p53 Orchestrates multiple pathways for tumor suppression.

p53, described initially in 1979, is one of the first tumor suppressor genes identified. p53 also is the most commonly mutated gene in human cancers, with over 50% of spontaneous tumors showing p53 mutations. In normal cells, p53 is quiescent, but following activation by cellular stresses, p53 binds to specific DNA sequences, where it induces expression of target genes that mediate tumor suppression. Multiple pathways are regulated by p53, including cell cycle arrest, DNA repair, inhibition of angiogenesis and metastasis, and apoptosis. Understanding the signaling networks that operate in p53-dependent activities may lead to the identification of novel genes and proteins amenable to therapies to overcome the p53 inactivation seen in human tumors. Mdm2, an E3 ubiquitin ligase, is one such regulator of p53 activity that may be a potential therapeutic target.

**********

Tumorigenesis is a process by which normal cells, through multiple genetic alterations, acquire many unique characteristics that disrupt the orderly regulation of cell growth and differentiation. For example, cells become competent to proliferate in the absence of growth factors or in the presence of anti-growth factors. Also cells acquire mechanisms to evade apoptosis, to induce angiogenesis, and to metastasize.

Naturally, the body has the means to combat tumorigenesis, and it does so through tumor-suppressor proteins. Examples include p53, RB (retinoblastoma), WT1 (Wilms' tumor), INK4A (melanoma), NF1 (neurofibromatosis), APC (adenomatous polyposis), and BRCA1 and 2 (breast and ovarian cancer).

The p53 tumor suppressor, identified in 1979, is a prototypical tumor suppressor. Through research over the years, p53 has earned its nickname as the "guardian of the genome." This paper reviews the varied mechanisms regulated by p53 that act to suppress tumor formation and growth.

p53 Is a Prototypical Tumor Suppressor

In general, tumor suppressor proteins function to inhibit the formation of tumors by various mechanisms. p53 mediates tumor suppression as a sequence-specific transcription factor. Upon genotoxic stress, p53 is activated to transactivate genes that serve to prevent tumor formation. For example, p53 upregulates genes that mediate cell cycle arrest, DNA repair, apoptosis, inhibition of angiogenesis and metastasis, and regulation of p53 itself.

While many genes can inhibit cell growth, specific criteria (termed Knudson's criteria) must be fulfilled for a gene to be classified as a tumor suppressor. p53 fulfills these criteria.

* First, p53 is recessive. Thus, one copy of p53 in a cell is sufficient to confer tumor suppression. However, the ability of p53 to function as a tumor suppressor is lost when the remaining wild-type allele is inactivated, resulting in loss of heterozygosity (LOH).

* Second, germline mutation of p53 results in the familial cancer syndrome Li-Fraumeni syndrome. Persons with this syndrome develop multiple soft-tissue sarcomas as early as 1 year of age, as well as breast and other tumor types.

* Third, p53 is frequently mutated or inactivated in spontaneous tumors. In fact, p53 is the most commonly mutated gene in human tumors, with over half of all spontaneous tumors containing mutations in the p53 gene.

The phenotype of the p53 knockout mouse also validates p53 as a tumor suppressor. Although most of these knockout mice are developmentally normal, all mice lacking p53 develop multiple spontaneous tumors at an early age. Interestingly, in mouse models of Li-Fraumeni syndrome, where mice are engineered to have one wild-type and one mutant p53 allele, the mice develop a spectrum of tumors similar to those of Li-Fraumeni patients. Importantly, studies using cells isolated from these mice confirm that mutant p53 does indeed possess gain-of-function oncogenic properties.

[GRAPHIC OMITTED]

Different p53 Domains Contribute to the Protein's Variable Functions

The p53 protein contains many functional domains: two transcriptional activation domains (AD1 and AD2), a sequence-specific DNA-binding domain, a nuclear localization signal, a tetramerization domain, and a C-terminal basic domain.

