Association of p19ARF with Mdm2 inhibits ubiquitin ligase activity of Mdm2 for tumor suppressor p53

Abstract

We have demonstrated previously that the oncoprotein Mdm2 has a ubiquitin ligase activity for the tumor suppressor p53 protein. In the present study, we characterize this ubiquitin ligase activity of Mdm2. We first demonstrate the ubiquitination of several p53 point mutants and deletion mutants by Mdm2. The point mutants, which cannot bind to Mdm2, are not ubiquitinated by Mdm2. The ubiquitination of the C-terminal deletion mutants, which contain so-called Mdm2-binding sites, is markedly decreased, compared with that of wild-type p53. The binding of Mdm2 to p53 is essential for ubiquitination, but p53's tertiary structure and/or C-terminal region may also be important for this reaction. DNA-dependent protein kinase is known to phosphorylate p53 on Mdm2-binding sites, where DNA damage induces phosphorylation, and p53 phosphorylated by this kinase is not a good substrate for Mdm2. This suggests that DNA damage-induced phosphorylation stabilizes p53 by inhibiting its ubiquitination by Mdm2. We further investigated whether the tumor suppressor p19(ARF) affects the ubiquitin ligase activity of Mdm2 for p53. The activity of p19(ARF)-bound Mdm2 was found to be lower than that of free Mdm2, suggesting that p19(ARF) promotes the stabilization of p53 by inactivating Mdm2.

Protein serine/threonine phosphatase-1 dephosphorylates p53 at Ser-15 and Ser-37 to modulate its transcriptional and apoptotic activities

“…One of the targets is the gene encoding the p19ARF, which is a key component of the p19ARF-MDM2-p53 network (Zhang et al, 1998;Zhu et al, 1999;Trimarchi and Lees, 2002). Activation of p19ARF inhibits MDM2, a p53 ubiquitin ligase, enhancing p53 stabilization (Momand et al, 1992;Kubbutat et al, 1997;Pomerantz et al, 1998;Stott et al, 1998;Zhang et al, 1998;Haupt et al, 1999;Honda and Yasuda, 1999;Weber et al, 1999;Llanos et al, 2001;Zindy et al, 2003). This explains why inhibition of PP-1 activity could lead to observed p53 upregulation in rabbit and rat lens epithelial cells (Li et al, 1998(Li et al, , 2001a.…”

ARF antagonizes the ability of Miz-1 to inhibit p53-mediated transactivation

“…Most, if not all, of these cellular responses are mediated through p53. Indeed, ARF stabilizes and activates p53 by directly interacting with its major negative regulator Mdm2, thereby inhibiting the ubiquitin ligase activity of Mdm2 and abolishing the inhibitory effect of Mdm2 on p53 (Kamijo et al, 1998;Pomerantz et al, 1998;Honda and Yasuda, 1999). Intriguingly, the mechanism by which ARF modulates Mdm2-p53 pathway is now appearing to be more complex than earlier conceptualized.…”

Mouse double minute antagonist Nutlin-3a enhances chemotherapy-induced apoptosis in cancer cells with mutant p53 by activating E2F1

“…This results in p53 stabilization, p21 induction, cell cycle arrest in G1 and G2 phases, apoptosis and growth inhibition of human tumor xenografts in nude mice. Although MDM2 has been extensively characterized as a regulator of p53, there is considerable evidence that MDM2 functionally interacts with a number of other proteins, including the tumor suppressor p14ARF (Honda and Yasuda, 1999), the retinoblastoma pRb (Xiao et al, 1995), the ribosomal protein L5 (Dai and Lu, 2004), E2F/DP1 (Martin et al, 1995) and p73a transcription factor (Zeng et al, 1999). These interactions suggest that MDM2 has p53-independent functions (Ganguli and Wasylyk, 2003).…”