Tetramerization Domain Of P53 Research Articles

Abstract 1176: Bypassing the dominant-negative effect of mutant p53 in cancer cells

Abstract The tumor suppressor p53 is mutated in more than 50% of all cancers. The transcriptional activity of p53 depends on localization to the nucleus and formation of p53 tetramers. Use of wild-type (wt) p53 for gene therapy is limited due to the dominant-negative effect of mutant (mut) p53 over wt p53. Mut p53 will hetero-oligomerize with wt p53 via the tetramerization domain (TD) found in p53, deactivating the tumor suppressor function of wt p53. In order to use p53 for gene therapy, an alternative oligomerization domain for p53 was investigated. The coiled-coil (CC) domain from Bcr (breakpoint cluster region), similar to the native TD of p53, can form an antiparallel dimer of dimers. The TD from p53 was then substituted with this CC, and the activity of this new p53 (p53-CC) was tested. We evaluated the transcriptional activity of p53-CC compared to wt p53 in a dual luciferase reporter gene assay. To demonstrate the ability of our p53-CC to bypass the dominant-negative effect prompted by hetero-oligomers, the reporter gene assay was carried in human breast adenocarcinoma (MCF-7) and human breast ductal carcinoma (T47D) cell lines. MCF-7 cells contain wt p53 population that is mislocalized to the cytoplasm, whereas, T47D cells contain mutant p53. Indeed, data from the reporter gene experiment shows a reduction in wt p53 activity introduced to T47D cells compared to MCF-7 cells, while the activity of p53-CC remains consistent regardless of the presence of an endogenous mut p53 population. We also tested the biological activity of p53-CC and its ability to induce apoptosis using 7-AAD and Annexin-V assays (late stage apoptosis). In addition, Caspase-3/7 assay (mid-stage apoptosis) was performed to further validate a p53-dependent apoptotic pathway. Data obtained from the aforementioned experiments suggest that p53-CC has a very similar biological activity to wt p53. Furthermore, we examined the gene expression profile of p53-CC using wt p53 as a reference. The assay was performed using real-time PCR array technology to analyze the expression of 84 genes related to p53-mediated signal transduction. Two negative controls were used; the first is a p53 construct that lacks its tetramerization domain (p53ΔTDC) and should not possess any transcriptional activity, and the second is the CC by itself. Indeed, p53-CC shows comparable gene expression profile to wt p53 and has the ability to transactivate the various p53 target genes. In summary, p53-CC retains p53 activity and excludes hetero-oligomer formation, overcoming a major barrier in using p53 for cancer gene therapy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1176. doi:1538-7445.AM2012-1176

Inhibition of tumor suppressor protein p53-dependent transcription by a tetramerization domain peptide via hetero-oligomerization

Tumor suppressor protein p53 induces cell cycle arrest, apoptosis, and senescence in response to cellular stresses. The p53 tetramer formation is essential for its functions. Despite of these crucial functions of p53 for integrity of genome, activation of the p53 signal pathway causes low induced pluripotent stem (iPS) cell generation efficiency. In this study, we report transient inhibition of p53-dependent transcription using a p53 tetramerization domain peptide that contains cell penetrating and nuclear localization signals. The peptide was efficiently introduced into cells and inhibited p21 expression via hetero-tetramerization with endogenous p53 protein. This method can be applied towards safe and efficient iPS cell generation.

On the Role of Flexibility in Protein-Ligand Interactions: the Example of p53 Tetramerization Domain

The recognition of protein surfaces by designed ligands has become an attractive approach in drug discovery. However, the variable nature and irregular behavior of protein surfaces defy this new area of research. The easy to understand "lock-and-key" model is far from being the ideal paradigm in biomolecular interactions and, hence, any new finding on how proteins and ligands behave in recognition events paves a step of the way. Herein, we illustrate a clear example on how an increase in flexibility of both protein and ligand can result in an increase in the stability of the macromolecular complex. The biophysical study of the interaction between a designed flexible tetraguanidinium-calix[4]arene and the tetramerization domain of protein p53 (p53TD) and its natural mutant p53TD-R337H shows how the floppy mutant domain interacts more tightly with the ligand than the well-packed wild-type protein. Moreover, the flexible calixarene ligand interacts with higher affinity to both wild-type and mutated protein domains than a conformationally rigid calixarene analog previously reported. These findings underscore the crucial role of flexibility in molecular recognition processes, for both small ligands and large biomolecular surfaces.

