PTEN: a New Guardian of the Genome

Home , Germline mutation, Major vault protein, PTEN (gene), Tumor suppressor gene

Y Yin and WH Shen

Department of Radiation Oncology, College of Physicians and Surgeons, Columbia University, New York, NY, USA

The phosphatase and tensin homolog deleted on chromo- and Peacocke, 1998). Hallmark features of Cowden some 10 (PTEN)tumor suppressor is a phosphatase that disease are hamartomatous overgrowths of various antagonizes the phosphoinositol-3-kinase/AKT signaling tissues, including skin, breast, intestine and brain, as pathway and suppresses cell survival as well as cell well as a predisposition to the development of breast proliferation. PTEN is the second most frequently and thyroid tumors (Liaw et al., 1997). Homozygous mutated gene in human cancer after p53. Germline deletion of Pten causes embryonic lethality, suggesting mutations of PTEN have been found in cancer suscept- that PTEN is essential for embryonic development (Di ibility syndromes, such as Cowden syndrome, in which Cristofano et al., 1998). Mice heterozygous for Pten over 80% of patients have mutations of PTEN. Homo- deletion develop tumors in multiple tissues comparable zygous deletion of Pten causes embryonic lethality, with those affected in humans (Di Cristofano et al., suggesting that PTEN is essential for embryonic devel- 1998; Suzuki et al., 1998; Podsypanina et al., 1999; opment. Mice heterozygous for Pten develop spontaneous Stambolic et al., 2000). These in vivo observations tumors in a variety of organs comparable with the indicate that the homeostatic balance has been disrupted spectrum of its mutations in human cancer. The mechan- and shifted toward oncogenesis, which is likely due to isms of PTEN functions in tumor suppression are lack of PTEN function in maintaining normal cellular currently under intense investigation. Recent studies processes. Indeed, studies in cell culture reveal a demonstrate that PTEN plays an essential role in the plethora of phenotypic changes in PTEN-deﬁcientcells, maintenance of chromosomal stability and that loss of such as increased proliferation (Sun et al., 1999; PTEN leads to massive alterations of chromosomes. The Backman et al., 2001), reduced apoptosis (Stambolic tumor suppressor p53 is known as a guardian of the et al., 1998) and enhanced migration (Tamura et al., genome that mediates the cellular response to environ- 1998; Liliental et al., 2000). In contrast, overexpression mental stress, leading to cell cycle arrest or cell death. of wild-type PTEN in cancer cells induces apoptosis and Through completely different mechanisms, PTEN also blocks cell-cycle progression, colony formation and cell protects the genome from instability. Thus, we propose migration (Gu et al., 1998; Li et al., 1998; Tamura et al., that PTEN is a new guardian of the genome. In this 1998; Weng et al., 2001a, b). Intensive efforts have been review, we will discuss new discoveries on the role of focused on how PTEN acts to regulate such a broad PTEN in tumor suppression and explore mechanisms by spectrum of phenotypic features, answers to which may which PTEN maintains genomic stability. come from a better understanding of its biochemical Oncogene (2008) 27, 5443–5453; doi:10.1038/onc.2008.241 features, structure, subcellular localization and signaling targets. Keywords: PTEN; p53; tumor suppressor; genome guardian; genomic instability; chromosome instability PTEN: structure and functional domains

PTEN as a powerful tumor suppressor There are two major domains of PTEN, the N-terminal phosphatase domain and the C-terminal domain (Lee Phosphatase and tensin homolog deleted on chromo- et al., 1999). Although the role of PTEN in tumor some 10 (PTEN)/MMAC1 was cloned in 1997 through suppression has been mostly attributed to its N-terminal its association with the human cancer susceptibility lipid phosphatase activity (Cantley and Neel, 1999), locus at10q23 (Li and Sun, 1997; Li et al., 1997; Steck more than 40% of PTEN tumorigenic mutations occur et al., 1997). PTEN is one of the most frequently in the C-terminal domain (Waite and Eng, 2002). This mutated genes in human cancers (Cantley and Neel, fact indicates that PTEN may have other functions 1999; Simpson and Parsons, 2001). Germline PTEN through its C terminal domain, which is significant in mutations are primarily found in the autosomal tumor suppression. The C-terminal domain contains dominanthamartomasyndrome Cowden disease (Eng both a C2 domain and a tail region that may be related to PTEN stability (Georgescu et al., 1999) and protein– protein interaction (Fanning and Anderson, 1999). The Correspondence: Dr Y Yin, Department of Radiation Oncology, College of Physicians and Surgeons, Columbia University, VC11-213, C2 domain has been implicated in PTEN stability 630 West 168th Street, New York, NY 10032, USA. (Georgescu et al., 2000) and its recruitment to phos- E-mail: [email protected] pholipid membranes (Das et al., 2003). Crystal structure PTEN maintains genomic stability through multiple mechanisms Y Yin and WH Shen 5444 analysis of this domain revealed a b-sandwich structure Cdc42, PTEN can also regulate cell motility by directly (Lee et al., 1999), suggesting a basis for its interaction targeting and dephosphorylating Shc kinase, thereby with DNA and other proteins. It was reported that inhibiting the mitogen-activated protein (MAP) kinase PTEN uses the C2 domain to interact with the signaling pathway (Gu et al., 1998; Tamura et al., 1998). centromere (Shen et al., 2007). As an important intracellular signaling pathway, the MAP kinase cascade provides multiple potential targets for PTEN. PTEN can inactivate multiple membrane- PTEN as a dual lipid and protein phosphatatse proximal proteins upstream of MAP kinase such as Ras and IRS-1 (Gu et al., 1998; Weng et al., 2001b). The PTEN is a lipid and protein phosphatase. PTEN protein prototypical MAP kinase, extracellular signal-regulated phosphatase can dephosphorylate protein substrates on kinase, is often affected by PTEN status. However, serine, threonine and tyrosine residues (Myers et al., PTEN is unable to directly dephosphorylate extracel- 1997). The lipid phosphatase activity of PTEN lular signal-regulated kinase (Myers et al., 1997), was reported for the substrate phosphoinositol 3,4,5- indicating that extracellular signal-regulated kinase is triphosphate (PIP3), a key signaling component of not a direct substrate of PTEN. The protein phospha- the phosphoinositol-3-kinase (PI3-kinase) pathway tase activity of PTEN has also been demonstrated to (Maehama and Dixon, 1998). The PI3-kinase/Akt inhibit FAK and extracellular signal-regulated kinase pathway is regarded as the primary physiological target and subsequently block the expression and secretion of of PTEN (Sulis and Parsons, 2003; Leslie and Downes, matrix metalloproteinase-9, which may contribute to the 2004; Sansal and Sellers, 2004). Phosphatase activity, suppression of glioblastoma invasion (Park et al., 2002). particularly lipid phosphatase activity, is believed to be A recent report reveals that JNK is a functional target of the most pertinent property of PTEN as a tumor PTEN (Vivanco et al., 2007), in which protein phos- suppressor. The role of PTEN as a lipid phosphatase in phatase activity of PTEN may not be directly involved. growth suppression can be distinguished by a germline PTEN can also inhibit phosphorylation of proteins missense mutation, G129E, which abrogates lipid downstream of MAP kinases. One prominent example is phosphatase but not protein phosphatase activity (Liaw ETS-2, a nuclear target of the MAP kinase pathway and et al., 1997). Most naturally occurring mutations, a transcription factor whose DNA-binding ability is however, are both lipid and protein phosphatase controlled by phosphorylation. PTEN may use its inactive, such as C124S (Maier et al., 1999). Double protein phosphatase activity to block ETS-2 phospho- phosphatase-deficient PTEN (C124S) (Maier et al., rylation in response to insulin-induced MAP kinase 1999) and lipid phosphatase-deficient PTEN (G129E) activation, in a PI3-kinase-independent manner (Weng (Gildea et al., 2004) have been shown to inhibit cell et al., 2002). PTEN can also regulate phosphorylation of invasion similar to wild type. These results imply another transcription factor Sp1, likely through its that PTEN may suppress tumors independent of its protein phosphatase activity (Kang-Park et al., 2003). phosphatase activity. In addition to intracellular signaling molecules, PTEN phosphatase also targets different proteins. receptor tyrosine kinases may also serve as direct Focal adhesion kinase (FAK), a nonreceptor protein protein targets of PTEN. PTEN physically associates tyrosine kinase, has been identified as a direct protein with the receptor of platelet-derived growth factor and target of PTEN (Tamura et al., 1998). Similarly, PTEN directly dephosphorylates the receptor, whereas the also reduces the tyrosine phosphorylation of p130Cas,a phosphatase-deficient PTEN mutant (C124S) not only FAK downstream effector (Tamura et al., 1998). By fails to dephosphorylate the platelet-derived growth targeting and dephosphorylating FAK and p130Cas, factor receptor but also acts in a dominant-negative PTEN regulates dynamic cell surface interactions and fashion to increase its phosphorylation (Mahimainathan inhibits cell migration and invasion. The lipid phospha- and Choudhury, 2004). tase activity of PTEN is dispensable for tyrosine Collectively, potential protein targets of PTEN range dephosphorylation of FAK, as the lipid phosphatase- from membrane-bound receptor tyrosine kinases and deficient G129E mutant retains this activity. In compar- cytoplasmic signaling molecules to transcription factors ison with PIP3 as a ubiquitous target of PTEN lipid in the nucleus. Nevertheless, the currently recognized phosphatase, FAK/p130Cas as targets of PTEN protein candidates are mostly identified from in vitro studies and phosphatase appear to be conditional. Although over- many of themare found in some butnotothercell expression of PTEN dephosphorylates FAK/p130Cas systems. Therefore, the identification of more direct (Tamura et al., 1998), their phosphorylation status functional protein substrates of the PTEN phosphatase remains unaffected in PtenÀ/À cells (Liliental et al., deserves further investigation. 2000), suggesting that PTEN is not a sole regulator of FAK/p130Cas. Interestingly, PtenÀ/À cells indeed exhibit increased cell motility, which is accompanied by induced PTEN mutations in human cancers: a database activities of two small GTPases, Rac1 and Cdc42. In this to be updated case, however, PTEN lipid phosphatase activity seems to be involved in the inhibition of cell motility and The cloning of the PTEN gene was accompanied by phosphorylation of both Rac1 and Cdc42 (Liliental detection of various types of mutations including et al., 2000). In addition to FAK/p130Cas, Rac1 and homozygous deletion, frameshift, inframe deletion,