[ILLUSTRATION OMITTED]

Activation domains AD1 and 2 are important for transactivation; these domains contain residues that contact the basal transcriptional machinery. Interestingly, AD1 and AD2 play key roles in the differential activation of p53 target genes. For example, AD1 is important for the induction of the cyclin-dependent kinase inhibitor p21, whereas AD2 is required for induction of the proapoptotic p53 target genes p53AIP1 and IGFBP3.

The DNA-binding domain recognizes the canonical p53 responsive element, consisting of two decamers ([RRRC A/T A/T GYYY) separated by a spacer of 0 to 13 basepairs (where R represents purine, G guanine, Y pyrimidine, and C cytidine).

[ILLUSTRATION OMITTED]

The C-terminal basic domain has been subjected to extensive analyses, which suggest that it is an important regulatory domain. This domain can both positively and negatively regulate transactivation of p53 target genes.

Although p53 exists predominantly as a single isoform, N- and C-terminally truncated forms have been identified for both human and mouse p53. N-terminal truncation of p53 yields [DELTA]Np53 and [DELTA]40p53 and occurs through use of an internal translational start site at codon 40 and 41, respectively. C-terminal truncation yields I9+p53 and p53-AS and occurs through alternative splicing of intron 9 and 10, respectively.

To date, the biological functions of these isoforms remain unclear; however, the realization that p53 functional domains influence p53-dependent activities sheds new light on the importance of these isoforms.

[ILLUSTRATION OMITTED]

p53 Family Members Share Similar Structures

Recently, p63 and p73 proteins have been identified as p53 homologues. Multiple isoforms, which differ in the amino and carboxyl termini, have been identified for both p63 and p73.

p53 family members share significant similarity at the amino acid level within the activation, DNA-binding, and tetramerization domains. Like p53, both p63 and p73 bind to the canonical p53 responsive element, transactivate p53 target gene expression, and induce apoptosis when overexpressed.

Unlike p53, the genes encoding p63 and p73 are rarely mutated in human cancers. Because p63 and p73 knockout mice demonstrate discrete developmental abnormalities, rather than a propensity for tumor formation as do p53 knockout mice, the relationship between p63 or p73 and tumor suppression has been unclear.

However, recent evidence suggests that p63 and p73 may play important roles in tumor suppression, as mice heterozygous for p63 ([p63.sup.+/-]) or p73 ([p73.sup.+/-]) develop spontaneous tumors. Interestingly, these tumors have undergone loss of heterozygosity, indicating that the presence of a functional p63 or p73 can confer tumor suppression. In addition, combined mutation of p53 with p63 or p73 mutation confers greater tumor susceptibility than p53 mutation alone.

[ILLUSTRATION OMITTED]

p53 is Regulated Tightly in Normal Cells

Because p53 regulates many genes that in turn effect tumor suppression, the cell must be able to keep p53 silent under normal conditions as well as activate p53 within a moment's notice. Thus, the half-life of the p53 protein is very short during unstressed conditions, but it can be extended through posttranslational mechanisms after genotoxic insults.

To date, Mdm2 is the most critical negative regulator of p53. Mdm2, an E3 ubiquitin ligase, regulates the activity, subcellular localization, and stability of p53 during unstressed conditions. By binding to the N-terminus of p53 between residues 17--27, Mdm2 directly obscures AD1 of p53 and interferes with p53's ability to transactivate target genes.

Mdm2 also mediates nuclear export and nuclear exclusion of p53, which physically separates p53 from its target DNA. Once the Mdm2--p53 complex is removed to the cytoplasm, Mdm2 functions to ubiquitinate the C-terminal lysine residues of p53, which in turn targets p53 for proteosomal degradation. Interestingly, Mdm2 is a p53 target gene and thus functions in a negative feedback loop to keep p53 in check.

