Nitric oxide-induced nuclear GAPDH activates p300/CBP and mediates apoptosis

Besides its role in glycolysis, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) initiates a cell death cascade. Diverse apoptotic stimuli activate inducible nitric oxide synthase (iNOS) or neuronal NOS (nNOS), with the generated nitric oxide (NO) S-nitrosylating GAPDH, abolishing its catalytic activity and conferring on it the ability to bind to Siah1, an E3-ubiquitin-ligase with a nuclear localization signal (NLS). The GAPDH-Siah1 protein complex, in turn, translocates to the nucleus and mediates cell death; these processes are blocked by procedures that interfere with GAPDH-Siah1 binding. Nuclear events induced by GAPDH to kill cells have been obscure. Here we show that nuclear GAPDH is acetylated at Lys 160 by the acetyltransferase p300/CREB binding protein (CBP) through direct protein interaction, which in turn stimulates the acetylation and catalytic activity of p300/CBP. Consequently, downstream targets of p300/CBP, such as p53 (Refs 10,11,12,13,14,15), are activated and cause cell death. A dominant-negative mutant GAPDH with the substitution of Lys 160 to Arg (GAPDH-K160R) prevents activation of p300/CBP, blocks induction of apoptotic genes and decreases cell death. Our findings reveal a pathway in which NO-induced nuclear GAPDH mediates cell death through p300/CBP.     

The WTX tumor suppressor enhances p53 acetylation by CBP/p300

WTX encodes a tumor suppressor, frequently inactivated in Wilms tumor, with both plasma membrane and nuclear localization. WTX has been implicated in β-catenin turnover, but its effect on nuclear proteins is unknown. We report an interaction between WTX and p53, derived from the unexpected observation of WTX, p53, and E1B 55K colocalization within the characteristic cytoplasmic body of adenovirus-transformed kidney cells. In other cells without adenovirus expression, the C-terminal domain of WTX binds to the DNA-binding domain of p53, enhances its binding to CBP, and increases CBP/p300-mediated acetylation of p53 at Lys 373/382. WTX knockdown accelerates CBP/p300 protein turnover and attenuates this modification of p53. In p53-reconstitution experiments, cell-cycle arrest, apoptosis, and p53 target-gene expression are suppressed by depletion of WTX. Together, these results suggest that WTX modulates p53 function, in part through regulation of its activator CBP/p300.     

Binding of p300/CBP co-activators by polyoma large T antigen.

Small DNA tumor viruses such as simian virus 40 (SV40) and polyomavirus (Py) take advantage of host cell proteins to transcribe and replicate their DNA. Interactions between the viral T antigens and host proteins result in cell transformation and tumor induction. Large T antigen of SV40 interacts with p53, pRb/p107/p130 family members, and the cyclic AMP-responsive element-binding protein (CREB)-binding protein (CBP)/p300. Py large T antigen is known to interact only with pRb and p300 among these proteins. Here we report that Py large T binds to CBP in vivo and in vitro. In co-transfection assays, Py large T inhibits the co-activation functions of CBP/p300 in CREB-mediated transactivation but not in NF-kappa B-mediated transactivation. p53 appears not to be involved in the functions of CREB-mediated transactivation and is not essential for large T:CBP interaction. Mutations introduced into a region of Py large T with homology to adenovirus E1A and SV40 large T prevent binding to the co-activators. These mutant large T antigens fail to inhibit CREB-mediated transactivation. The CBP/p300-binding Py mutants are able to transform established rat embryo fibroblasts but are restricted in their ability to induce tumors in the newborn mouse, indicating that interaction of large T with the co-activators may be essential for virus replication and spread in the intact host.     

p300/CBP/p53 interaction and regulation of the p53 response.

Substantial evidence points to a critical role for the p300/CREB binding protein (CBP) coactivators in p53 responses to DNA damage. p300/CBP and the associated protein P/CAF bind to and acetylate p53 during the DNA damage response, and are needed for full p53 transactivation as well as downstream p53 effects of growth arrest and/or apoptosis. Beyond this simplistic model, p300/CBP appear to be complex integrators of signals that regulate p53, and biochemically, the multipartite p53/p300/CBP interaction is equally complex. Through physical interaction with p53, p300/CBP can both positively and negatively regulate p53 transactivation, as well as p53 protein turnover depending on cellular context and environmental stimuli, such as DNA damage.