C-terminal ubiquitination of p53 contributes to nuclear export.

The growth inhibitory functions of p53 are controlled in unstressed cells by rapid degradation of the p53 protein. One of the principal regulators of p53 stability is MDM2, a RING finger protein that functions as an E3 ligase to ubiquitinate p53. MDM2 promotes p53 nuclear export, and in this study, we show that ubiquitination of the C terminus of p53 by MDM2 contributes to the efficient export of p53 from the nucleus to the cytoplasm. In contrast, MDM2 did not promote nuclear export of the p53-related protein, p73. p53 nuclear export was enhanced by overexpression of the export receptor CRM1, although no significant relocalization of MDM2 was seen in response to CRM1. However, nuclear export driven by CRM1 overexpression did not result in the degradation of p53, and nuclear export was not essential for p53 degradation. These results indicate that MDM2 mediated ubiquitination of p53 contributes to both nuclear export and degradation of p53 but that these activities are not absolutely dependent on each other.

Contribution of two independent MDM2-binding domains in p14(ARF) to p53 stabilization.

The MDM2 protein targets the p53 tumor suppressor for ubiquitin-dependent degradation [1], and can function both as an E3 ubiquitin ligase [2] and as a regulator of the subcellular localization of p53 [3]. Oncogene activation stabilizes p53 through expression of the ARF protein (p14(ARF) in humans, p19(ARF) in the mouse) [4], and loss of ARF allows tumor development without loss of wild-type p53 [5] [6]. ARF binds directly to MDM2, and prevents MDM2 from targeting p53 for degradation [6] [7] [8] [9] by inhibiting the E3 ligase activity of MDM2 [2] and preventing nuclear export of MDM2 and p53 [10] [11]. Interaction between ARF and MDM2 results in the localization of both proteins to the nucleolus [12] [13] [14] through nucleolar localization signals (NoLS) in ARF and MDM2 [11] [12] [13] [14]. Here, we report a new NoLS within the highly conserved amino-terminal 22 amino acids of p14(ARF), a region that we found could interact with MDM2, relocalize MDM2 to the nucleolus and inhibit the ability of MDM2 to degrade p53. In contrast, the carboxy-terminal fragment of p14(ARF), which contains the previously described NoLS [11], did not drive nucleolar localization of MDM2, although this region could bind MDM2 and weakly inhibit its ability to degrade p53. Our results support the importance of nucleolar sequestration for the efficient inactivation of MDM2. The inhibition of MDM2 by a small peptide from the amino terminus of p14(ARF) might be exploited to restore p53 function in tumors.

Regulation and function of the p53-related proteins: same family, different rules.

The tumour-suppressor protein p53 has recently been shown to belong to a family that includes two structurally related proteins, p63 and p73. Although all three proteins share similar transcriptional functions and the ability to induce apoptosis, each of them appears to play a distinct role in development and tumour suppression. In order for cell division to occur, the antiproliferative activities of these proteins must be tightly controlled, and exciting advances have been made in our understanding of the pathways involved in regulating p53 activity.

Regulation and activation of p53 and its family members.

Regulation of the p53 tumor suppressor protein occurs to a large extent through control of protein stability, and the MDM2 protein has been shown to play a key role in targeting p53 for degradation. Stress signals that activate the p53 response lead to stabilization of p53 through inhibition of MDM2 mediated degradation, and it is becoming evident that a number of mechanisms exist to abrogate this activity of MDM2. Other members of the p53 protein family may also be regulated through protein stability, although MDM2 is not responsible for the degradation of p73. Nevertheless, interactions of p63 and p73 with MDM2 or p53 have been described, suggesting that each of the p53-related proteins can play some role in regulating the activity of the others