Inhibition of osteoclast formation is a key mechanism by which estrogen (E2) prevents bone loss. The anti-osteoclastogenic effect of E2 results, in part, from the ability of the sex steroid to block the production of TNF from bone marrow monocytes. However, the mechanism by which E2 regulates TNF production remains unknown. The applicants' preliminary data suggest that in RAW 264.7 cells, a monocytic line, E2 represses cytokine-induced TNF gene expression by decreasing the activity of the Jun N terminal kinase (JNK). The resulting diminished phosphorylation of c-Jun and JunD at their N termini decreases the ability of these nuclear proteins to autostimulate the expression of the c-Jun and JunD genes, thus leading to decreased production of c-Jun and JunD. The resulting decrease in the nuclear levels of c-Jun and JunD leads to decreased binding of c-Jun/c-Fos and JunD/c-Fos heterodimers to the AP-1 consensus sequence (TRE site) in the TNF promoter and, thus, to decreased transactivation of the TNF gene. Therefore, it is hypothesized the E2 down-regulates c-Jun and JunD gene expression via an effect mediated by AP-1 binding sites in the c-Jun and JunD promoters, and that E2 decreases c-Jun and JunD gene expression by down-regulating JNK activity and the resulting phosphorylation of c-Jun and JunD in N terminus. In the first Specific Aim, the applicants wish to determine the mechanism by which E2 down-regulates c-Jun and JunD gene expression. Thus, they will investigate whether E2 affects c-Jun and JunD gene transcription or message stability. If, as expected, transcriptional inhibition is proved to be the key regulatory mechanism, they will analyze the c-Jun and the JunD promoters so as to identify the regions which confer E2 responsiveness.
In Specific Aim 2, they propose to determine the effects of E2 on the phosphorylation of c-Jun and JunD and the mechanism by which E2 modulates the phosphorylation of these proteins. This will be pursued by (a) determining the effects of E2 on the phosphorylation of c-Jun and JunD in both C- and N- termini, (b) investigating the mechanism by which E2 regulates the C terminal phosphorylation of c-Jun and JunD, and (c) investigating the mechanism by which E2 modulates the N terminal phosphorylation of c-Jun and JunD. It is suggested that the relevance of the proposed studies is high because identification of the mechanism by which E2 blocks the monocytic production of TNF may help in clarifying how E2 prevents postmenopausal bone loss.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK055746-05
Application #
6616243
Study Section
Orthopedics and Musculoskeletal Study Section (ORTH)
Program Officer
Malozowski, Saul N
Project Start
2000-08-01
Project End
2005-07-31
Budget Start
2003-08-01
Budget End
2005-07-31
Support Year
5
Fiscal Year
2003
Total Cost
$255,360
Indirect Cost
Name
Emory University
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
066469933
City
Atlanta
State
GA
Country
United States
Zip Code
30322
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