Estrogen decreases TNF gene expression by blocking JNK activity and the resulting production of c-Jun and JunD
Sunil Srivastava, … , Stuart Adler, Roberto Pacifici
Sunil Srivastava, … , Stuart Adler, Roberto Pacifici
Published August 15, 1999
Citation Information: J Clin Invest. 1999;104(4):503-513. https://doi.org/10.1172/JCI7094.
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Article

Estrogen decreases TNF gene expression by blocking JNK activity and the resulting production of c-Jun and JunD

Abstract

Central to the bone-sparing effect of estrogen (E2) is its ability to block the monocytic production of the osteoclastogenic cytokine TNF-α (TNF). However, the mechanism by which E2 downregulates TNF production is presently unknown. Transient transfection studies in HeLa cells, an E2 receptor–negative line, suggest that E2 inhibits TNF gene expression through an effect mediated by estrogen receptor β (ERβ). We also report that in RAW 264.7 cells, an E2 receptor–positive murine monocytic line, E2 downregulates cytokine-induced TNF gene expression by decreasing the activity of the Jun NH2-terminal kinase (JNK). The resulting diminished phosphorylation of c-Jun and JunD at their NH2-termini decreases the ability of these nuclear proteins to autostimulate the expression of the c-Jun and JunD genes, thus leading to lower production of c-Jun and JunD. The consequent decrease in the nuclear levels of c-Jun and JunD leads to diminished binding of c-Jun/c-Fos and JunD/c-Fos heterodimers to the AP-1 consensus sequence in the TNF promoter and, thus, to decreased transactivation of the TNF gene.

Authors

Sunil Srivastava, M. Neale Weitzmann, Simone Cenci, F. Patrick Ross, Stuart Adler, Roberto Pacifici

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(a) E2 decreases JNK activity in RAW 264.7 cells. RAW 264.7 cells were t...
(a) E2 decreases JNK activity in RAW 264.7 cells. RAW 264.7 cells were treated with either E2 or vehicle for 24 hours and then stimulated with IL-1 and TNF for 0–15 minutes. Cell extracts were then immunoprecipitated by anti-JNK antibody. Equal amounts of immunoprecipitation material were recovered and incubated with recombinant GST-c-Jun and [γ-32P]ATP. Phosphorylated material was resolved by SDS-PAGE and viewed by autoradiography. Representative data from 1 experiment. (b) E2 has no effects on total JNK levels. Western blot studies were conducted as described in the text using an antibody that recognizes the dephosphorylated and the phosphorylated species of both JNK1 and JNK2. Representative data from 1 of 3 experiments. (c) E2 decreases the level of phosphorylated (active JNK). Western blot analysis was conducted using cell lysates from control- and E2-treated RAW 264.7 cells stimulated with IL-1β and TNF for 5 minutes, and an antibody specific for phospho JNK. Representative data from 1 of 3 experiments.