Our long term objective is to develop a detailed understanding of erythroid cell differentiation. As such, the specific aims of the project have been to purify, and characterize nuclear factors that interact with the alpha- beta-globin genes. The purification of three identified factors has been published. A fourth factor, found only in nuclear extracts prepared from erythroid cells, has also been purified. Preliminary results show that all factors modulate transcription of globin gene promoters in vitro. Other factors have been identified, but not purified. We propose to extend our results by purifying these additional erythroid cell factors. We also propose to begin an extensive physical characterization of factors already purified. Physical characterization will include peptide mapping, amino acid sequence determination, characterization of factor subunit composition and structure, and determining if factors are phosphorylated or glycosylated. In addition, we intend to prepare polyclonal antisera against each purified factor. Antisera will be used to determine factor distribution, synthesis and turn-over rates, and potential associations with other cellular proteins. Lastly, we will continue to employ our erythroid cell derived in vitro transcription system to assess further the role each purified factor plays in mediating globin gene transcription. Our analysis will help us obtain a deeper understanding of how nuclear proteins regulate cell growth and differentiation. In this regard it is of interest to note that MEL cells are transformed erythroid precursors arrested in differentiation at the CFU-e state. It is not unreasonable to speculate that a detailed understanding of the physical structure and function of each identified factor might lead to a deeper appreciation of how nuclear factors transduce environmental signals that regulate growth and differentiation. These insights might provide novel strategies for selectively altering factor activities with important implications for approaches aimed at altering the uncontrolled growth and lack of differentiation characterization of many tumor cells.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK037513-05
Application #
3236483
Study Section
Hematology Subcommittee 2 (HEM)
Project Start
1986-08-01
Project End
1992-07-31
Budget Start
1990-08-01
Budget End
1991-07-31
Support Year
5
Fiscal Year
1990
Total Cost
Indirect Cost
Name
Sloan-Kettering Institute for Cancer Research
Department
Type
DUNS #
064931884
City
New York
State
NY
Country
United States
Zip Code
10065
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Zhong, F; Swendeman, S L; Popik, W et al. (1994) Evidence that levels of the dimeric cellular transcription factor CP2 play little role in the activation of the HIV-1 long terminal repeat in vivo or following superinfection with herpes simplex virus type 1. J Biol Chem 269:21269-76
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Sheffery, M; Kim, C G; Barnhart, K M (1989) Purification of four erythroid cell proteins that bind the promoters of the murine globin genes. Prog Clin Biol Res 316A:343-57
Barnhart, K M; Kim, C G; Banerji, S S et al. (1988) Identification and characterization of multiple erythroid cell proteins that interact with the promoter of the murine alpha-globin gene. Mol Cell Biol 8:3215-26
Kim, C G; Barnhart, K M; Sheffery, M (1988) Purification of multiple erythroid cell proteins that bind the promoter of the alpha-globin gene. Mol Cell Biol 8:4270-81