Analysis of cell cycle modulating proteins in the process of polyploidization of megakaryocytes.

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Mechanism of polypoidization of megakaryocyte remain largely unclarified. To clarify them further, we used a human megakaryoblastic cell line, Meg-J, which showed prominent polyploidization and augmented platelet glycoprotein (GP) Ib expression after incubation with K252a (an indolocarbasole derivative). In the process of K252a-induced polyploidizaton, levels of both cdc2 and cyclin B1 were elevated transiently and subsequently decreased. This suggests that the polyploidization process in Meg-J cells is at least in part associated with a transient elevation and subsequent decrease in the expression of cdc2/cyclin B1 complex. Next, we analyzed the expression of the transcription factors and observed that the expression of NF-E2 p45, but not those of GATA-1, GATA-2, Tal-1/SCL, Evi-1, and MafK, was increased after TPO and K252a stimulation. Gel-shift assay confirmed the enhanced binding activity to the NF-E2 site. The abolishment of NF-E2 p45 with NF-E2 antisense oligomers inhibited TPO plus K252a-induced polyploidization. These findings suggest that NF-E2 p45 is essential for the polyploidization of megakaryocytic cells.