The general goal of this project is to better understand the multistage process of cancer development in differentiating epithelia. Because established carcinomas are generally refractory to conventional treatments, knowledge of the genetic defects and cellular markers of carcinoma stages is important for early cancer detection and for the rational design of chemopreventive and therapeutic strategies.
The aims are based on the following hypotheses: 1) p53 protein forms control G1/S and G2/M decision points in epidermal cells, selecting among pathways of terminal differentiation, growth arrest or apoptosis after DNA damage. Normal and malignant cells will be tested for activation of each pathway and dependence on p53 expression by immunoblotting, flow cytometry and indirect immunofluorescence. 2) Kinetically and functionally distinct p53 protein forms respond to DNA damage: a transient response of p53 latent for DNA binding carries out transcriptional repression of growth- associated genes and DNA damage-sensing functions while a sustains response of a p53 form(s) active for sequence-specific DNA binding specializes in transcriptional activation. These functions may be differentially lost during malignant conversion or progression. The principal investigator has found that p53 (regular splice) and p53 as (endogenous alternative splice in mouse cells and (tissues) are induced with different kinetics after treatment of cells with DNA damaging agents. The two proteins also have different DNA binding efficiencies and cell cycle associations, suggesting different functions. These forms and functions will be measured by assays of sequence specific and non-specific DNA binding, and transcriptional modulation of reporter plasmids with or without p53 binding sites, immunoblotting of p53 and p53as and of protein products of downstream target genes. 3) Expression of wt p53 protein in active forms or in active complexes occurs due to cellular factors which form complexes with p53 and regulate its activity. Experiments will include the characterization of two cellular factors, a p80 (which appears to be wild type conformation-specific and is not mdm2) and a p40 (which appears to be p53 conformation independent) which have been coimmunoprecipitated with p53 proteins from cells and which bind to GST/p53 fusion proteins; the characterization will include microsequencing, obtaining or constructing expression vectors in order to verify their association with p53 nd to characterize their role, if any, in p53 activity and contribute to benign tumor formation or malignant conversion. Three transcripts selected by differential display in cells at these transformation stages will be verified by northern blotting, their genes identified and expression vectors used to examine roles in differentiation, cell growth control and cotransformation of cells with a defective p53 gene. Knowledge of different functional p53 protein forms in cells and cooperating factors may improve the resolution, prognosis or treatment plan for human tumors with selective loss of distinct p53 functions.
|Liu, Y; Kulesz-Martin, M F (2006) Sliding into home: facilitated p53 search for targets by the basic DNA binding domain. Cell Death Differ 13:881-4|
|Horn, Elizabeth J; Albor, Amador; Liu, Yuangang et al. (2004) RING protein Trim32 associated with skin carcinogenesis has anti-apoptotic and E3-ubiquitin ligase properties. Carcinogenesis 25:157-67|
|Liu, Yuangang; Lagowski, James P; Vanderbeek, Gretchen E et al. (2004) Facilitated search for specific genomic targets by p53 C-terminal basic DNA binding domain. Cancer Biol Ther 3:1102-8|
|McAllister, Shane C; Hansen, Scott G; Ruhl, Rebecca A et al. (2004) Kaposi sarcoma-associated herpesvirus (KSHV) induces heme oxygenase-1 expression and activity in KSHV-infected endothelial cells. Blood 103:3465-73|
|Knights, Chad D; Liu, Yuangang; Appella, Ettore et al. (2003) Defective p53 post-translational modification required for wild type p53 inactivation in malignant epithelial cells with mdm2 gene amplification. J Biol Chem 278:52890-900|
|Wang, Zhiping; Liu, Yuangang; Mori, Motomi et al. (2002) Gene expression profiling of initiated epidermal cells with benign or malignant tumor fates. Carcinogenesis 23:635-43|
|Huang, Hua; Kaku, Shinsuke; Knights, Chad et al. (2002) Repression of transcription and interference with DNA binding of TATA-binding protein by C-terminal alternatively spliced p53. Exp Cell Res 279:248-59|
|Liu, Y; Asch, H; Kulesz-Martin, M F (2001) Functional quantification of DNA-binding proteins p53 and estrogen receptor in cells and tumor tissues by DNA affinity immunoblotting. Cancer Res 61:5402-6|
|Liu, Y; Kulesz-Martin, M (2001) p53 protein at the hub of cellular DNA damage response pathways through sequence-specific and non-sequence-specific DNA binding. Carcinogenesis 22:851-60|
|Davis, T L; Rabinovitz, I; Futscher, B W et al. (2001) Identification of a novel structural variant of the alpha 6 integrin. J Biol Chem 276:26099-106|
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