Abstract 1836: Distinct tumor spectrum and age of onset in a Brazilian cohort of p.R337H TP53 mutation carriers

Abstract The Li-Fraumeni syndrome (LFS) and its variant, Li-Fraumeni-like (LFL), are associated with high risk of early-onset tumors, typically breast cancer (BC), soft tissue sarcoma (STS), adrenocortical tumor (ADR) and brain cancer (BT). The major challenge in management of LFS/LFL is to provide screening for early detection due to the wide tumor spectrum and variable age of onset. Germline mutations in the TP53 gene are the only known genetic defect underlying LFS/LFL, detected in about 70% LFS and 20% LFL families. Most mutations occur in the exons encoding the DNA binding domain of p53 protein, whereas mutations outside these exons are rare and may have partial penetrance. A particular germline mutation at codon 337 (c.1010G>1, p.R337H) within the C-terminal, tetramerization domain of p53 is found in 0.3% of South and Southeastern Brazilian populations due to a founder effect. The aim of this study is to compare tumor spectrum in p.R337H carriers to that of other TP53 mutations. A cohort of 345 patients from 131 families was tested for germline TP53 mutations by sequencing the coding region (exons 2-11). Deletions were assessed by Multiple Ligand Polymorphism Assay. Ninety-one patients from 35 families harbored a TP53 mutation and are followed at the Department of Oncogenetics, Hospital A.C. Camargo in São Paulo, Brazil. Mutations were detected in 35/131 families (26.5%) and 88 tumors were documented in TP53 mutation carriers. Subjects and tumor′s data were managed and analyzed with a specifically designed software (HH-LFS) running on Excel (MicrosoftTM) and Visual Basic programming. The most common mutation was the founder p.R337H mutation (FM) p.R337H, present in 54.3% (19/35) of all families. Among 91 subjects found to carry a mutation, 70.3% (64/91) carried FM. Other mutations in the DNA binding domain (OM) were detected in 27 patients and large deletions occurred in two patients. The types of tumors in carriers of FM were similar to OM. However, there were significant differences in prevalence and/or age of occurrence. In both groups, the most common tumor was breast cancer, accounting for 35.4% in FM and 25% in OM. STS were detected in 14,5% in FM and 25% in OM. Interestingly, the third most common tumor among FM was thyroid cancer (TC) (14.5%), present in 7 patients from 3 distinct families whereas no TC were found in OM (p<0.001). BT and ADR were found in both groups with similar frequencies. Age at diagnosis varied between the two groups. Tumors were detected in 22% of FM before the age of 20 compared to 39.5% in OM (p=0.05). Conversely, 15.2% cancers in FM were above age 60 compared to 7.9% in OM. Gender distribution showed a similar predominance of tumors in females in both groups. The tendency of FM to develop a higher proportion of adult cancers as compared to OM suggests specific screening strategies for FM group. Moreover, the high prevalence of TC advocates for anual thyroid ultrasound screening in carriers of the p.R337H mutation. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1836. doi:10.1158/1538-7445.AM2011-1836

Probing phenylalanine environments in oligomeric structures with pentafluorophenylalanine and cyclohexylalanine

Stabilization of protein structures and protein-protein interactions are critical in the engineering of industrially useful enzymes and in the design of pharmaceutically valuable ligands. Hydrophobic interactions involving phenylalanine residues play crucial roles in protein stability and protein-protein/peptide interactions. To establish an effective method to explore the hydrophobic environments of phenylalanine residues, we present a strategy that uses pentafluorophenylalanine (F5Phe) and cyclohexylalanine (Cha). In this study, substitution of F5Phe or Cha for three Phe residues at positions 328, 338, and 341 in the tetramerization domain of the tumor suppressor protein p53 was performed. These residues are located at the interfaces of p53-p53 interactions and are important in the stabilization of the tetrameric structure. The stability of the p53 tetrameric structure did not change significantly when F5Phe-containing peptides at positions Phe328 or Phe338 were used. In contrast, the substitution of Cha for Phe341 in the hydrophobic core enhanced the stability of the tetrameric structure with a T(m) value of 100 degrees C. Phe328 and Phe338 interact with each other through pi-interactions, whereas Phe341 is buried in the surrounding alkyl side-chains of the hydrophobic core of the p53 tetramerization domain. Furthermore, high pressure-assisted denaturation analysis indicated improvement in the occupancy of the hydrophobic core. Considerable stabilization of the p53 tetramer was achieved by filling the identified cavity in the hydrophobic core of the p53 tetramer. The results indicate the status of the Phe residues, indicating that the "pair substitution" of Cha and F5Phe is highly suitable for probing the environments of Phe residues.

