title: p38 MAPK regulates IL-1beta induced IL-6 expression through mRNA stability in osteoblasts.
abstract: Osteoblast-derived IL-6 functions in coupled bone turnover by supporting osteoclastogenesis favoring bone resorption instead of bone deposition. Gene regulation of IL-6 is complex occurring both at transcription and post-transcription levels. The focus of this paper is at the level of mRNA stability, which is important in IL-6 gene regulation. Using the MC3T3-E1 as an osteoblastic model, IL-6 secretion was dose dependently decreased by SB203580, a p38 MAPK inhibitor. Steady state IL-6 mRNA was decreased with SB203580 (2 microM) ca. 85% when stimulated by IL-1beta (1-5 ng/ml). These effects require de novo protein synthesis as they were inhibited by cycloheximide. p38 MAPK had minor effects on proximal IL-6 promoter activity in reporter gene assays. A more significant effect on IL-6 mRNA stability was observed in the presence of SB203580. Western blot analysis confirmed that SB203580 inhibited p38 MAP kinase, in response to IL-1beta in a dose dependent manner in MC3T3-E1 cells. Stably transfected MC3T3-E1 reporter cell lines (MC6) containing green fluorescent protein (GFP) with the 3'untranslated region of IL-6 were constructed. Results indicated that IL-1beta, TNFalpha, LPS but not parathyroid hormone (PTH) could increase GFP expression of these reporter cell lines. Endogenous IL-6 and reporter gene eGFP-IL-6 3'UTR mRNA was regulated by p38 in MC6 cells. In addition, transient transfection of IL-6 3'UTR reporter cells with immediate upstream MAP kinase kinase-3 and -6 increased GFP expression compared to mock transfected controls. These results indicate that p38 MAPK regulates IL-1beta-stimulated IL-6 at a post transcriptional mechanism and one of the primary targets of IL-6 gene regulation is the 3'UTR of IL-6.
authors.affiliation: Department of Oral Biology, State University of New York at Buffalo, Buffalo, New York, USA.
authors.forename: Chetan
authors.initials: C
authors.lastname: Patil
authors.forename: Xinsheng
authors.initials: X
authors.lastname: Zhu
authors.forename: Carlos
authors.initials: C
authors.lastname: Rossa
authors.forename: Young Joon
authors.initials: YJ
authors.lastname: Kim
authors.forename: Keith L
authors.initials: KL
authors.lastname: Kirkwood
mesh_terms: D020413:3' Untranslated Regions / Q000235:genetics; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects*; D005822:Genetic Vectors / Q000235:genetics; D007375:Interleukin-1 / Q000494:pharmacology*; D015850:Interleukin-6 / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D010006:Osteoblasts / Q000187:drug effects* / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics; D020871:RNA Stability / Q000187:drug effects*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D048051:p38 Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*
chemical_list: D020413:3' Untranslated Regions; D004791:Enzyme Inhibitors; D007375:Interleukin-1; D015850:Interleukin-6; D012333:RNA, Messenger; D011994:Recombinant Proteins; D048051:p38 Mitogen-Activated Protein Kinases