The objective of the proposed study is to understand the malignant transformation of hematopoietic cells by identifying the biological functions of Ikaros proteins. Ikaros is essential for hematopoietic development and acts as a tumor suppressor. The Ikaros gene is alternately spliced to generate multiple zinc finger proteins involved in gene regulation and chromatin remodeling. Our preliminary data show that: 1) Ikaros is phosphorylated at multiple evolutionarily conserved sites by CK2 and other kinases during G1 and S phase of the cell cycle 2) Phosphorylation of Ikaros at specific amino acids regulates its DNA-binding ability and subcellular localization. 3) Ikaros binds to the Bcl-xL gene promoter in vivo and disregulation of Ikaros activity is associated with upregulation of Bcl-xL gene expression.
The specific aims of our proposals are:
Specific Aim #1 : To identify the specific phosphorylation sites responsible for Ikaros function in the regulation of transcription, cellular proliferation and differentiation. We hypothesize that phosphorylation of Ikaros interferes with its function in transcriptional regulation and chromatin remodeling and influences cellular proliferation. We will define phosphorylation sites that are critical for Ikaros function in DNA-binding, subcellular localization, and protein-protein interaction. The role of Ikaros phosphorylation in controlling cell cycle progression will be studied using a murine leukemia cellular system derived from Ikaros deficient mice. These cells will be transduced with wild type or Ikaros phosphomimetic mutants to define sites that are critical for Ikaros'function in cell cycle control. The role of Ikaros' phosphorylation in regulating T cell differentiation and T cell proliferation will be studied in vivo. Murine stem cells from mice with targeted disruption of Ikaros will be infected with retroviral vectors containing wild type Ikaros or phosphomimetic Ikaros mutants and transplanted into sublethally irradiated Ikaros knockout mice. The ability of phosphomimetic Ikaros mutants to restore normal T cell differentiation will be compared to that of wild type Ikaros.
Specific aim #2 : To dissect the mechanism by which Ikaros regulates Bcl-xL expression. Previous studies suggest that decreased Ikaros activity leads to overexpression of the Bcl-xL gene. We hypothesize that Ikaros exerts its tumor suppressor activity by negatively regulating Bcl-xL expression. To test this hypothesis we will determine whether increased Ikaros expression downregulates Bcl-xL transcription in human lymphoma cells and we will map the regions of the Bcl-xL upstream regulatory element (URE) that are critical for Ikaros-modulated control of Bcl-xL expression. These studies will provide the first detailed functional analysis of the signal transduction pathways that control the tumor suppressor function of Ikaros. Our research will provide new and important information on the mechanisms controlling the proliferation of hematopoietic cells and will yield insights into the pathophysiology and treatment of leukemia.

Public Health Relevance

The goal of the proposed project is to identify the function of Ikaros gene. Altered activity of the Ikaros gene is associated with the development of childhood acute lymphoblastic leukemia (ALL), infant leukemia and juvenile chronic lymphocytic leukemia, as well as with several types of adult leukemia and lymphoma. Thus, our results will help to gain insights into the mechanism of development of leukemia. Further elucidation of the mechanism of the malignant transformation process will aid in providing novel and more effective treatment options for patients with leukemia/lymphoma.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL095120-05
Application #
8296355
Study Section
Cancer Molecular Pathobiology Study Section (CAMP)
Program Officer
Thomas, John
Project Start
2009-07-01
Project End
2014-06-30
Budget Start
2012-07-01
Budget End
2013-06-30
Support Year
5
Fiscal Year
2012
Total Cost
$377,941
Indirect Cost
$130,441
Name
Pennsylvania State University
Department
Pediatrics
Type
Schools of Medicine
DUNS #
129348186
City
Hershey
State
PA
Country
United States
Zip Code
17033
Payne, Kimberly J; Dovat, Sinisa (2014) G0S2--a new player in leukemia. Leuk Res 38:147-8
Wang, Haijun; Ouyang, Hongsheng; Lai, Liangxue et al. (2014) Pathogenesis and regulation of cellular proliferation in acute lymphoblastic leukemia - the role of Ikaros. J BUON 19:22-8
Iempridee, Tawin; Reusch, Jessica A; Riching, Andrew et al. (2014) Epstein-Barr virus utilizes Ikaros in regulating its latent-lytic switch in B cells. J Virol 88:4811-27
Wang, Haijun; Song, Chunhua; Gurel, Zafer et al. (2014) Protein phosphatase 1 (PP1) and Casein Kinase II (CK2) regulate Ikaros-mediated repression of TdT in thymocytes and T-cell leukemia. Pediatr Blood Cancer 61:2230-5
Gowda, Chandrika; Dovat, Sinisa (2013) Genetic targets in pediatric acute lymphoblastic leukemia. Adv Exp Med Biol 779:327-40
Li, Zhanjun; Song, Chunhua; Ouyang, Hongsheng et al. (2012) Cell cycle-specific function of Ikaros in human leukemia. Pediatr Blood Cancer 59:69-76
Goldman, Frederick D; Gurel, Zafer; Al-Zubeidi, Duha et al. (2012) Congenital pancytopenia and absence of B lymphocytes in a neonate with a mutation in the Ikaros gene. Pediatr Blood Cancer 58:591-7
Dovat, Sinisa; Song, Chunhua; Payne, Kimberly J et al. (2011) Ikaros, CK2 kinase, and the road to leukemia. Mol Cell Biochem 356:201-7
Yang, Dongqin; Yaguchi, Takahiro; Nagata, Tetsu et al. (2011) AMID mediates adenosine-induced caspase-independent HuH-7 cell apoptosis. Cell Physiol Biochem 27:37-44
Payne, Kimberly J; Dovat, Sinisa (2011) Ikaros and tumor suppression in acute lymphoblastic leukemia. Crit Rev Oncog 16:3-12

Showing the most recent 10 out of 11 publications