TUMOR-NECROSIS-FACTOR ACTIVATES ANGIOTENSINOGEN GENE-EXPRESSION BY THE REL-A TRANSACTIVATOR

Angiotensinogen encodes the only known precursor of angiotensin II, a critical regulator of the cardiovascular system. Transcriptional contr ol of angiotensinogen in hepatocytes is an important regulator of circ ulating angiotensinogen concentrations. Angiotensinogen transcription is increased by the inflammatory cytokine tumor necrosis factor (TNF)- alpha by a nuclear factor-kappa B-like protein binding to an inducible enhancer called the acute-phase response element. By gel mobility shi ft assays, we observe two specific acute-phase response element-bindin g complexes, C1 and C2. The abundance of C2 is not changed by TNF trea tment. In contrast, C1 is faintly detected in untreated cells, and its abundance increases by fivefold after stimulation. We identify the nu clear factor-kappa B subunits in these complexes using subunit-specifi c antibodies in the gel mobility ''supershift'' assay. The transcripti onally inert nuclear factor-kappa B DNA-binding subunit NF-kappa B1 is present in both control and stimulated hepatocyte nuclei. Its abundan ce changes weakly upon TNF stimulation. In contrast, the potent transa ctivating protein Rel A is not found in unstimulated hepatocyte nuclei and is recruited by TNF-alpha into the C1 DNA-binding complex. Overex pression of Rel A results in acute-phase response element transcriptio n. Cotransfection of a chimeric GAL4-Rel A protein with GAL4 DNA-bindi ng sites is a strategy that allows for selective study of Rel A. The G AL4:Rel A chimera is a TNF-alpha-inducible transactivator. Deletion of the amino-terminal 254 amino acids of Rel A produces a constitutive a ctivator (that is no longer TNF-alpha inducible). The cytokine inducti on of Rel A, then, is mediated through its amino-terminal 254 amino ac ids. We conclude that Rel A:NF-kappa B1 is a crucial cytokine-inducibl e transcription factor complex regulating angiotensinogen gene synthes is in hepatocytes and may be involved in controlling the activity of t he renin-angiotensin system.