The objective is to understand the molecular signals and events which govern cellular proliferation and differentiation. More specifically, those processes that control human monocytic differentiation (using the established HL-60 cell line as a model system) constitute the central theme of this laboratory and thus of this research proposal. In preliminary experiments designed to study the expression of genes specific for the stem and terminally differentiated cell type in the human promyelocytic cell line, we have identified a protein (p30) which is commonly induced by various agents (chemical or cell-derived) that cause the HL-60 cells to traverse a myelocytic differentiation pathway and result in the acquisition of phenotypes characteristic of human macrophages. Initial studies on the properties of p30 (based on its subcellular localization and binding to affinity resins) are consistent with its general participation in cell proliferation/differentiation and a specific role in either the recruitment and/or the template activity of differentiation stage-specific mRNA. The detailed physiochemical and biological characterization of p30 by HL-60 monocytic differentiation-inducing agents (either singly or in combination) is the focal point of this four year research proposal.
Our specific aims are to biochemically isolate p30 by conventional and affinity chromatographic procedures. The availability of partially purified and homogeneous preparation of p30 will permit the extensive biochemical and biological characterization of p30. Biological functions of p30 will be investiagated by examining its ability to inhibit proliferation of HL-60 and various other tumor cell lines. Proliferation will be ascertained by different methods such as (3H)thymidine incorporation. Cell counts and analysis of cell cycle distribution by laser flow cytometry. The possible effects of p30 on differentiation stage-specific mRNA expression will be studied in vitro using either homologous (derived from HL-60 cells) or heterologous (derived from wheat germ or rabbit reticulocytes) cell-free translating systems. The preparation of a p30-monospecific antibody is also proposed as a first step towards a detailed understanding of p30 expression. These studies are expected to provide a better understanding of the molecular features underlying leukemic cell differentiation which will in turn provide more effective approaches for eradication of certain forms of cancer.
| Miyamoto, S; Wu, J M (1990) Effect of staurosporine on the induction of actin/gelsolin in PMA-treated HL-60 cells. Biochem Int 22:427-33 |
| Fraioli, M; Wu, J M (1989) Differential induction of 2',5'-oligoadenylate synthetase by IFN-beta ser and IFN-alpha 2 in serum-supplemented and serum-free HL-60 leukemic cell cultures. J Biol Regul Homeost Agents 3:112-7 |