Cellular stress signals triggered by DNA damage, hypoxia, ionizing and UV radiation, or other genotoxic stresses lead to post-translational modifications of p53 that inhibit Mdm2 binding and result in activation of the p53 protein. These modifications include phosphorylation, acetylation, sumoylation, methylation, and proteinprotein interactions.

For example, ataxia telangiectasia-mutated (ATM), DNA-dependent protein kinase (DNA-PK), and Chk2 phosphorylate and stabilize p53 in response to ionizing radiation. ATR (ataxia-telangiectasia Rad3-related), Chk1, and homeo-domain-interacting protein kinase 2 (HIPK2) phosphorylate and stabilize p53 in response to UV radiation. N-terminal phosphorylation of p53 abolishes the interaction between p53 and Mdm2, preventing degradation of p53 via the proteosome pathway.

[ILLUSTRATION OMITTED]

p53 C-terminal lysine residues are acetylated by p300 and CBP. Acetylation of p53 prevents ubiquitination by Mdm2 as well as activates p53 DNA-binding activity. p53 also is regulated by many different protein-protein interactions that serve to influence p53 target gene transactivation and protein stability.

Interestingly, in addition to these many mechanisms of regulation, p53 also activates genes that in turn positively or negatively regulate the level and activity of p53 itself.

p53 Regulates Cell Cycle Arrest by Targeting Cyclins 

The cell cycle is a tightly controlled process in which a cell replicates its DNA and then divides into two daughter cells. To prevent transmission of genetically unstable material to daughter cells, progression through the cell cycle is regulated by a series of checkpoints: the G1/S, S, and G2/M checkpoints.

The G1/S and G2/M checkpoints center on the regulation and activation of cyclin-dependent kinase (CDK) complexes. Phosphorylation of target proteins by CDKs, which are serine/threonine kinases, leads to cell cycle progression.

Because CDK levels are stable throughout the cell cycle, CDK activity is regulated by their cyclin partners. In response to various stress signals, p53 inhibits cell cycle progression by transactivating target genes that initiate G1, S, and G2 arrest.

The G1/S checkpoint is regulated by the G1 cyclins (D-type and E-type cyclins) and CDKs 2, 4, and 6. A cell is maintained in G1 phase when the retinoblastoma protein (pRB) is in its active hypo-phosphorylated state. pRB is hyperphosphorylated by the G1 cyclin D/ CDK4/6 complex. Active pRB functions to inhibit the E2F transcription factor. The G1/S transition phase begins in late G1 when pRB is inactivated via hyper-phosphorylation (ppRB) by cyclin E/CDK4. Inactive ppRB releases E2F, which then induces genes required for entry into S phase.

Inhibition of the G1/S transition is mainly effected by cyclin-dependent kinase inhibitors (CKIs). CKIs inhibit CDKs by binding to cyclins, CDKs, or cyclin-CDK complexes. Without cyclin/CDK activity, E2F cannot induce genes required for S phase entry.

The G2/M checkpoint centers on activation of the cyclin B/cdc2 complex. The activation is regulated at many levels. The first level involves formation of the cyclin B/cdc2 complex. During G2, cyclin B is synthesized and forms an inactive cyclin B/cdc2 complex. The cyclin B/ cdc2 complex is activated by cdc25 phosphatase and CDK-activating kinase (CAK), which removes inhibitory phosphates at T14/Y15 and adds an activating phosphate at T161, respectively.

The cyclin B/cdc2 complex also is regulated by its subcellular localization. The complex remains inactive through nuclear export until the cell is ready to enter mitosis and nuclear export is blocked. Cyclin B/cdc2 levels then accumulate in the nucleus and initiate the G2/M transition.

