The major objective of this application is to determine the molecular mechanisms by which the RNA binding proteins TIA1 and TIAR (TIA1/R) regulate the production of TNF-alpha, a major mediator of inflammation in systemic rheumatic diseases. TIAR been reported to bind the TNF-alpha mRNA 3' untranslated region (UTR) AU-rich element (ARE), which has been implicated in mediating TNF-alpha mRNA destabilization, deadenylation, and translational repression. Preliminary date revealed that macrophages deficient in TIA1 (TIA1^-/-) or TIAR (TIAR^-/-) secreted 2 to 3- fold higher amounts of TNF protein after LPS stimulation than wild type macrophages. TIA1 and TIAR sequester poly A+ mRNA into sites of translation suppression during cellular stress. The central hypothesis is that TIA1 and TIAR regulate TNF-alpha protein production by affecting one or more of the following processes: TNF-alpha mRNA stability, polyadenylation, and trans- lational efficiency. The initial experiments of the application will define which of these processes is affected by TIA1 and/or TIAR. The molecular mechanisms regulating that process will then be pursued.
The first aim of this application is to compare the TNF-alpha mRNA half-life and poly A tail length in the LPS-stimulated TIA1^-/- and TIAR^-/- macrophages. The mRNA sequence elements and the proteins with which TIA1 and TIAR interact to mediate these processes will be sought.
The second aim of this application will determine whether TIA1 and TIAR affect the translational efficiency of TNF-alpha mRNA and establish the relative proportion of polysome-associated TNF-alpha mRNA in knockout versus wild type cells. Direct translational suppression would be a novel means of regulating TNF-alpha production. The protein-protein and protein-RNA interactions that mediate this process will be characterized. The results of this application will identify new targets for anti-inflammatory therapies.
| Pinz, Ilka; Wax, Stephen D; Anderson, Paul et al. (2008) Low over-expression of TNFalpha in the mouse heart increases contractile performance via TNFR1. J Cell Biochem 105:99-107 |
| Wax, Stephen D; Nakamura, Hideki; Anderson, Paul J (2005) The tumor necrosis factor-alpha AU-rich element inhibits the stable association of the 40S ribosomal subunit with RNA transcripts. Biochem Biophys Res Commun 333:1100-6 |
| Wax, Stephen; Piecyk, Monica; Maritim, Beatrice et al. (2003) Geldanamycin inhibits the production of inflammatory cytokines in activated macrophages by reducing the stability and translation of cytokine transcripts. Arthritis Rheum 48:541-50 |