Oncogene PTEN maintains genomic stability through multiple mechanisms Y Yin and WH Shen 5445 truncation, point mutation and so on (Li et al., 1997; yan–Zonana syndrome) and C124S. The affected Shin et al., 2002). Over the last decade following the residues are located either in the catalytic motif (H123, discovery of this powerful tumor suppressor, numerous C124 and G129) or a conserved a-helix that is critical for PTEN mutations have been identified in a wide range of maintaining secondary structure of the phosphatase sporadic malignancies and ata high frequency in cancer- (L57, G165, T167 and S70) (Myers et al., 1997). susceptibility syndromes. PTEN mutations occur most Unfortunately, there are no reported PTEN mutations frequently in three types of human cancer: glioblastoma, that are only deficient for protein phosphatase activity. endometrial and prostate cancer (Supplementary Table Nevertheless, these naturally occurring PTEN muta- 1). The firstreporton PTEN mutation in primary tions have contributed significantly to our understand- glioblastoma revealed a high mutational frequency ing of PTEN function. (44%) among 34 cases of glioblastoma multiforme Germline PTEN mutations are associated with 80% of (Wang et al., 1997). Subsequentmutationalanalyses Cowden syndrome cases and 60% of Bannayan–Riley– using different sample numbers also indicate glioblasto- Ruvalcaba syndrome cases (Eng, 2003). Both of these ma as one of the cancer types bearing frequent PTEN cancer susceptibility syndromes are now referred to mutations (Table 1). Owing to limited sample sizes in collectively as the PTEN hamartoma tumor syndrome. most of these studies, the frequencies of PTEN The above-mentioned PTEN mutations are mostly mutations vary from 9 to 44%. A summary of all these located in the N-terminal region of the PTEN molecule, studies gives rise to a frequency of 28.8% with PTEN which conveys its phosphatase catalytic activity. How- mutation in a total of 605 glioblastomas. Similarly, ever, over 40% of the PTEN mutations are found in the information about the frequencies of PTEN mutation in C-terminal region (Waite and Eng, 2002). Moreover, other types of human cancers has also been summarized many C-terminal truncated forms of PTEN have been in Table 1. In addition to the frequency, consolidation found in Cowden syndrome, due to the generation of a of information on position and type of each detected stop codon from a single base substitution (Iida et al., mutation would help in identifying hot spots in different 1998, 2000; Kanaseki et al., 2002; McGarrity et al., 2003; cancers, and understanding how PTEN functions in the Agrawal et al., 2005). These C-terminal truncation etiology and molecular pathogenesis of human cancer. It PTEN mutations are particularly important for func- has been over 10 years since PTEN was discovered. A tional characterization of the C-terminal domain of mutational database for this second most frequently PTEN. Recent studies have revealed the essential role of mutated tumor suppressor is warranted. the C-terminal region of PTEN in determining sub- Mutational studies have indeed revealed some critical cellular PTEN localization (Trotman et al., 2007; Wang residues for PTEN function. The most prominent et al., 2007) and maintaining genomic stability (Shen mutation, G129E, was found in Cowden disease kindred et al., 2007). The C-terminal region is required for PTEN (Liaw et al., 1997). This famous mutation, in fact, led to function to stabilize centromeres, although the phospha- the milestone finding that PTEN has a lipid phosphatase tase activity of PTEN is dispensable (Shen et al., 2007). activity because the mutant is deficient specifically in its Interestingly, phosphatase-deficient PTEN mutants, ability to dephosphorylate PIP3 (Myers et al., 1998). C124S and G129E, retain the ability to localize in the Interestingly, this mutation loses its specificity when the nucleus and physically associate with the kinetochore same glycine residue is mutated to arginine (G129R) component CENP-C, whereas a C-terminal truncated instead of glutamate (G129E) (Myers et al., 1997, 1998). PTEN mutant, PTEN189, fails to do so (Shen et al., G129R is also a naturally occurring mutation in 2007). Similarly, a C-terminal point mutation, K289E, glioblastoma (Li et al., 1997). Whereas G129E retains confers a nuclear importdefect,although itdoes not the activity of dephosphorylating protein substrates, affect phosphatase activity (Trotman et al., 2007). These G129R loses both lipid phosphatase and protein important C-terminal mutations have been very helpful phosphatase activity (Myers et al., 1997, 1998). In in identifying the new functions of PTEN, particularly of addition to G129R, many missense point mutations of nuclear PTEN. PTEN found in primary human cancers have similar Identification and characterization of these PTEN functional deficiencies, including H123Y (detected in mutations in various types of human cancer further endometrial cancer), L57W (glioblastoma), G165R supports the need for a PTEN mutational database. (glioblastoma), T167P (breast cancer), S70R (Banna- This database could greatly contribute to the continuing

Table 1 PTEN mutations in different tumor types Tumor types Mutation frequency (%) Specimens with PTEN mutation Specimens analyzed

Glioblastoma 28.8 174 605 Endometrial carcinoma 34.6 148 428 Melanoma 12.1 17 141 Prostate cancer 11.8 18 152 Breastcancer 3.5 9 254

Mutation frequencies summarized in Supplementary Table 1 are on the basis of currently available reports cited in Supplementary information.