Enhancement of transcriptional activity of mutant p53 tumor suppressor protein through stabilization of tetramer formation by calix[6]arene derivatives

Li–Fraumeni syndrome, a hereditary disorder characterized by familial clusters of early-onset multiple tumors, is caused by mutation of the TP53 gene, which encodes the p53 tumor suppressor protein. Mutation of Arg337 to histidine in the tetramerization domain of p53 is most frequently observed in Li–Fraumeni syndrome. This mutation is reported to destabilize the tetrameric structure of p53. We designed and synthesized calix[6]arene derivatives, which have six imidazole or pyrazole groups at the upper rim. In this study, we report, for the first time, the enhancement of the in vivo transcriptional activity of the most common Li–Fraumeni p53 mutant by imidazole-calix[6]arene through stabilization of the oligomer formation.

Structural evolution of p53, p63, and p73: Implication for heterotetramer formation

Oligomerization of members of the p53 family of transcription factors (p53, p63, and p73) is essential for their distinct functions in cell-cycle control and development. To elucidate the molecular basis for tetramer formation of the various family members, we solved the crystal structure of the human p73 tetramerization domain (residues 351-399). Similarly to the canonical p53 tetramer, p73 forms a tetramer with D(2) symmetry that can be described as a dimer of dimers. The most striking difference between the p53 and p73 tetramerization domain is the presence of an additional C-terminal helix in p73. This helix, which is conserved in p63, is essential for stabilizing the overall architecture of the tetramer, as evidenced by the different oligomeric structures observed for a shortened variant lacking this helix. The helices act as clamps, wrapping around the neighboring dimer and holding it in place. In addition, we show by mass spectrometry that the tetramerization domains of p63 and p73, but not p53, fully exchange, with different mixed tetramers present at equilibrium, albeit at a relatively slow rate. Taken together, these data provide intriguing insights into the divergent evolution of the oligomerization domain within the p53 family, from the ancestral p63/p73-like protein toward smaller, less promiscuous monomeric building blocks in human p53, allowing functional separation of the p53 pathway from that of its family members.

P53 tetramerization domain mutations: germline R342X and R342P, and somatic R337G identified in pediatric patients with Li–Fraumeni syndrome and a child with adrenocortical carcinoma

Germline p53 mutations are associated with Li-Fraumeni syndrome (LFS) and other familial cancer phenotypes not fulfilling the definition for LFS. The majority of germline p53 mutations cluster in exons 5-8, corresponding to a DNA binding domain. We report the identification of two germline mutations and a somatic mutation in a tetramerization domain (TD), a rare site for mutations. The germline mutation, R342X (16915C>T), and the novel mutation, R342P (16916G>C), were found in a child with adrenocortical carcinoma and in a LFS pediatric patient with multiple primaries. The novel somatic mutation, R337G (16900C>G), was discovered in myelodysplastic syndrome with transformation to acute myeloblastic leukemia, developing as the third primary in the LFS child. These findings add further information on p53 TD mutations and TD contribution to tumorigenesis.

