The application is directed towards understanding the process of endomitosis, which is a hallmark of megakaryocytopoiesis and occurs also in other cells such as embryonic cells and vascular smooth muscle cells exposed to hypertension. The investigator has previously found that megakaryocytes do not continuously synthesize DNA but rather undergo a cell cycle consisting of Gap/S phases. She also demonstrated by antisense experiments that the G1 phase cyclin, cyclin D3, is essential for megakaryopoiesis in vitro. She has also cloned from megakaryocytes a CDNA for a protein which has high sequence homology to the family of Never in Mitosis (NIMA) kinases, which were originally identified by their ability to rescue mutant fungal cells arrested at entry to mitosis, and undergo repeated rounds of DNA synthesis. She has called this protein MegNIMA-like. She has data showing that both cyclin D3 and MegNIMA-like are regulated by thrombopoietin. The effect on CD3 is transcriptional. The goal of this proposal is to further the understanding of the regulation of these two genes in megakaryocytopoiesis.
The specific aims are: 1) To isolate the full-length MegNIMA-like CDNA and express and characterize the protein it encodes.2) To characterize CD3 genomic regulatory elements responsible for gene activation in megakaryocytes as well as the TPO-responsive DNA elements. 3) To determine the roles of MegNIMA-like, NIMA kinases and CD3 in megakaryocytopoiesis and thrombocytopoiesis in cultured cells and in vivo. The in vitro studies will be done with both rat megakaryocytes and with cell lines.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL058547-04
Application #
6184400
Study Section
Hematology Subcommittee 2 (HEM)
Project Start
1997-08-01
Project End
2002-07-31
Budget Start
2000-08-01
Budget End
2001-07-31
Support Year
4
Fiscal Year
2000
Total Cost
$351,954
Indirect Cost
Name
Boston University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
604483045
City
Boston
State
MA
Country
United States
Zip Code
02118
Nguyen, Hao G; Ravid, Katya (2010) Polyploidy: mechanisms and cancer promotion in hematopoietic and other cells. Adv Exp Med Biol 676:105-22
Ravid, K (2010) The value of a native milieu: mutated non-muscle myosin IIA does lead to thrombocytopenia. J Thromb Haemost 8:2241-2
Ravid, Katya (2009) Megakaryocytes survive without survivin. Blood 114:4
Ravid, Katya (2009) MAL: not just a leukemia inducer. Blood 114:3977-8
Nguyen, Hao G; Yu, Guangyao; Makitalo, Maria et al. (2005) Conditional overexpression of transgenes in megakaryocytes and platelets in vivo. Blood 106:1559-64
Zhang, Ying; Nagata, Yuka; Yu, Guangyao et al. (2004) Aberrant quantity and localization of Aurora-B/AIM-1 and survivin during megakaryocyte polyploidization and the consequences of Aurora-B/AIM-1-deregulated expression. Blood 103:3717-26
Zhang, Ying; Sun, Shishinn; Wang, Zhengyu et al. (2002) Signaling by the Mpl receptor involves IKK and NF-kappaB. J Cell Biochem 85:523-35
Chinnappan, D; Zhang, Y; Ravid, K (2002) AIM-1 transgenic mice with a curly tail phenotype and its chromosome location. Cytogenet Genome Res 98:231A
Ravid, Katya; Lu, Jun; Zimmet, Jeffrey M et al. (2002) Roads to polyploidy: the megakaryocyte example. J Cell Physiol 190:7-20
Kaluzhny, Yulia; Hechler, Beatrice; Lu, Jun et al. (2002) A selective effect of Mpl ligand on mRNA stabilization during megakaryocyte differentiation. FEBS Lett 527:279-83

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