In the event of a stress signal such as that from DNA damage, G1 and G2 arrest occurs. p53 mediates G1 arrest by inducing genes that inhibit cell cycle progression, specifically genes that inhibit the G1 cyclin/CDK complexes. The major mediator of p53-dependent G1 arrest is p21, a cyclin-dependent kinase inhibitor (CKI) of the Cip/Kip family. High levels of p21, induced by p53, stoichiometrically overwhelm the cyclin/CDK complexes and inhibit their activity. p53 also inhibits cyclin E by inducing hCDC4b, an F box protein and component of the SCF ubiquitin ligase complex, that targets cyclin E for proteosomal degradation.

p53 regulates the G2/M transition mainly by regulating genes that target the cyclin B/cdc2 complex. Interestingly, p53 is thought to repress cdc2, cyclin B, and cdc25c. Each of these proteins plays an important role during the G2/M transition. p21, the major mediator of G1 arrest, also plays a role in G2 arrest by inhibiting of the phosphorylation of cdc2 at T161.

In addition, p53 induces the growth arrest and DNA damage-inducible protein (GADD45), 14-3-3[sigma], MCG10, Reprimo, and B99. GADD45, the first identified p53 target gene, inhibits cyclin B/cdc2 activity by binding to cdc2 and preventing cyclin B/cdc2 complex formation. 14-3-3[sigma], a scaffold protein, removes cyclin B/cdc2 from the nucleus to physically separate cyclin B/cdc2 from its target proteins. MCG10, Reprimo, and B99 facilitate G2/M arrest through unknown mechanisms.

p53 Operates in Multiple DNA Repair Pathways

DNA repair helps to maintain genome fidelity through several pathways--for example, excision repair (consisting of both nucleotide excision repair [NER] and base excision repair [BER]), mismatch repair (MMR), and doublestrand break (DSB) repair. Each pathway uses unique enzymatic machinery, and as a guardian of the genome, p53 plays a role in each pathway.

[ILLUSTRATION OMITTED]

p53 participates in NER by inducing genes for xeroderma pigmentosum group E (XPE), XP group C (XPC), and GADD45. A defective NER pathway is causative of xeroderma pigmentosum (XP), an autosomal recessive disorder. Due to an inability to repair damaged DNA, patients with XP are extremely sensitive to UV light and have excessive skin cancers. At least seven genes, XPA to XPG, have been implicated in XP.

The p53 target gene XPE yields a protein that is a subunit of the UV-damaged DNA-binding protein (UV-DDB). XPE localizes to damaged DNA and enhances removal of UV photoproducts, such as cyclobutane pyrimidine dimers and 6-4 photoproducts. The protein XPC localizes to areas of UV-induced damage, where its interaction with damaged DNA is enhanced by XPE.

GADD45 is involved in NER, as GADD45-/- murine keratinocytes show impaired thymidine dimer repair and increased sensitivity to UV radiation.

[ILLUSTRATION OMITTED]

Interestingly, the p53 protein itself has been implicated in both NER and BER pathways. p53 interacts with several factors that play a role in NER as well as with DNA pol[beta]to stabilize the interaction between damaged DNA and the BER machinery.

p53 acts in the mismatch repair (MMR) pathway through the induction of p53R2. p53R2, which has homology to the R2 regulatory subunit of ribonucleotide reductase, functions in a general manner to increase the pool of free deoxyribonucleotides when repair is needed.

In the MMR pathway, p53 also acts by inducing human MutS homologue 2 (hMSH2) and proliferating cell nuclear antigen (PCNA). hMSH2 is integral to the MMR pathway, where it aids in mismatch recognition by binding to mismatched bases. Due to an inability to repair damaged DNA, mutations in the hMSH2 gene result in the hereditary nonpolyposis colorectal cancer (HNPCC) syndrome.

PCNA, a cofactor for DNA polymerase [delta], interacts with hMSH2 and facilitates hMSH2 transfer to mismatched bases.

p53 Inhibits Tumor Neovascularization

An important step in the growth of any tumor beyond a few millimeters is the generation of new blood supplies that feed and nurture the malignant cells. p53 regulates the process of angiogenesis through activation of genes that inhibit neovascularization, such as TSP-1, GD-AiF, and BAI-1, as well as repression of VEGF, an important gene that promotes neovascularization.