A Peptide-Based Dendrimer That Enhances the Splice-Redirecting Activity of PNA Conjugates in Cells

The full therapeutic potential of oligonucleotide (ON)-based agents has been hampered by cellular delivery challenges. Cell-penetrating peptides (CPP) represent promising delivery vectors for nucleic acids, and their potential has recently been evaluated using a functional splicing redirection assay, which capitalizes on the nuclear delivery of splice-correcting steric-block ON analogues such as peptide nucleic acids (PNA). Despite encouraging in vitro and in vivo data with arginine-rich CPP-steric block conjugates, mechanistic studies have shown that entrapment within the endosome/lysosome compartment after endocytosis remains a limiting factor. Previous work from our group has shown that CPP oligomerization greatly improves cellular delivery and increases transfection of plasmid DNA. We now report the chemical synthesis and the evaluation of multivalent CPP-PNA constructs incorporating monomeric (p53(mono)) and dendrimer-like tetrameric (p53(tet)) forms of the p53 tetramerization domain containing peptide, a 10 arginine CPP domain (R10), and a splice redirecting PNA (PNA705). These CPP-PNA conjugates were termed R10p53(tet)-PNA705 and R10p53(mono)-PNA705, referring to their oligomerization state. The present study demonstrates that the splicing redirection efficiency of PNA705 is much greater in the context of the tetrameric R10p53(tet)-PNA705 construct than for the monomeric and occurs at nanomolar concentrations, demonstrating that multivalency is an important factor in delivering PNA into cells.

TOE1 interacts with p53 to modulate its transactivation potential

The TOE1 gene was discovered as a target of the Egr1 transcription factor that participates in cell growth regulation through the upregulation of p21 and a cell cycle delay at the G2/M phase. We report here that TOE1 is able to bind to the p53 tumor suppressor protein, specifically interacting with the C terminal tetramerization domain of p53. We have further characterized this interaction through determination of binding kinetics using nanoporous optical interferometry and demonstrated that this interaction is capable of enhancing the transcriptional activity of p53-dependent gene targets. These results suggest a mechanistic role for TOE1 as a co-regulator of p53. Structured summary MINT- 7144895: P53 (uniprotkb: P04637) physically interacts (MI: 0914) with TOE1 (uniprotkb: Q96GM8) by pull down (MI: 0096) MINT- 7144912: TOE1 (uniprotkb: Q96GM8) binds (MI: 0407) to P53 (uniprotkb: P04637) by biophysical (MI: 0013) MINT- 7144884: P53 (uniprotkb: P04637) physically interacts (MI: 0914) with TOE1 (uniprotkb: Q96GM8) by anti bait coimmunoprecipitation (MI: 0006)

Modulation of the Oligomerization State of p53 by Differential Binding of Proteins of the S100 Family to p53 Monomers and Tetramers

We investigated the ways S100B, S100A1, S100A2, S100A4, and S100A6 bind to the different oligomeric forms of the tumor suppressor p53 in vitro, using analytical ultracentrifugation and multiangle light scattering. It is established that members of the S100 protein family bind to the tetramerization domain (residues 325-355) of p53 when it is uncovered in the monomer, and so binding can disrupt the tetramer. We found a stoichiometry of one dimer of S100 bound to a monomer of p53. We discovered that some S100 proteins could also bind to the tetramer. S100B bound the tetramer and also disrupted the dimer by binding monomeric p53. S100A2 bound monomeric p53 as well as tetrameric, whereas S100A1 only bound monomeric p53. S100A6 bound more tightly to tetrameric than to monomeric p53. We also identified an additional binding site for S100 proteins in the transactivation domain (1-57) of p53. Based on our results and published observations in vivo, we propose a model for the binding of S100 proteins to p53 that can explain both activation and inhibition of p53-mediated transcription. Depending on the concentration of p53 and the member of the S100 family, binding can alter the balance between monomer and tetramer in either direction.

Epstein–Barr virus nuclear antigen 3C targets p53 and modulates its transcriptional and apoptotic activities

The p53 tumor suppressor gene is one of the most commonly mutated genes in human cancers and the corresponding encoded protein induces apoptosis or cell-cycle arrest at the G1/S checkpoint in response to DNA damage. To date, previous studies have shown that antigens encoded by human tumor viruses such as SV40 large T antigen, adenovirus E1A and HPV E6 interact with p53 and disrupt its functional activity. In a similar fashion, we now show that EBNA3C, one of the EBV latent antigens essential for the B-cell immortalization in vitro, interacts directly with p53. Additionally, we mapped the interaction of EBNA3C with p53 to the C-terminal DNA-binding and the tetramerization domain of p53, and the region of EBNA3C responsible for binding to p53 was mapped to the N-terminal domain of EBNA3C (residues 130-190), previously shown to interact with a number of important cell-cycle components, specifically SCF(Skp2), cyclin A, and cMyc. Furthermore, we demonstrate that EBNA3C substantially represses the transcriptional activity of p53 in luciferase based reporter assays, and rescues apoptosis induced by ectopic p53 expression in SAOS-2 (p53(-/-)) cells. Interestingly, we also show that the DNA-binding ability of p53 is diminished in the presence of EBNA3C. Thus, the interaction between the p53 and EBNA3C provides new insights into the mechanism(s) by which the EBNA3C oncoprotein can alter cellular gene expression in EBV associated human cancers.

