CYCLIN D3 IS ESSENTIAL FOR MEGAKARYOCYTOPOIESIS

A normal cell cycle in most eukaryotic cells consists of a tightly reg ulated sequence of phases including DNA synthesis (S) followed by a ga p (G2), mitosis (M), and a gap (G1). In the megakaryocytic lineage, th e cells undergo endomitosis, which involves DNA synthesis in the absen ce of mitosis, thus giving rise to polyploid cells. We aimed at defini ng whether the megakaryocytic cell cycle consists of a continuous S ph ase or of G1/S phases and at determining which cyclins are involved in this process, Studies were performed in primary cultures of mouse bon e marrow cells. DNA synthesis in megakaryocytes was followed by determ ining incorporation of a DNA precursor, bromodeoxyuridine (BrdU), into the cells by in situ staining for BrdU, These experiments showed that no more than 15% of the recognizable megakaryocytes in normal bone ma rrow are in the process of endomitosis, including S phases interrupted by short gaps. Using immunohistochemistry, we showed that mature mega karyocytes express the G1 phase cyclin and cyclin D3, but not the mito tic cyclin, cyclin B1. Under culture conditions that selectively promo te megakaryocytopoiesis, antisense oligonucleotides designed to suppre ss cyclin D3 expression, but not sense oligonucleotides or antisense o ligonucleotides to cyclin B1, dramatically suppress endomitosis and ab rogate megakaryocyte development. Our results indicate that endoredupl ication in megakaryocytes is associated with low levels of or the abse nce of cyclin B1, whereas progression through this process depends on the G1 phase for which cyclin D3 is crucial. (C) 1995 by The American Society of Hematology.