Thrombospondin-1 (TSP-1), an extracellular matrix glycoprotein, is secreted by many cell types and binds several cellular receptors. TSP-1 inhibits angiogenesis by interacting with the CD36 receptor on microvascular endothelial cells. This interaction culminates in apoptosis of the microvascular endothelial cell.

p53 inhibits angiogenesis in the brain by inducing glioma-derived angiogenesis inhibitory factor (GD-AiF) and brain specific angiogenesis inhibitor-1 (BAI-1). GD-AiF is a secreted inhibitor of angiogenesis, and BAI-1 is a seven transmembrane protein that suppresses tumor angiogenesis.

p53 represses vascular endothelial derived growth factor (VEGF). VEGF binds to the VEGF receptor on endothelial cells and enhances endothelial cell growth.

[ILLUSTRATION OMITTED]

p53 Inhibits Metastasis by Blocking ECM Degradation

Tissue invasion and metastasis involve a multistep process of invasion, intravasation, extravasation, and growth at a secondary site. Invasion and metastasis occur when genes that normally function to hold a cell in place lose their ability to do so.

Cell-cell adhesion is mediated by cadherins and cell-adhesion molecules (CAMS). Cadherins are transmembrane proteins that mediate [Ca.sup.2+]-dependent adhesion, and CAMS are cell surface proteins. The adhesion of cells to the extracellular matrix (ECM) is mediated by integrins, a family of transmembrane proteins. When cell-cell and cell-matrix interactions are disrupted, tumor cells break lose from each other and migrate.

Altered gene expression in tumor cells can lead to disruption of these interactions. For example, tumor cells may have increased extracellular protease activity and decreased protease inhibitor activity. ECM-degrading proteases, which include matrix metalloproteinases (MMPs) and serine proteases, degrade matrix proteins such as collagen, laminin, and fibronectin. MMPs are regulated by tissue inhibitors of MMPs (TIMPs). Serine proteases are regulated by serpins.

p53 inhibits metastasis by inducing genes that block ECM degradation and by repressing genes that degrade the ECM. p53 induces two serpins, plasminogen activator inhibitor-1 (PAI-1) and maspin. PAI-1 limits the metastatic potential of tumors by blocking ECM degradation. Specifically, PAI-1 inhibits urokinase-type plasminogen activator (u-PA), which initiates a cascade of cleavages that result in the activation of plasmin. Plasmin degrades a wide variety of ECM proteins, such as fibrin, fibronectin, and laminin.

Maspin, a serpin, does not utilize its protease inhibitor activity to inhibit migration or metastasis. Instead, maspin may increase cell adhesion to the ECM, as over-expression of maspin in a highly invasive mouse mammary tumor inhibits tumor growth and metastasis.

KAI-1/CD82 and Nm23-H1 are metastasis suppressor genes induced by p53. Furthermore, p53 also represses MMP1 and MMP13 (collagenases), both of which are involved in degrading the ECM.

[ILLUSTRATION OMITTED]

p53 Mediates Apoptosis via Two Pathways

Apoptosis, or programmed cell death, occurs when a cell is triggered, by either external or internal factors, to undergo a type of cell death that occurs without inflammation or necrosis. The characteristics of an apoptotic cell are nuclear condensation and fragmentation, membrane blebbing, and mitochondrial swelling.

Apoptosis can be initiated by either the extrinsic (death receptor) pathway or the intrinsic (mitochondrial) pathway, both of which involve p53-mediated processes.

Briefly, the extrinsic pathway is activated when an extracellular ligand binds a cell surface death receptor to signal the cleavage and activation of the initiator caspase 8. Caspases, a family of cysteine-dependent aspartate-directed proteases, remain quiet until they are activated by proteolytic cleavage.

In the intrinsic pathway, alteration of the mitochondrial membrane potential facilitates the release of cytochrome c and the subsequent activation of the initiator caspase 9.