Oncogene PTEN maintains genomic stability through multiple mechanisms Y Yin and WH Shen 5446 discoveries of novel functions of PTEN and to a with centromeres. Second, PTEN may be necessary for comprehensive understanding of how this versatile DNA repair because loss of PTEN results in a high tumor suppressor safeguards important cellular frequency of double-strand breaks. PTEN affects machineries againsttumorigenesis. double-strand breaks through regulation of Rad51, a key componentfor homologous recombinationrepair of DNA double-strand breaks. Consistently, regulation of Maintenance of genomic stability: novel functions PTEN nuclear localization has also been reported of PTEN (Trotman et al., 2007; Wang et al., 2007). Ithas been demonstrated that PTEN physically associates with an Genome or chromosome instability is a hallmark of integral component of centromeres in the nucleus, cancers. Tumor suppressors play roles in maintaining disruption of which causes centromere breakage and genome stability, and loss of function of these tumor massive chromosomal aberrations (Shen et al., 2007). suppressors therefore leads to genomic instability. These new findings revealed the novel function of PTEN Genetic instability represents an inevitable consequence in guarding the genome and propelled PTEN research of the loss of tumor suppressors. Indeed, the frequent into the ‘nuclear age’ (Baker, 2007). occurrence of PTEN mutation and genetic instability is The cytoplasm has been considered as the primary site found in a large range of PTEN-deficienthuman cancers for PTEN to elicit its tumor-suppressive function, and (Cantley and Neel, 1999; Simpson and Parsons, 2001). the ability of PTEN to block the PI3-kinase pathway However, the function of PTEN in maintaining genetic through its phosphatase activity has been regarded as stability had not been recognized and demonstrated the key mechanism by which PTEN suppresses carcino- until recently. The first link between PTEN deficiency genesis (Parsons, 2004; Cully et al., 2006). However, the and genetic instability was revealed by the pioneer work discoveries of PTEN localization and novel functions in of the Parsons’ group (Puc et al., 2005). Pten-null the nucleus have expanded its role in tumor suppression. embryonic stem cells were shown to exhibit DNA repair Although the cellular distribution of PTEN varies in checkpoint defects in response to ionizing radiation, differenttissues,endogenous PTEN in neurons, gliomas which results in the accumulation of unrepaired and cells of the thyroid, pancreas and skin is found chromosomes with DNA double-strand gaps and mostly in the nuclear compartment (Sano et al., 1999; breaks. Further mechanistic study suggested that the Gimm et al., 2000; Lachyankar et al., 2000; Perren et al., observed G2 checkpointdefectsmay have resultedfrom 2000; Whiteman et al., 2002). Growing evidence has functional impairment of an important checkpoint suggested that malignancies may be accompanied protein, CHK1, due to lack of PTEN. PTEN deficiency by translocation of PTEN from the nucleus to the directly elevates AKT kinase activity, which triggers cytoplasm (Gimm et al., 2000; Perren et al., 2000; CHK1 phosphorylation (Figure 1). Phosphorylated Whiteman et al., 2002). The function of nuclear PTEN CHK1 undergoes ubiquitination, which prevents its may deviate from its role in regulating PIP3 at the entry into the nucleus. Sequestering CHK1 in the plasma membrane, but is consistent with an alternative cytoplasm impairs its normal function in initiating a role in controlling genetic stability and involving DNA repair checkpoint. In addition to G2 checkpoint chromatin function. In addition, high expression levels defects, CHK1 inactivation in Pten-deficientcells can of nuclear PTEN have recently been associated with cell- lead to the accumulation of DNA double-strand breaks cycle arrestatthe G0/G1 phase (Ginn-Pease and Eng, (Puc and Parsons, 2005). These in vitro observations 2003), indicating a likely role of nuclear PTEN in cell have also been demonstrated in vivo. Examination of growth inhibition. Similarly, multiple aspects of the CHK1 localization in a large panel of primary human anti-oncogenic function of PTEN, such as regulation of breast carcinomas indicates an increased cytoplasmic cell growth and migration, are found independent of its level of CHK1 in tumor cells with lower expression of phosphatase activity (Freeman et al., 2003; Raftopoulou PTEN and elevated AKT phosphorylation. Further- et al., 2004; Okumura et al., 2005; Liu et al., 2005b; more, aneuploidy was frequently observed in both Tang and Eng, 2006; Leslie et al., 2007). Indeed, somatic human breastcarcinomas withlow expression of PTEN PTEN mutations do occur outside the phosphatase and prostatic intraepithelial neoplasia from Pten þ /À domain (Eng, 2003), implying that PTEN can function mice (Puc and Parsons, 2005). These consistent in vitro in tumor suppression through additional activities and in vivo observations suggest that PTEN deficiency besides antagonism of the PI3-kinase pathway. These may initiate an oncogenic signaling process by causing earlier observations form a growing pool of evidence dysfunction of important checkpoint proteins. that supports the idea that PTEN may possess other The essential role of nuclear PTEN in the main- functions beyond those exhibited in the cytoplasm and tenance of chromosomal stability has been demon- those dependent upon its phosphatase activity. As strated recently in both mouse and human systems shown in Figure 1, PTEN localizes atcentromeres and (Shen et al., 2007). As shown in Figure 1, nuclear PTEN interacts with CENP-C, an integral kinetochore compo- may utilize two novel mechanisms to maintain chromo- nent, which is crucial for protection against chromosome integrity. First, PTEN interacts with centromeres some instability. Importantly, it is found that physical and maintains their stability. It is believed that PTEN association between PTEN and centromeres/kineto- does so through its C2 domain, as mutant PTEN chores does not require the phosphatase activity of without this C2 domain loses the capability to interact PTEN, because a phosphatase-deficient PTEN mutant

Oncogene PTEN maintains genomic stability through multiple mechanisms Y Yin and WH Shen 5447

Figure 1 A model of PTEN as a guardian of the genome. PTEN maintains genomic stability through multiple mechanisms. In the cytoplasm, PTEN antagonizes phosphoinositol-3-kinase by dephosphorylating PIP3 and prevents Akt activation and Chk1 phosphorylation, releasing Chk1 into the nucleus for DNA repair. PTEN can shuttle between the cytoplasm and the nucleus through an ubiquitin-dependent mechanism, in which NEDD-4-mediated monoubiquitination of PTEN promotes its nuclear import. In the nucleus, PTEN physically associates with centromeres through CENP-C and maintains chromosome integrity. PTEN also acts on chromatin to coordinate with E2F-1 for the transcriptional regulation of Rad51, thus resulting in the control of DNA double-strand break repair.

