Deregulated Expression of c-myc in Megakaryocytes of Transgenic Mice Increases Megakaryopoiesis and Decreases Polyploidization

Alexander Thompson,Ying Zhang,Dimitry Kamen,Carl W Jackson,Robert D Cardiff,Katya Ravid

doi:10.1074/jbc.271.38.22976

Abstract

Platelets, essential for vascular integrity and hemostasis, fragment from polyploid megakaryocytes, characterized by their endomitotic cell cycle. We studied the influence of overexpression of c-myc oncogene on megakaryopoiesis and endomitosis in vivo, using transgenic mice carrying c-myc fused to the estrogen receptor under the control of the platelet factor 4 (PF4) megakaryocyte-specific promoter. The rationale behind this strategy was to obtain controlled overexpression of an active c-Myc, depending on the estrogen level in the mouse circulation. Analysis of these transgenic mice revealed that the bone marrow of female transgenic mice or of estrogen-injected male transgenic mice, but not of age-matched transgenic males nor nontransgenic females, contained frequent immature myeloid cells and an increased number of megakaryocytes. Deregulated expression of c-Myc shifted the normal ploidy profile of megakaryocytes due to a significant increase in proliferating megakaryocytes and a decrease in the fraction of ploidizing cells. These transgenic mice represent a novel in vivo model for a Myc-induced myeloproliferative disorder which can be controlled.

Highlights

Platelet precursors, the megakaryocytes, undergo an endomitotic cell cycle whereby they replicate DNA but do not undergo cytokinesis
We studied the influence of overexpression of c-myc oncogene on megakaryopoiesis and endomitosis in vivo, using transgenic mice carrying c-myc fused to the estrogen receptor under the control of the platelet factor 4 (PF4) megakaryocyte-specific promoter
Plasmids—The plasmid PF4MERGH was constructed by using gene fragments from the following PUC based plasmids: pPF4GH (Ravid et al, 1991b) which contains the rat 1.1-kb PF4 promoter linked to human growth hormone gene (HGH); pMV-7MER (Eilers et al, 1989) which contains exons 2 and 3 of human myc fused to the estrogen receptor (MER). pPF4GH has a unique NdeI site at the 5Ј end of the PF4 promoter, a unique EcoRI site at the 3Ј end of the HGH gene and a unique BanII site 20 bp downstream of the transcriptional start

Summary

Introduction

The megakaryocytes, undergo an endomitotic cell cycle whereby they replicate DNA but do not undergo cytokinesis. Expression of Myc is induced following mitogenic stimulation, is shut off during entry into quiescence, and its expression is deregulated in various neoplasias (Cole, 1986; Marcu et al, 1992) These effects of Myc were attributed to its ability to bind growth suppressors such as the retinoblastoma gene product (Rustgi et al, 1991), to act as a sequence-specific transcriptional regulator (Amin et al, 1993), and to induce DNA replication (Classon et al, 1987). Since megakaryocyte precursor proliferation occurs prior to megakaryocyte polyploidization, and the cell undergoing polyploidization is more differentiated than the cell undergoing cell division, we hypothesized that c-Myc overexpression in early megakaryocytes may increase precursor proliferation and, decrease the fraction of polyploid cells We have tested this hypothesis by using a transgenic model with constitutive overexpression of c-Myc in the megakaryocytic lineage in vivo. Myc Oncogene in Megakaryocyte Development that c-Myc overexpression induces megakaryocyte precursor proliferation at the expense of polyploidization

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Conclusion