The intrinsic and extrinsic pathways converge with the activation of effector caspases 3, 6, and 7 by the initiator caspases 8 and 9. The cleavage of nuclear and cytoskeletal structural proteins by effector caspases culminates in apoptosis.

Although many p53 target genes have been identified that play a role in p53-mediated apoptosis, no single essential gene for apoptosis has been identified. Because p53 regulates all known apoptotic pathways, it is likely that any given apoptotic target gene is only necessary for a specific apoptotic pathway.

To activate the death receptor pathway, p53 upregulates the cell surface death receptors Fas/ CD95 and KILLER/DR5, the Fas ligand (termed FasL/CD95L), as well as certain caspases. Both Fas and KILLER/DR5 are members of the tumor necrosis factor (TNF)/ nerve growth factor receptor superfamily. FasL/CD95L is a member of the TNF family.

Binding of FasL to Fas induces receptor trimerization and clustering of the intracellular death domain region of the receptor. By homotypic death domain interactions, Fas recruits the adaptor protein Fas-associated death domain (FADD). FADD contains a death effector domain. Through homotypic death effector domain interactions, FADD recruits procaspase 8 to form the death-induced signaling complex (DISC). Activation of the initiator caspase-8, another p53 target gene, eventually leads to the activation of effector caspases and the induction of apoptosis.

As more pro-caspase-8 molecules accumulate at the DISC, procaspase-8 undergoes transcatalysis, resulting in the activation of a large pool of the initiator caspase-8. p53 also has been shown to induce the expression of caspase-1 and caspase-6.

To regulate the amplitude of the apoptotic signal, p53 induces the inhibitor genes TRUNDD/TRAIL-R4, TRID/TRAIL-R3, and cFLIP. TRUNDD/TRAIL-R4 and TRIDD/TRAIL-R3 are TRAIL decoy death receptors that lack the death domain. Therefore, on ligand binding and receptor trimerization, the decoy receptors cannot recruit death effector domain-containing proteins required to initiate the caspase cascade. Cellular FLICE-like inhibitory protein (cFLIP) inhibits caspase-8 and prevents the formation of an active DISC.

In the mitochondrial pathway of apoptosis, p53 regulates many genes whose proteins localize to the mitochondria and regulate the release of cytochrome c. p53 target genes include Bcl-2 family proteins as well as non-Bcl-2 family members. In general, all Bcl-2 family members, which include both proand antiapoptotic proteins, contain Bcl-2 homology (BH) domains that are similar to four of the seven [alpha]-helices of Bcl-[X.sub.L], another Bcl-2 family member.

p53 target genes that belong to the Bcl-2 family include Bax, Noxa, p53-upregulated modulator of apoptosis (PUMA), BID, Bcl-2, and Bcl-x.

Bax is a proapoptotic Bcl-2 protein that contains BH1, BH2, and BH3 domains. The mechanism of Bax-induced apoptosis is complex. Bax translocates from the cytosol to the inner mitochondrial membrane and facilitates the release of cytochrome c from the mitochondria. In the cytosol, cytochrome c combines with Apaf-1 and caspase-9 to form the apoptosome, which activates effector caspases.

Noxa and PUMA are BH3-only containing Bcl-2 proteins. Through homotypic BH3 domain interactions, PUMA binds to Bcl-2 and Bcl-[X.sub.L] and induces cytochrome c release.

Recently, BID has been identified to bridge the extrinsic and intrinsic pathways for apoptosis. Cleavage of BID by caspase-8 exposes an N-terminal glycine that is subject to myristoylation. The cleaved, modified BID translocates to the mitochondria to participate in the mitochondrial apoptotic pathway.

Interestingly, p53 represses the antiapoptotic Bcl-2 family members Bcl-2 and Bcl-xL. These proteins function to protect the cell from apoptosis by stabilizing the mitochondrial membrane potential.