Conformational detection of p53’s oligomeric state by FlAsH Fluorescence

The p53 tumor suppressor protein is a critical checkpoint in prevention of tumor formation, and the function of p53 is dependent on proper formation of the active tetramer. In vitro studies have shown that p53 binds DNA most efficiently as a tetramer, though inactive p53 is predicted to be monomeric in vivo. We demonstrate that FlAsH binding can be used to distinguish between oligomeric states of p53, providing a potential tool to explore p53 oligomerization in vivo. The FlAsH tetra-cysteine binding motif has been incorporated along the dimer and tetramer interfaces in the p53 tetramerization domain to create reporters for the dimeric and tetrameric states of p53, though the geometry of the four cysteines is critical for efficient FlAsH binding. Furthermore, we demonstrate that FlAsH binding can be used to monitor tetramer formation in real-time. These results demonstrate the potential for using FlAsH fluorescence to monitor protein–protein interactions in vivo.

Molecular basis of Pirh2-mediated p53 ubiquitylation

Pirh2 (p53-induced RING-H2 domain protein; also known as Rchy1) is an E3 ubiquitin ligase involved in a negative-feedback loop with p53. Using NMR spectroscopy, we show that Pirh2 is a unique cysteine-rich protein comprising three modular domains. The protein binds nine zinc ions using a variety of zinc coordination schemes, including a RING domain and a left-handed beta-spiral in which three zinc ions align three consecutive small beta-sheets in an interleaved fashion. We show that Pirh2-p53 interaction is dependent on the C-terminal zinc binding module of Pirh2, which binds to the tetramerization domain of p53. As a result, Pirh2 preferentially ubiquitylates the tetrameric form of p53 in vitro and in vivo, suggesting that Pirh2 regulates protein turnover of the transcriptionally active form of p53.

Members of the S100 family bind p53 in two distinct ways

p53 binds to some members of the S100 family (S100B, S100A4, S100A2, and S100A1). We previously showed that both S100B and S100A4 bind to the p53 tetramerization domain, and consequently control its oligomerization state, but only S100B binds to the C-terminal negative regulatory domain (NRD). Here, we investigate other binding partners for p53 within the S100 family (S100A6 and S100A11), and show that binding to the p53 tetramerization domain seems to be a general feature of the S100 family, while binding to the NRD is a characteristic of a subset of the family.

Oxidation of methionine residue at hydrophobic core destabilizes p53 tetrameric structure

The tumor suppressor protein p53 is a tetrameric phosphoprotein that induces cell cycle, development, and differentiation by regulating the expression of target genes. The tetramerization of p53 is essential for its tumor suppressor functions. It has been known that oxidation of proteins affects their structure and function. A methionine residue (Met340) is located at the hydrophobic core in p53 tetramerization domain. Here, we demonstrated that Met340 residue can be oxidized to methionine sulfoxide under oxidative conditions and investigated effects of the oxidation of p53 tetramerization domain on its stability and oligomerization state by CD measurement and gel filtration. The oxidation of Met340 drastically induced destabilization of the p53 tetramer by 22.8 kJ/mol of DeltaDeltaG(Tm), while retaining the identical conformation as that of the wild-type peptide. Trypsin digestion experiments also showed that oxidation of Met340 allowed the peptide to form locally loose structure and become more sensitive to enzyme degradation. The tetrameric structure may be destabilized because the oxidation of Met340 induces charge repulsion and/or steric hindrance between the sulfoxide groups. These results taken together suggested that oxidation of methionine residues in the p53 protein might be one of the inactivation mechanisms of p53 transcriptional function under conditions of oxidative stress.