(C124S) can still interact with CENP-C. Interestingly, a regulates cell cycle and apoptosis (Chung et al., 2005). few C-terminal PTEN mutants lacking association Moreover, cell cycle-dependentnuclear PTEN localiza- with centromeres still cause centromere instability, even tion and function can be regulated by its interaction though they retain phosphatase activity (Shen et al., with the major vault protein mediated by Ca2 þ signaling 2007). These data suggest that the physical association (Minaguchi et al., 2006). A recent study suggests that of nuclear PTEN with centromeres, rather than its oxidative stress can also regulate PTEN nuclear phosphatase activity, is crucial for its fundamental role localization in a manner independent of phosphatase in maintaining chromosomal integrity. activity but dependent upon phosphorylation of PTEN (Chang et al., 2008). Taken together, it appears that multiple mechanisms may direct PTEN nuclear localiza- Nuclear PTEN at the center of the stage tion and affect its nuclear function. Although nuclear transport of PTEN may be a The nuclear localization of PTEN has now obtained phosphatase-independent process, PTEN could still mechanistic support. Although PTEN lacks a typical need its phosphatase activity to exert multiple functions nuclear localization sequence (NLS), ubiquitination of in the nucleus. It is known that PTEN controls a DNA speciﬁc sites in its C-terminal region enhances transport repair process by regulating expression of Rad51 (Shen of PTEN into the nucleus (Trotman et al., 2007). et al., 2007). Consistently, lack of PTEN also impairs Consistent with our data, the nuclear import process CHK1 function and results in the accumulation of seems to be unrelated to the phosphatase activity of double-stranded DNA breaks (Puc and Parsons, 2005). PTEN because mutation of a critical ubiquitination site, Transcriptional modulation is obviously another novel K289, impairs nuclear importbutdoes notaffect function of PTEN in the nucleus, although the detailed catalytic activity (Trotman et al., 2007). Besides the mechanism remains to be elucidated. The function of ubiquitin-mediated mechanism, nuclear import of PTEN in transcriptional regulation is found to be PTEN may involve other mechanisms, such as passive phosphatase-dependent, because the phosphatase-deﬁ- transport by diffusion (Liu et al., 2005a) and active cient PTEN mutant (C124S) is unable to induce Rad51 transport through NLS-like signals (Chung et al., 2005). in PTEN-null cancer cells, whereas a catalytically active PTEN subcellular localization can exhibit a cell- C-terminal mutant can mimic full-length PTEN in cycle-dependent pattern (Liu et al., 2005b) and regulation of Rad51 (Shen and Yin, unpublished data). nuclear-cytoplasmic partitioning of PTEN differentially Although the C-terminal truncated PTEN mutant

Oncogene PTEN maintains genomic stability through multiple mechanisms Y Yin and WH Shen 5448 retains the ability to induce Rad51, it can no longer gene and its pathway can be directly targeted by BRCA1 associate with centromeres, causing chromosomal in- dysfunction, which may explain why basal-like breast stability. As illustrated in Figure 1, these results suggest cancer often develops due to BRCA1 deficiency. Given that different mechanisms may be responsible for the the critical roles of both PTEN and BRCA1 in distinct functions of nuclear PTEN: maintenance of maintaining chromosomal stability and proper DNA centromere stability (phosphatase-independent, through repair, successive hits on these important tumor CENP-C) and control of DNA repair (phosphatase- suppressors give rise to an increasing chance of tumor dependent, through Rad51). Although the PI3-kinase development and may interpret the aggressiveness and pathway may continue to serve as an important target genetically unstable feature of this specific type of for PTEN in the cytoplasm, identification of more breast cancer. Interestingly, the p53 gene, in addition nuclear-specific targets of PTEN will advance the field to BRCA1 and PTEN, is also frequently mutated in to a better understanding of PTEN function in tumor basal-like breastcancer (Borresen-Dale, 2003; Langerod suppression. Interestingly, major elements of the PTEN- et al., 2007), which further intrigues the need to exploit targeted PI3-kinase pathway in the cytoplasm, including how tumor suppressors coordinate with each other and the PI3-kinase itself and AKT, have now been found in form a complementary and cooperative network in the nucleus (Trotman et al., 2006). Whether or not its protecting against tumorigenesis. phosphatase activity is indispensable and whether the PI3-kinase/AKT serves as a target pathway, the important and exciting news is that ‘PTEN enters the Tumor suppressor network: p53, PTEN and beyond nuclear age’ (Baker, 2007). The nuclear PTEN and cytoplasmic PTEN may function complementarily p53 is the most frequently mutated gene in human in multiple pathways stabilizing the genome. cancers (Vogelstein et al., 2000; Levine et al., 2004). In Previous studies using cDNA microarray revealed terms of overall frequency, PTEN is the second most global changes in gene expression profiles associated commonly mutated gene in human cancers (Cantley and with alternate PTEN status (Hong et al., 2000; Neel, 1999; Simpson and Parsons, 2001). The mutation Matsushima-Nishiu et al., 2001), suggesting that PTEN spectrums of p53 and PTEN in human cancers, may be involved in gene transcription regulation. however, are different. Mutations of p53 occur at high Indeed, PTEN can negatively modulate the transcrip- frequencies in lung, colon and breastcancers, whereas tional activity of the androgen receptor (Nan et al., PTEN mutations are mostly found in glioblastoma, 2003), c-Met(Abounader et al., 2004), nuclear factor-kB endometrial cancer, malignant melanoma and prostate (Mayo et al., 2002; Moon et al., 2004), CRE binding cancer. There are germline p53 mutations in a familial protein (Mayo and Donner, 2002) and AP-1 (Koul syndrome of breastcancer, sarcomas and other et al., 2007). PTEN also downregulates the promoter neoplasms (Malkin et al., 1990). p53 is mutated in activity of MDM2 (Chang et al., 2004), insulin-like Li–Fraumeni syndrome, which is a familial cancer growth factor II (Kang-Park and Lee, 2003) and matrix susceptibility syndrome that develops disease at an early metalloproteinase-9 (Moon et al., 2004). The transcrip- onset(Bischoff et al., 1991; Strong et al., 1992; Yin et al., tion activity of hypoxia-inducible factor 1 is also 1992). Germline mutations of PTEN are associated with regulated by PTEN in the nucleus (Emerling et al., some inherited hamartoma-cancer syndromes (Eng and 2008). A directrole for PTEN in transcriptional Peacocke, 1998). Mice null for p53 develop normally regulation has been recognized just recently in a report and exhibit spontaneous tumors, mainly including showing that PTEN regulates the TFIIIB-centered basic lymphoma and sarcoma (Donehower et al., 1992). In transcription machinery by interrupting formation of contrast, homozygous deletion of Pten results in mouse functional TFIIIB complexes (Woiwode et al., 2008). embryonic lethality, suggesting that PTEN is necessary PTEN is physically associated with the promoter of for embryonic development(Di Cristofano et al., 1998; Rad51 and cooperates with the E2F-1 transcriptional Suzuki et al., 1998). Mice heterozygous for Pten develop factor in synergistic regulation of Rad51 (Shen et al., spontaneous tumors and conditional tissue-specific 2007). These data suggest that PTEN acts on chromatin disruption of Pten leads to different tumors in the and regulates transcription of important genes involved affected tissues (Di Cristofano and Pandolfi, 2000; in critical cellular processes such as cell cycle control and Stambolic et al., 2000; Kimura et al., 2003; Backman DNA repair. et al., 2004). Therefore, PTEN is a powerful and distinct As one of the most frequently mutated genes tumor suppressor. Regulation of p53 protein stability is associated with cancer, PTEN becomes the victim of a major mechanism to control levels of the p53 protein defective DNA repair due to the mutation of other (Shmueli and Oren, 2005). Mdm2 is a mediator for p53 DNA repair genes, such as BRCA1. Loss of BRCA1 ubiquitination, and MDM2 promotes the shuttle of p53 leads to recurrent gross mutation of PTEN (Saal et al., toward nuclear export, which leads to proteasome- 2008). Various structural mutations on chromosome mediated degradation of p53 (Shmueli and Oren, 2004). 10q23.31 atthe PTEN locus were frequently found in a p53 functions as a transcription factor that regulates very aggressive type of breast cancer, basal-like breast target genes (Levine, 1997). p53 controls cell- cancer. More importantly, gross mutations of PTEN in cycle progression, apoptosis and genomic stability this type of breast tumor are highly correlated with (Yonish-Rouach et al., 1991; Kastan et al., 1992; BRCA1 mutations. These results suggest that the PTEN Livingstone et al., 1992; Yin et al., 1992; Lowe et al.,