Non-Bcl-2 family genes targeted by p53 include mtCLIC/CLIC4 and p53AIP-1. mtCLIC, an organellular chloride channel protein of the CLIC family of intracellular chloride channels, reduces mitochondrial membrane potential. p53-apoptosis inducing factor 1 (p53AIP-1) localizes to the mitochondria and interacts with Bcl-2 to facilitate the release of cytochrome c from the mitochondria.

In addition to the mitochondrial proteins, p53 also induces intracellular regulators of the intrinsic pathway, i.e., apoptosis protease-activating factor 1 (Apaf-1) and the effector caspase-6. Apaf-1 combines with cytochrome c and caspase-9 to form the apoptosome.

Recently, studies have shown that in addition to p53 target genes, the p53 protein itself localizes to the mitochondria and perhaps alters the mitochondrial membrane potential. Thus, both p53 target genes and p53 protein facilitate induction of the intrinsic pathway.

[ILLUSTRATION OMITTED]

p53-Directed Strategies May Hold Therapeutic Potential

Because p53 function is lost in most human cancers, restoration of wild-type p53 activity in tumor cells is a promising therapeutic approach. Strategies include the use of gene replacement to introduce wild-type p53 into tumor cells with mutant p53; use of p53 status to destroy tumor cells that lack a functional p53; use of small molecules that restore a wild-type, active conformation to mutant p53; and use of agents that prevent the degradation of p53.

Adenoviral and retroviral vectors have been used to test the efficacy of p53 gene replacement as a cancer therapy. Unfortunately, clinical trials using p53 gene replacement have met limited success. This is likely due to incomplete restoration of wild-type p53 within the entire tumor. In addition, it is possible that viral gene transfer of p53 endows a low-level of p53 expression that facilitates cell cycle arrest but not apoptosis.

Despite the limited success of clinical trials to date, activation of wild-type p53 function in tumor cells remains an attractive potential therapy. With the increasing sensitivity and specificity of cancer diagnostics, it may become possible to detect malignancies at earlier stages of tumorigenesis. Strategies to turn on wild-type p53 activity during the early stages of tumorigenesis in these tumor cells may prove more effective.

A promising therapy focuses on inhibiting the p53--Mdm2 interaction. Inhibition of the p53--Mdm2 interaction promotes p53 stability as well as combats overexpression of Mdm2, which is found in many tumors. While many peptides have been identified that block the p53--Mdm2 interaction, the recent characterization of small molecule inhibitors, termed nutlins, has provided evidence that blocking the p53--Mdm2 interaction may be pharmaceutically feasible. Oral administration of nutlin to mice bearing established tumor xenografts of a human osteosarcoma cell line inhibited tumor growth by 90%.

Since the identification of p53 in 1979, many discoveries have changed the way scientists think about and study p53. For example, for the first 10 years after its identification, p53 was thought to be an immortalizing oncogene instead of a tumor suppressor. Thus, the discovery that p53 could suppress cell transformation was revolutionary.

The identification of p53 isoforms as well as the p53 family proteins p63 and p73 offer new areas for research. The identification and characterization of novel p53 target genes will help to dissect the complex signaling pathways of p53 in tumor suppression and may uncover means to harness the activity of this extraordinary protein.

DNA Repair

NER (nucleotide excision repair) repairs bulky lesions, such as UV-induced pyrimidine dimers. NER begins when the repairosome, a multi-enzyme complex, recognizes a bulky lesion. It cuts the DNA on both sides of the lesion, removes an oligonucleotide stretch, and then fills and seals the gap.

BER (base excision repair) occurs when NER neglects a modified base. Unlike NER, which removes an oligonucleotide stretch, BER removes only the damaged base.

MMR (mismatch repair) repairs mismatched bases, which occur as a result of spontaneous hydrolysis or incorrect incorporation during DNA replication. During MMR, a single mismatched nucleotide is replaced with the correct one.

