Unscheduled growth signals stimulated by a number of activated oncogenes induce the ARF tumor suppressor protein. ARF can activate a p53 response resulting in cell cycle arrest or apoptotic cell death. The ARF-p53 tumor suppressor pathway is one of the cell's major defense mechanisms against cancer induced by activating oncogenes and is dysfunctional in a large number of human tumor cells. The ARF signaling pathway to p53 remains to be elucidated. A number of the small DNA viruses induce ARF, but also specify proteins that bind to and directly inactivate p53 function. The polyoma virus (Py) activating oncogene, middle T-antigen (PyMT), induces ARF but none of the Py proteins bind to and inactivate p53. We have shown that the polyoma virus small T-antigen (PyST) targets the ARF signaling pathway to p53 so that p53 is not upregulated when ARF is activated. This inhibition of p53 induction requires the Protein Phosphatase 2A (PP2A) binding domain of PyST revealing a previously unrecognized role of PP2A in the ARF-p53 signaling pathway. The PP2A binding domain is required for PyST to substitute for the PP2A B regulatory subunit and bind to the PP2A A scaffolding subunit and C catalytic subunit dimer to form a new PP2A complex. We have observed that PyST activates the MAP Kinase cascade, whereas the similar SV40 small T-antigen (SV40 ST) activates Protein Kinase B (AKT) also in a PP2A dependent manner. Thus PyST through its interaction with PP2A is highlighting a new aspect of the important ARF-p53 tumor-suppressor and MAP Kinase signaling pathways. We plan to use proteomic, biological and biochemical approaches to determine how PyST influences PP2A and PP2A cellular targets to affect these important cellular pathways.
The specific AIMs of this proposal are 1) To determine the effect of the incorporation of PyST into the PP2A holoenzyme by identifying new cellular targets of PP2A complexes containing PyST as well as the PP2A B subunits displaced by PyST 2) To identify PP2A subunits whose inhibition by siRNA mimics the effects of PyST 3) Determine the effect of PyST on the phosphorylation of proteins involved in the ARF-p53 pathway 4) Determine the differential effects of PyST and SV40 ST on the ARF-p53 pathway.
These AIMs will collectively serve to characterize the effect of PP2A on ARF signaling to p53 and potentially suggest new therapeutic approaches to restoring p53 function and exploiting p53 dysfunction in cancer cells. The inappropriate activation of cellular signaling pathways involved in growth control can lead to the generation of a cancer cell. One of the cell's major defense mechanisms against such improper growth signals is to activate the ARF gene leading to stimulation of the p53 tumor suppressor protein that can block cancer formation by causing cell death or cell cycle arrest. Our goal is to determine the molecular mechanisms underlying ARF signaling to p53, which is dysfunctional in a number of human cancers.
|Rodriguez-Viciana, P; Collins, C H; Moule, M G et al. (2006) Chromosomal instability at a mutational hotspot in polyoma middle T-antigen affects its ability to activate the ARF-p53 tumor suppressor pathway. Oncogene 25:1454-62|
|Rodriguez-Viciana, Pablo; Collins, Crista; Fried, Mike (2006) Polyoma and SV40 proteins differentially regulate PP2A to activate distinct cellular signaling pathways involved in growth control. Proc Natl Acad Sci U S A 103:19290-5|
|Hunt, Abigail E; Moule, Madeleine G; Fried, Mike (2006) The rat ARF protein is translated from two closely spaced ATG start codons and can transcriptionally activate p53 in the absence of p53 protein stabilization. Cell Cycle 5:1324-30|
|O'Shea, Clodagh C; Fried, Mike (2005) Modulation of the ARF-p53 pathway by the small DNA tumor viruses. Cell Cycle 4:449-52|
|Moule, Madeleine G; Collins, Crista H; McCormick, Frank et al. (2004) Role for PP2A in ARF signaling to p53. Proc Natl Acad Sci U S A 101:14063-6|