ASGSB 1999 Annual Meeting Abstracts

PROSTAGLANDIN E₂-INDUCED UPREGULATION OF C-FOS MRNA IS PRIMARILY MEDIATED BY CAMP IN MC3T3-E1 OSTEOBLASTS. M.Hughes-Fulford, V.Gilbertson and J.Fitzgerald, Laboratory of Cell Growth, VAMC and UCSF, San Francisco, CA.

We have learned from BioRack experiments that signal transduction is changed in microgravity. In spaceflight experiments, synthesis of PGE₂ is up-regulated for both the 0-G and 1-G onboard samples.  Signal transduction in response to PGE₂ is also changed in microgravity.  We have investigated the mechanism behind these changes. On Earth, the mechanism by which the proto-oncogene, c-fos, is up-regulated in response to PGE₂ in the mouse osteoblastic (MC3T3-E₁) cell line was investigated using RT-PCR. c-fos mRNA up-regulation by dmPGE₂ is rapid, starting 10 minutes post-stimulation and transient.  The specific protein kinase A (PKA) inhibitor, H89, inhibited c-fos induction. Moreover, down-regulation of protein kinase C (PKC) activity by chronic TPA treatment had no effect on the induction of c-fos by dmPGE₂.  We conclude that up-regulation of c-fos by dmPGE₂ is primarily dependent on PKA in MC3T3-E₁ osteoblasts. In S49 lymphoma wild type but not S49 cyc^- cells, which are deficient in cAMP signaling, dmPGE2 up-regulates c-fos and increases cell growth compared to unstimulated cells.  Thus, in S49 lymphoma cells, c-fos induction by PGE2 is also dependent on cAMP signaling.   The minimal c-fos promoter region required for dmPGE₂-induced expression was identified by transfecting c-fos promoter deletion constructs coupled to the chloramphenicol acetyltransferase (CAT) reporter gene into Vero cells.  Transfection of a plasmid containing 99bp c-fos proximal promoter was sufficient to direct c-fos/CAT expression following stimulation with dmPGE₂.  Since induction of c-fos is mediated by cAMP these data are consistent with activation of c-fos via the CRE/ATF cis element.  Together, these studies suggest that basic signal transduction in osteoblasts may be changed at the cellular level

Supported by NASA: NAGW 1244 and NAG-2-1086

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