DSB repair corrects double-strand breaks, which are induced by ionizing radiation or oxidative damage. DSB repair occurs by non-homologous end-joining or homologous recombination mechanisms.

Other p53-Dependent Processes

In addition to the cellular stress responses, p53 regulates genes that play roles in other p53-dependent activities. For example:

* p53 regulates many genes that function in a positive or negative feedback loop to regulate p53 stability and activity.

* p53 regulates genes that are involved in development.

* p53 regulates genes involved in cell death mediated by reactive oxygen species (ROS).

Publication date: 28 June 2005

RECENT REVIEWS

L.J. Ko and Carol Prives: p53: puzzle and paradigm. Genes Dev 10:1054-1072, 1 May 1996.

Douglas Hanahan and Robert A. Weinberg: The hallmarks of cancer. Cell 100:57-70, 7 Jan 2000.

Amy Willis and Xinbin Chen: The promise and obstacle of p53 as a cancer therapeutic agent. Curr Mol Med 2(4):329-345, 2002.

Kelly Harms, Susan Nozell, and Xinbin Chen: The common and distinct target genes of the p53 family transcription factors. Cell Mol Life Sci 61: 822-842, Apr 2004.

ORIGINAL PAPERS

Xinbin Chen, Gang Liu, Jianhui Zhu, et al: Isolation and characterization of fourteen novel putative and nine known target genes of the p53 family. Cancer Biol Ther 2:55-62, Jan-Feb 2003.

Lyubomir T. Vassilev, Binh T. Vu, Bradford Graves, et al: In vivo activation of the p53 pathway by small-molecule antagonists of MDM2. Science 303:844-848, 6 Feb 2004.

Kelly Lynn Harms and Xinbin Chen: The C terminus of p53 family proteins is a cell fate determinant. Mol Cell Biol 25:2014-2030, Mar 2005.

KELLY LYNN HARMS is a doctoral student in the Department of Cell Biology, and XINBIN CHEN is Professor of Cell Biology and Co-director of the Tumor Biology Program at the UAB Comprehensive Cancer Center, University of Alabama at Birmingham, Birmingham, Alabama.

[e-mail]

1979--p53 identified (considered
        an oncogene)

1989--p53 identified as a
        tumor suppressor

1990--Identified as DNA-binding
        transcription factor

1992--First p53 knockout mouse

1993--p53 negatively regulated
        by Mdm2
      p53 induces apoptosis
      p53 induces p21 for
        cell cycle arrest

1997--p73 identified

1998--p63 identified

2005

p53 GERMLINE MUTATIONS

% OF ALL TUMORS

BREAST            (27%)
SOFT TISSUE     (14.7%)
BRAIN           (13.9%)
BONE            (12.5%)
ADRENAL GLAND    (9.7%)
OTHER              (8%)
HEMATOLOGIC      (3.9%)
LUNG             (2.8%)
COLORECTUM       (1.9%)
STOMACH          (1.9%)
OVARY            (1.6%)
TESTIS          (0.63%)
KIDNEY          (0.47%)
LIVER           (0.31%)
HEAD & NECK     (0.31%)
PROSTATE        (0.31%)

Note: Table made from bar graph.

p53 SPONTANEOUS MUTATIONS

ESOPHAGUS       (46.9%)
OVARY           (46.4%)
COLORECTUM        (44%)
HEAD & NECK     (41.5%)
PANCREAS        (40.9%)
LUNG            (38.1%)
SKIN            (35.6%)
STOMACH         (30.3%)
BLADDER         (30.1%)
BRAIN           (27.6%)
LIVER           (26.2%)
BREAST          (25.6%)
UTERUS          (20.8%)
LYMPH NODES       (20%)
ENDOCRINE       (17.3%)
BONE            (16.7%)
SOFT TISSUES    (16.5%)
PROSTATE        (13.6%)
HEMATOPOEITIC   (13.2%)
CERVIX           (5.6%)

Note: Table made from bar graph.