Oncogene PTEN maintains genomic stability through multiple mechanisms Y Yin and WH Shen 5449 1993; Tlsty et al., 1994). p53 is the first to be proposed as activation of the p53 pathway is consistent with a a guardian of the genome (Lane, 1992). PTEN functions cellular senescence phenotype upon Pten deletion, and as a phosphatase that inhibits PI-3 kinase/AKT signal- interestingly, combined deletion of p53 led to increased ing by converting PIP3 to PIP2. PTEN exerts tumor cell proliferation and escape from senescence (Chen suppressor function by regulation of proliferation and et al., 2005). Consistently, combined inactivation of Pten survival, cell migration, invasion and angiogenesis. As a and p53 greatly accelerated tumor development, which guardian of the genome, PTEN maintains genomic can be interpreted by deterioration of Pten deficiency- stability through multiple pathways. induced tumorigenesis due to lack of p53-dependent As an autonomous surveillance and defense system cellular senescence (Chen et al., 2005). These data against genomic instability and tumorigenesis, tumor supporta ‘one-by-one’ hitmodel for tumordevelopment suppressors act in coordination with each other to form due to sequential loss of major tumor suppressor genes. a regulatory network. The relationship between PTEN In agreement, upregulation of p53 and senescence-like and p53 has attracted extensive interest and enthusiasm. growth arrest in different human cells containing Comparison of their spectra of mutations revealed that inactivated PTEN were observed (Trotman et al., somatic mutations of PTEN and p53 are usually 2007). These data suggest a coordinate and complemen- independentand even mutually exclusive (Fujisawa tary pattern of interaction between PTEN and the p53 et al., 2000; Kato et al., 2000; Koul et al., 2002; Kurose pathway and would have therapeutic as well as et al., 2002), suggesting that these two guardians of the prognostic significance. genome evolved to guard distinct aspects of homeostasis Besides p53, PTEN may coordinate with another and cellular processes in a complementary mode. A tumor suppressor, Rb, in the prevention of tumorigen- similar complementary mode can also be implicated esis. Rb plays an important role in controlling cell cycle from their distinct patterns of expression and action. progression (Friend et al., 1986; Weinberg, 1992). PTEN The p53 protein is maintained at low levels in normal controls the cell cycle (Li and Sun, 1998; Ramaswamy cells and its induction requires stimulation upon cellular et al., 1999) and Rb has been reported to be essential for stress. Therefore, the sensitivity of p53 activation in PTEN-mediated cell cycle arrest and growth suppres- response to stress signals, rather than its steady sion (Paramio et al., 1999). There are two forms of Rb: expression levels, is crucial for its action against phosphorylated Rb, which is an inactive form, and tumorigenesis. In comparison, PTEN expression levels dephosphorylated Rb, which is activated. Rb is phos- are steady and high in normal cells and the maintenance phorylated and inactivated by cyclin D-dependent of its abundance is important to sustain its function to kinases CDK4 and CDK6. Rb can be activated through safeguard the genome. Besides PTEN mutation, loss or dephosphorylation by CDK inhibitors, such as p16 and reduction of PTEN expression are often found in human p18, thus retaining its growth-suppressive state (Guan cancers (Leslie and Downes, 2004), suggesting that et al., 1994; Medema et al., 1995; Quelle et al., 1995). PTEN function depends on its abundant expression and Interestingly, knockout of p18Ink4c in the Pten þ /À back- that insufficient PTEN expression may permit tumori- ground accelerates tumor development with a wider genesis. p53 action is dynamic and impulsive, whereas spectrum than Pten þ /À single deletion (Bai et al., 2006), PTEN functions in a static and continuous fashion. indicating a functional collaboration between these two PTEN was reported as a downstream transcriptional tumor suppressors. target of p53 in mediating apoptosis (Backman et al., Protection against tumorigenesis is maintained by an 2001). Other studies suggest that PTEN may act innate defense system comprised of a number of tumor upstream of p53 to regulate its expression levels and suppressors, each with special properties in controlling activity (Mayo and Donner, 2002; Freeman et al., 2003). one or more cellular processes through their specific PTEN can increase p53 stability and its DNA binding pathways. It is crucial that a network exists to integrate activity through physical association with p53 (Freeman signals and to coordinate molecular events in response et al., 2003). Furthermore, this cooperative interaction to environmental challenges. As p53 and PTEN are both between PTEN and p53 was applied to an animal model guardians of the genome, interaction and cooperation in which loss of one allele of Pten dramatically between their respective pathways are necessary for accelerated tumor onset in p53 þ /À mice (Freeman proper actions in protection of the genome. As p53 and et al., 2003). The physical association between PTEN PTEN are distinct in many ways as discussed above, and p53 was confirmed by other studies, which give they have different roles in guarding the genome. further evidence of their functional cross talk (Tang and Because PTEN expression is high in cells and tissues, Eng, 2006; Flores-Delgado et al., 2007). Recently, PTEN may be on duty during normal peaceful times, PTEN was found to physically associate with p300 in acting like a police force. However, p53 is mainly the nucleus, which maintains p53 in a highly acetylated responsible for dealing with emergency, such as DNA state in response to DNA damage (Bilbao et al., 2006). damage, as p53 levels are usually extremely low but All these data support a positive regulatory relationship highly increased following DNA damage and genotoxic between PTEN and p53. stress. Consistently, p53 mediates cell cycle checkpoints However, p53 and PTEN may have other types of and cell death, depending upon the type and nature of interactions. For example, deletion of Pten resulted in damage incurred. However, our preliminary studies enhanced expression of p53 as well as p21, a direct indicate that PTEN mainly functions in mitosis, which is transcription target of p53 (Xiong et al., 1993). The always active during the normal growth stage. There-

Oncogene PTEN maintains genomic stability through multiple mechanisms Y Yin and WH Shen 5450 fore, PTEN and p53 function in different aspects of life, contribute to the development of new therapeutic but together they protect cells against damage all the strategies for cancer treatment. time. Given that frequent mutations often give rise to functional deﬁciencies of important tumor suppressors, Acknowledgements a failsafe mechanism should exist among these tumor suppressors and their downstream effectors to maintain We are grateful to Drs C Walsh and HB Lieberman for their genomic integrity and to initiate protective responses. critical comments on the manuscript. This work was supported Understanding how they coordinate with each other by a grant from the National Institutes of Health (R01 will reveal the mechanism of tumor suppression and CA102447 to YY).

