Thrombopoietin (TPO) induces c- myc expression through a PI3K- and MAPK-dependent pathway that is not mediated by Akt, PKC ζ or mTOR in TPO-dependent cell lines and primary megakaryocytes

Abstract

Thrombopoietin (TPO) and its receptor (c-Mpl) are the major regulators of megakaryocyte and platelet production and serve a critical and non-redundant role in hematopoietic stem cell (HSC) biology. TPO signals through the Jak-STAT, Ras-Raf-MAPK, and PI3K pathways, and promotes survival, proliferation, and polyploidization in megakaryocytes. The proto-oncogene c- myc also plays an important role in many of these same processes. In this work we studied the regulated expression of c- myc in megakaryocytic cell lines and primary cells by quantitative real-time RT-PCR. We found that TPO induced expression of c- myc in 1 h in both hematopoietic cell lines (UT-7 and BaF3/Mpl) and mature murine megakaryocytes. The TPO-induced expression of c- myc was blocked by a phosphatidylinositol 3-kinase (PI3K) inhibitor, suggesting that TPO stimulated c- myc expression through a PI3K-dependent pathway. Of interest, our study showed that overexpression of active Akt did not rescue the effect of PI3K blockade on c- myc expression, rather, enhanced it. In addition, inhibitors of protein kinase C (PKC) ζ and the target of rapamycin (mTOR) also failed to affect c- myc mRNA expression, while c- myc mRNA expression was reduced by inhibition of the mitogen activated protein kinase (MAPK) pathway. Therefore, we conclude that TPO stimulates c- myc expression in primary megakaryocytes through a PI3K- and MAPK-dependent pathway that is not mediated by Akt, PKC ζ or mTOR.