BAF60a Interacts with p53 to Recruit the SWI/SNF Complex

To understand the tumor-suppressing mechanism of the SWI/SNF chromatin remodeling complex, we investigated its molecular relationship with p53. Using the pREP4-luc episomal reporter, we first demonstrated that p53 utilizes the chromatin remodeling activity of the SWI/SNF complex to initiate transcription from the chromatin-structured promoter. Among the components of the SWI/SNF complex, we identified BAF60a as a mediator of the interaction with p53 by the yeast two-hybrid assay. p53 directly interacted only with BAF60a, but not with other components of the SWI/SNF complex, such as BRG1, SRG3, SNF5, or BAF57. We found out that multiple residues at the amino acid 108-150 region of BAF60a were involved in the interaction with the tetramerization domain of p53. The N-terminal fragment of BAF60a containing the p53-interacting region as well as small interfering RNA for baf60a inhibited the SWI/SNF complex-mediated transcriptional activity of p53. The uncoupling of p53 with the SWI/SNF complex resulted in the repression of both p53-dependent apoptosis and cell cycle arrest by the regulation of target genes. These results suggest that the SWI/SNF chromatin remodeling complex is involved in the suppression of tumors by the interaction with p53.

Activation of p53-dependent responses in tumor cells treated with a PARC-interacting peptide

We tested the activity of a p53 carboxy-terminal peptide containing the PARC-interacting region in cancer cells with wild type cytoplasmic p53. Peptide delivery was achieved by fusing it to the TAT transduction domain (TAT-p53-C-ter peptide). In a two-hybrid assay, the tetramerization domain (TD) of p53 was necessary and sufficient to bind PARC. The TAT-p53-C-ter peptide disrupted the PARC–p53 complex. Peptide treatment caused p53 nuclear relocation, p53-dependent changes in gene expression and enhancement of etoposide-induced apoptosis. These studies suggest that PARC-interacting peptides are promising candidates for the enhancement of p53-dependent apoptosis in tumors with wt cytoplasmic p53.

Using peptides to study the interaction between the p53 tetramerization domain and HIV‐1 tat

Peptides are valuable tools for studying protein-protein interactions, especially in cases of isolated protein domains and natively unfolded proteins. Here, we used peptides to quantitatively characterize the interaction between the natively unfolded HIV-1 Tat protein and the tetramerization domain of the cellular tumor suppressor protein p53. We used peptide mapping, fluorescence anisotropy, and NMR spectroscopy to perform a detailed structural and biophysical characterization of the interaction between the two proteins and elucidate its molecular mechanism, which have so far been studied using cell-based methods. We show that the p53 tetramerization domain, p53(326-355), binds directly to residues 1-35 and 47-57 in Tat. We have characterized the interaction between p53(326-355) and Tat(47-57) in detail. The p53 residues that are mainly involved in binding to Tat(47-57) are E343 and E349, which bind to the positively charged arginine-rich motif of Tat by a partly electrostatic mechanism. All oligomerization states of p53(326-355) bind Tat(47-57) without inhibiting p53 tetramerization, since the residues in p53(326-355) that bind Tat(47-57) face away from the tetramerization interface. We conclude that p53 is able to bind Tat as a transcriptionally active tetramer.

Regulation of p53 tetramerization and nuclear export by ARC

Inactivation of the transcription factor p53 is central to carcinogenesis. Yet only approximately one-half of cancers have p53 loss-of-function mutations. Here, we demonstrate a mechanism for p53 inactivation by apoptosis repressor with caspase recruitment domain (ARC), a protein induced in multiple cancer cells. The direct binding in the nucleus of ARC to the p53 tetramerization domain inhibits p53 tetramerization. This exposes a nuclear export signal in p53, triggering Crm1-dependent relocation of p53 to the cytoplasm. Knockdown of endogenous ARC in breast cancer cells results in spontaneous tetramerization of endogenous p53, accumulation of p53 in the nucleus, and activation of endogenous p53 target genes. In primary human breast cancers with nuclear ARC, p53 is almost always WT. Conversely, nearly all breast cancers with mutant p53 lack nuclear ARC. We conclude that nuclear ARC is induced in cancer cells and negatively regulates p53.