References

Abounader R, Reznik T, Colantuoni C, Martinez-Murillo F, Donehower LA, Harvey M, Slagle BL, McArthur MJ, Montgomery Jr CA, Rosen EM, Laterra J. (2004). Regulation of c-Met-dependent gene Butel JS et al. (1992). Mice deficientfor p53 are developmentally expression by PTEN. Oncogene 23: 9173–9182. normal but susceptible to spontaneous tumours. Nature 356: Agrawal S, Pilarski R, Eng C. (2005). Different splicing defects lead to 215–221. differential effects downstream of the lipid and protein phosphatase Emerling BM, Weinberg F, Liu JL, Mak TW, Chandel NS. (2008). activities of PTEN. Hum Mol Genet 14: 2459–2468. PTEN regulates p300-dependent hypoxia-inducible factor 1 tran- Backman SA, Ghazarian D, So K, Sanchez O, Wagner KU, scriptional activity through Forkhead transcription factor 3a Hennighausen L et al. (2004). Early onsetof neoplasia in the (FOXO3a). Proc Natl Acad Sci USA 105: 2622–2627. prostate and skin of mice with tissue-specific deletion of Pten. Proc Eng C. (2003). PTEN: one gene, many syndromes. Hum Mutat 22: Natl Acad Sci USA 101: 1725–1730. 183–198. Backman SA, Stambolic V, Suzuki A, Haight J, Elia A, Pretorius J Eng C, Peacocke M. (1998). PTEN and inherited hamartoma-cancer et al. (2001). Deletion of Pten in mouse brain causes seizures, ataxia syndromes. Nat Genet 19: 223. and defects in soma size resembling Lhermitte–Duclos disease. Fanning AS, Anderson JM. (1999). Protein modules as organizers of Nat Genet 29: 396–403. membrane structure. Curr Opin Cell Biol 11: 432–439. Bai F, Pei XH, Pandolfi PP, Xiong Y. (2006). p18 Ink4c and Pten Flores-Delgado G, Liu CW, Sposto R, Berndt N. (2007). A limited constrain a positive regulatory loop between cell growth and cell screen for protein interactions reveals new roles for protein cycle control. Mol Cell Biol 26: 4564–4576. phosphatase 1 in cell cycle control and apoptosis. J Proteome Res Baker SJ. (2007). PTEN enters the nuclear age. Cell 128: 25–28. 6: 1165–1175. Bilbao C, Rodriguez G, Ramirez R, Falcon O, Leon L, Chirino R Freeman DJ, Li AG, Wei G, Li HH, Kertesz N, Lesche R et al. (2003). et al. (2006). The relationship between microsatellite instability and PTEN tumor suppressor regulates p53 protein levels and activity PTEN gene mutations in endometrial cancer. Int J Cancer 119: through phosphatase-dependent and -independent mechanisms. 563–570. Cancer Cell 3: 117–130. Bischoff FZ, Strong LC, Yim SO, Pratt DR, Siciliano MJ, Giovanella BC FriendSH,BernardsR,RogeljS,WeinbergRA,RapaportJM,AlbertDM et al. (1991). Tumorigenic transformation of spontaneously et al. (1986). A human DNA segment with properties of the gene immortalized fibroblasts from patients with a familial cancer that predisposes to retinoblastoma and osteosarcoma. Nature 323: syndrome. Oncogene 6: 183–186. 643–646. Borresen-Dale AL. (2003). TP53 and breastcancer. Hum Mutat 21: Fujisawa H, Reis RM, Nakamura M, Colella S, Yonekawa Y, 292–300. Kleihues P et al. (2000). Loss of heterozygosity on chromosome 10 Cantley LC, Neel BG. (1999). New insights into tumor suppression: is more extensive in primary (de novo) than in secondary PTEN suppresses tumor formation by restraining the phosphoinosi- glioblastomas. Lab Invest 80: 65–72. tide 3-kinase/AKT pathway. Proc Natl Acad Sci USA 96: Georgescu MM, Kirsch KH, Akagi T, Shishido T, Hanafusa H. 4240–4245. (1999). The tumor-suppressor activity of PTEN is regulated Chang CJ, Freeman DJ, Wu H. (2004). PTEN regulates Mdm2 by its carboxyl-terminal region. Proc Natl Acad Sci USA 96: expression through the P1 promoter. J Biol Chem 279: 29841–29848. 10182–10187. Chang CJ, Mulholland DJ, Valamehr B, Mosessian S, Sellers WR, Wu Georgescu MM, Kirsch KH, Kaloudis P, Yang H, Pavletich NP, H. (2008). PTEN nuclear localization is regulated by oxidative stress Hanafusa H. (2000). Stabilization and productive positioning roles and mediates p53-dependent tumor suppression. Mol Cell Biol 28: of the C2 domain of PTEN tumor suppressor. Cancer Res 60: 3281–3289. 7033–7038. Chen Z, Trotman LC, Shaffer D, Lin HK, Dotan ZA, Niki M et al. Gildea JJ, Herlevsen M, Harding MA, Gulding KM, Moskaluk CA, (2005). Crucial role of p53-dependentcellular senescence in Frierson HF et al. (2004). PTEN can inhibit in vitro organotypic suppression of Pten-deficient tumorigenesis. Nature 436: 725–730. and in vivo orthotopic invasion of human bladder cancer cells Chung JH, Ginn-Pease ME, Eng C. (2005). Phosphatase and tensin even in the absence of its lipid phosphatase activity. Oncogene 23: homologue deleted on chromosome 10 (PTEN) has nuclear 6788–6797. localization signal-like sequences for nuclear import mediated by Gimm O, Perren A, Weng LP, Marsh DJ, Yeh JJ, Ziebold U et al. major vaultprotein. Cancer Res 65: 4108–4116. (2000). Differential nuclear and cytoplasmic expression of PTEN in Cully M, You H, Levine AJ, Mak TW. (2006). Beyond PTEN normal thyroid tissue, and benign and malignant epithelial thyroid mutations: the PI3K pathway as an integrator of multiple inputs tumors. Am J Pathol 156: 1693–1700. during tumorigenesis. Nat Rev Cancer 6: 184–192. Ginn-Pease ME, Eng C. (2003). Increased nuclear phosphatase and Das S, Dixon JE, Cho W. (2003). Membrane-binding and activation tensin homologue deleted on chromosome 10 is associated with G0- mechanism of PTEN. Proc Natl Acad Sci USA 100: 7491–7496. G1 in MCF-7 cells. Cancer Res 63: 282–286. Di Cristofano A, Pandolfi PP. (2000). The multiple roles of PTEN in Gu J, Tamura M, Yamada KM. (1998). Tumor suppressor PTEN tumor suppression. Cell 100: 387–390. inhibits integrin- and growth factor-mediated mitogen-activated Di Cristofano A, Pesce B, Cordon-Cardo C, Pandolfi PP. (1998). Pten protein (MAP) kinase signaling pathways. JCellBiol143: 1375–1383. is essential for embryonic development and tumour suppression. Guan KL, Jenkins CW, Li Y, Nichols MA, Wu X, O’Keefe CL et al. Nat Genet 19: 348–355. (1994). Growth suppression by p18, a p16INK4/MTS1- and

