Mutational inactivation of the retinoblastoma (Rb) gene predisposes young children to malignant tumors of the eye, and peripheral osteosarcomas as adults. The genesis of a variety of additional human cancers, including small cell lung, bladder and breast carcinomas, is also associated with functional inactivation of the Rb gene. As such, Rb is believed to function in normal cells to limit or constrain cell proliferation and/or facilitate cell differentiation. In recent years, Rb has been shown to physically and functionally interact with a number of cellular transcription factors and to regulate the expression of a handful of growth-related genes. The Rb protein is also subject to a complex series of cell-cycle regulated phosphorylation and dephosphorylation reactions. Is it believed, although not as yet proved, that these cyclic changes in post-translational modification result in alteration of Rb function. Taken together, it is likely that Rb functions as a """"""""molecular switch"""""""", transducing intra- and extracellular signals into differential rates of transcription of key growth-regulating genes. Our long term goals are to precisely define the mechanism(s) by which Rb function is regulated, to identify targets of Rb function, and to relate this knowledge to processes such as cell-cycle progression, cellular senescence, and differentiation. Our experimental approach is to use in vitro assays, such as affinity chromatography and protein-DNA binding, to identify, clone, and characterize cellular proteins that physically interact with the Rb protein or Rb-regulated promoter elements. Thus, in this proposal our Specific Aims are: 1. Identification, cloning, biochemical characterization, and functional analysis of cDNAs encoding proteins that bind specifically to the amino- terminal one-third of the human and murine Rb proteins. 2. Cloning and/or characterization of two nuclear proteins that specifically-bind Rb-regulated elements within the c-fos, c-myc, and TGF- beta1 promoters in vitro and functional analysis of Rb-mediated transcriptional regulation.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
2R01CA053248-04A1
Application #
2095260
Study Section
Molecular Cytology Study Section (CTY)
Project Start
1990-12-12
Project End
1999-04-30
Budget Start
1994-08-02
Budget End
1995-04-30
Support Year
4
Fiscal Year
1994
Total Cost
Indirect Cost
Name
Duke University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
071723621
City
Durham
State
NC
Country
United States
Zip Code
27705
Spengler, Mary L; Kennett, Sarah B; Moorefield, K Scott et al. (2005) Sumoylation of internally initiated Sp3 isoforms regulates transcriptional repression via a Trichostatin A-insensitive mechanism. Cell Signal 17:153-66
Moorefield, K Scott; Fry, Sarah J; Horowitz, Jonathan M (2004) Sp2 DNA binding activity and trans-activation are negatively regulated in mammalian cells. J Biol Chem 279:13911-24
Kennett, Sarah B; Moorefield, K Scott; Horowitz, Jonathan M (2002) Sp3 represses gene expression via the titration of promoter-specific transcription factors. J Biol Chem 277:9780-9
Sterner, J M; Dew-Knight, S; Musahl, C et al. (1998) Negative regulation of DNA replication by the retinoblastoma protein is mediated by its association with MCM7. Mol Cell Biol 18:2748-57
Tao, Y; Kassatly, R F; Cress, W D et al. (1997) Subunit composition determines E2F DNA-binding site specificity. Mol Cell Biol 17:6994-7007
Kennett, S B; Udvadia, A J; Horowitz, J M (1997) Sp3 encodes multiple proteins that differ in their capacity to stimulate or repress transcription. Nucleic Acids Res 25:3110-7
Rogers, K T; Higgins, P D; Milla, M M et al. (1996) DP-2, a heterodimeric partner of E2F: identification and characterization of DP-2 proteins expressed in vivo. Proc Natl Acad Sci U S A 93:7594-9
Sterner, J M; Tao, Y; Kennett, S B et al. (1996) The amino terminus of the retinoblastoma (Rb) protein associates with a cyclin-dependent kinase-like kinase via Rb amino acids required for growth suppression. Cell Growth Differ 7:53-64
Sterner, J M; Murata, Y; Kim, H G et al. (1995) Detection of a novel cell cycle-regulated kinase activity that associates with the amino terminus of the retinoblastoma protein in G2/M phases. J Biol Chem 270:9281-8
Udvadia, A J; Templeton, D J; Horowitz, J M (1995) Functional interactions between the retinoblastoma (Rb) protein and Sp-family members: superactivation by Rb requires amino acids necessary for growth suppression. Proc Natl Acad Sci U S A 92:3953-7

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