)
- OSTI ID:
- 5823305
- Journal Information:
- Mol. Cell. Biol.; (United States), Vol. 9:5
- Country of Publication:
- United States
- Language:
- English
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GENE REPRESSORS
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CHLORAMPHENICOL
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ENZYME INDUCTION
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550200* - Biochemistry
550400 - Genetics