Oncogene PTEN maintains genomic stability through multiple mechanisms Y Yin and WH Shen 5451 p14INK4B/MTS2-related CDK6 inhibitor, correlates with wild- Li DM, Sun H. (1997). TEP1, encoded by a candidate tumor type pRb function. Genes Dev 8: 2939–2952. suppressor locus, is a novel protein tyrosine phosphatase regulated Hong TM, Yang PC, Peck K, Chen JJ, Yang SC, Chen YC et al. by transforming growth factor beta. Cancer Res 57: 2124–2129. (2000). Profiling the downstream genes of tumor suppressor PTEN Li DM, Sun H. (1998). PTEN/MMAC1/TEP1 suppresses the in lung cancer cells by complementary DNA microarray. Am J tumorigenicity and induces G1 cell cycle arrest in human Respir Cell Mol Biol 23: 355–363. glioblastoma cells. Proc Natl Acad Sci USA 95: 15406–15411. Iida S, Nakamura Y, Fujii H, Kimura M, Moriwaki K. (1998). Li J, Simpson L, Takahashi M, Miliaresis C, Myers MP, Tonks N A heterozygous germline mutation of the PTEN/MMAC1 gene in a et al. (1998). The PTEN/MMAC1 tumor suppressor induces cell patient with Cowden disease. Int J Mol Med 1: 565–568. death that is rescued by the AKT/protein kinase B oncogene. Iida S, Ono A, Sayama K, Hamaguchi T, Fujii H, Nakajima H et al. Cancer Res 58: 5667–5672. (2000). Accelerated decline of blood glucose after intravenous Li J, Yen C, Liaw D, Podsypanina K, Bose S, Wang SI et al. (1997). glucose injection in a patient with Cowden disease having a PTEN, a putative protein tyrosine phosphatase gene mutated in heterozygous germline mutation of the PTEN/MMAC1 gene. human brain, breast, and prostate cancer. Science 275: 1943–1947. Anticancer Res 20: 1901–1904. Liaw D, Marsh DJ, Li J, Dahia PL, Wang SI, Zheng Z et al. (1997). Kanaseki T, Torigoe T, Hirohashi Y, Hirai I, Himi T, Sato N. (2002). Germline mutations of the PTEN gene in Cowden disease, an Identification of germline mutation of PTEN gene and analysis of inherited breast and thyroid cancer syndrome. Nat Genet 16: 64–67. apoptosis resistance of the lymphocytes in a patient with Cowden Liliental J, Moon SY, Lesche R, Mamillapalli R, Li D, Zheng Y et al. disease. Pathobiology 70: 34–39. (2000). Genetic deletion of the Pten tumor suppressor gene Kang-Park S, Lee YI. (2003). PTEN modulates insulin-like growth promotes cell motility by activation of Rac1 and Cdc42 GTPases. factor II (IGF-II)-mediated signaling; the protein phosphatase Curr Biol 10: 401–404. activity of PTEN downregulates IGF-II expression in hepatoma Liu F, Wagner S, Campbell RB, Nickerson JA, Schiffer CA, Ross AH. cells. FEBS Lett 545: 203–208. (2005a). PTEN enters the nucleus by diffusion. J Cell Biochem 96: Kang-Park S, Lee YI, Lee YI. (2003). PTEN modulates insulin-like 221–234. growth factor II (IGF-II)-mediated signaling; the protein phospha- Liu JL, Sheng X, Hortobagyi ZK, Mao Z, Gallick GE, Yung WK. tase activity of PTEN downregulates IGF-II expression in (2005b). Nuclear PTEN-mediated growth suppression is indepen- hepatoma cells. FEBS Lett 545: 203–208. dentof Aktdown-regulation. Mol Cell Biol 25: 6211–6224. Kastan MB, Zhan Q, el-Deiry WS, Carrier F, Jacks T, Walsh WV Livingstone LR, White A, Sprouse J, Livanos E, Jacks T, Tlsty TD. et al. (1992). A mammalian cell cycle checkpoint pathway utilizing (1992). Altered cell cycle arrest and gene amplification potential p53 and GADD45 is defective in ataxia-telangiectasia. Cell 71: accompany loss of wild-type p53. Cell 70: 923–935. 587–597. Lowe SW, Schmitt EM, Smith SW, Osborne BA, Jacks T. (1993). p53 Kato H, Kato S, Kumabe T, Sonoda Y, Yoshimoto T, Kato S et al. is required for radiation-induced apoptosis in mouse thymocytes. (2000). Functional evaluation of p53 and PTEN gene mutations in Nature 362: 847–849. gliomas. Clin Cancer Res 6: 3937–3943. Maehama T, Dixon JE. (1998). The tumor suppressor, PTEN/ Kimura T, Suzuki A, Fujita Y, Yomogida K, Lomeli H, Asada N et al. MMAC1, dephosphorylates the lipid second messenger, phospha- (2003). Conditional loss of PTEN leads to testicular teratoma tidylinositol 3,4,5-trisphosphate. J Biol Chem 273: 13375–13378. and enhances embryonic germ cell production. Development 130: Mahimainathan L, Choudhury GG. (2004). Inactivation of platelet- 1691–1700. derived growth factor receptor by the tumor suppressor PTEN Koul A, Willen R, Bendahl PO, NilbertM, Borg A. (2002). Distinct provides a novel mechanism of action of the phosphatase. J Biol sets of gene alterations in endometrial carcinoma implicate alternate Chem 279: 15258–15268. modes of tumorigenesis. Cancer 94: 2369–2379. Maier D, Jones G, Li X, Schonthal AH, Gratzl O, Van Meir EG Koul D, Shen R, Shishodia S, Takada Y, BhatKP, Reddy SA et al. et al. (1999). The PTEN lipid phosphatase domain is not (2007). PTEN down regulates AP-1 and targets c-fos in human required to inhibit invasion of glioma cells. Cancer Res 59: glioma cells via PI3-kinase/Aktpathway. Mol Cell Biochem 300: 5479–5482. 77–87. Malkin D, Li FP, Strong LC, Fraumeni Jr JF, Nelson CE, Kim DH Kurose K, Gilley K, Matsumoto S, Watson PH, Zhou XP, Eng C. et al. (1990). Germ line p53 mutations in a familial syndrome of (2002). Frequent somatic mutations in PTEN and TP53 are breastcancer, sarcomas, and otherneoplasms. Science 250: mutually exclusive in the stroma of breast carcinomas. Nat Genet 1233–1238. 32: 355–357. Matsushima-Nishiu M, Unoki M, Ono K, Tsunoda T, Minaguchi T, Lachyankar MB, Sultana N, Schonhoff CM, Mitra P, Poluha W, Kuramoto H et al. (2001). Growth and gene expression profile LambertS et al. (2000). A role for nuclear PTEN in neuronal analyses of endometrial cancer cells expressing exogenous PTEN. differentiation. J Neurosci 20: 1404–1413. Cancer Res 61: 3741–3749. Lane DP. (1992). Cancer. p53, guardian of the genome. Nature 358: Mayo LD, Donner DB. (2002). The PTEN, Mdm2, p53 tumor 15–16. suppressor-oncoprotein network. Trends Biochem Sci 27: 462–467. Langerod A, Zhao H, Borgan O, Nesland JM, Bukholm IR, Ikdahl T Mayo MW, Madrid LV, Westerheide SD, Jones DR, Yuan XJ, et al. (2007). TP53 mutation status and gene expression profiles are Baldwin Jr AS et al. (2002). PTEN blocks tumor necrosis factor- powerful prognostic markers of breast cancer. Breast Cancer Res induced NF-kappa B-dependent transcription by inhibiting the 9: R30. transactivation potential of the p65 subunit. J Biol Chem 277: Lee JO, Yang H, Georgescu MM, Di Cristofano A, Maehama T, Shi Y 11116–11125. et al. (1999). Crystal structure of the PTEN tumor suppressor: McGarrity TJ, Wagner Baker MJ, Ruggiero FM, Thiboutot DM, implications for its phosphoinositide phosphatase activity and Hampel H, Zhou XP et al. (2003). GI polyposis and glycogenic membrane association. Cell 99: 323–334. acanthosis of the esophagus associated with PTEN mutation Leslie NR, Downes CP. (2004). PTEN function: how normal cells positive Cowden syndrome in the absence of cutaneous manifesta- control it and tumour cells lose it. Biochem J 382: 1–11. tions. Am J Gastroenterol 98: 1429–1434. Leslie NR, Yang X, Downes CP, Weijer CJ. (2007). PtdIns(3,4,5)P(3)- Medema RH, Herrera RE, Lam F, Weinberg RA. (1995). Growth dependent and -independent roles for PTEN in the control of cell suppression by p16ink4 requires functional retinoblastoma protein. migration. Curr Biol 17: 115–125. Proc Natl Acad Sci USA 92: 6289–6293. Levine AJ. (1997). p53, the cellular gatekeeper for growth and division. Minaguchi T, Waite KA, Eng C. (2006). Nuclear localization of PTEN Cell 88: 323–331. is regulated by Ca(2+) through a tyrosil phosphorylation-indepen- Levine AJ, Finlay CA, Hinds PW. (2004). P53 is a tumor suppressor dent conformational modification in major vault protein. Cancer gene. Cell 116: S67–S69, 61 p following S69. Res 66: 11677–11682.

Oncogene PTEN maintains genomic stability through multiple mechanisms Y Yin and WH Shen 5452 Moon SK, Kim HM, Kim CH. (2004). PTEN induces G1 cell cycle Shmueli A, Oren M. (2005). Life, death, and ubiquitin: taming the arrest and inhibits MMP-9 expression via the regulation of mule. Cell 121: 963–965. NF-kappaB and AP-1 in vascular smooth muscle cells. Arch Simpson L, Parsons R. (2001). PTEN: life as a tumor suppressor. Biochem Biophys 421: 267–276. Exp Cell Res 264: 29–41. Myers MP, Pass I, Batty IH, Van der Kaay J, Stolarov JP, Hemmings BA Stambolic V, Suzuki A, de la Pompa JL, Brothers GM, et al. (1998). The lipid phosphatase activity of PTEN is critical Mirtsos C, Sasaki T et al. (1998). Negative regulation of PKB/ for its tumor supressor function. Proc Natl Acad Sci USA 95: Akt-dependent cell survival by the tumor suppressor PTEN. Cell 95: 13513–13518. 29–39. Myers MP, Stolarov JP, Eng C, Li J, Wang SI, Wigler MH et al. Stambolic V, Tsao MS, Macpherson D, Suzuki A, Chapman WB, (1997). P-TEN, the tumor suppressor from human chromosome Mak TW. (2000). High incidence of breastand endometrial 10q23, is a dual-specificity phosphatase. Proc Natl Acad Sci USA neoplasia resembling human Cowden syndrome in pten+/À mice. 94: 9052–9057. Cancer Res 60: 3605–3611. Nan B, Snabboon T, Unni E, Yuan XJ, Whang YE, Marcelli M. Steck PA, Pershouse MA, Jasser SA, Yung WK, Lin H, Ligon AH (2003). The PTEN tumor suppressor is a negative modulator of et al. (1997). Identification of a candidate tumour suppressor gene, androgen receptor transcriptional activity. J Mol Endocrinol 31: MMAC1, at chromosome 10q23.3 that is mutated in multiple 169–183. advanced cancers. Nat Genet 15: 356–362. Okumura K, Zhao M, DePinho RA, Furnari FB, Cavenee WK. Strong LC, Williams WR, Tainsky MA. (1992). The Li–Fraumeni (2005). PTEN: a novel anti-oncogenic function independent of syndrome: from clinical epidemiology to molecular genetics. Am J phosphatase activity. Cell Cycle 4: 540–542. Epidemiol 135: 190–199. Paramio JM, Navarro M, Segrelles C, Gomez-Casero E, Jorcano JL. Sulis ML, Parsons R. (2003). PTEN: from pathology to biology. (1999). PTEN tumour suppressor is linked to the cell cycle control Trends Cell Biol 13: 478–483. through the retinoblastoma protein. Oncogene 18: 7462–7468. Sun H, Lesche R, Li DM, Liliental J, Zhang H, Gao J et al. (1999). Park MJ, Kim MS, Park IC, Kang HS, Yoo H, Park SH et al. (2002). PTEN modulates cell cycle progression and cell survival by PTEN suppresses hyaluronic acid-induced matrix metalloprotei- regulating phosphatidylinositol 3,4,5,-trisphosphate and Akt/ nase-9 expression in U87 MG glioblastoma cells through focal protein kinase B signaling pathway. Proc Natl Acad Sci USA 96: adhesion kinase dephosphorylation. Cancer Res 62: 6318–6322. 6199–6204. Parsons R. (2004). Human cancer, PTEN and the PI-3 kinase Suzuki A, de la Pompa JL, Stambolic V, Elia AJ, Sasaki T, del Barco pathway. Semin Cell Dev Biol 15: 171–176. Barrantes I et al. (1998). High cancer susceptibility and embryonic Perren A, Komminoth P, Saremaslani P, Matter C, Feurer S, Lees JA lethality associated with mutation of the PTEN tumor suppressor et al. (2000). Mutation and expression analyses reveal differential gene in mice. Curr Biol 8: 1169–1178. subcellular compartmentalization of PTEN in endocrine pancreatic Tamura M, Gu J, Matsumoto K, Aota S, Parsons R, Yamada KM. tumors compared to normal islet cells. Am J Pathol 157: 1097–1103. (1998). Inhibition of cell migration, spreading, and focal adhesions PodsypaninaK,EllensonLH,NemesA,GuJ,TamuraM,YamadaKM by tumor suppressor PTEN. Science 280: 1614–1617. et al. (1999). Mutation of Pten/Mmac1 in mice causes neoplasia in Tang Y, Eng C. (2006). PTEN autoregulates its expression by multiple organ systems. Proc Natl Acad Sci USA 96: 1563–1568. stabilization of p53 in a phosphatase-independent manner. Cancer Puc J, Keniry M, Li HS, Pandita TK, Choudhury AD, Memeo L et al. Res 66: 736–742. (2005). Lack of PTEN sequesters CHK1 and initiates genetic Tlsty TD, White A, Livanos E, Sage M, Roelofs H, Briot A et al. instability. Cancer Cell 7: 193–204. (1994). Genomic integrity and the genetics of cancer. Cold Spring Puc J, Parsons R. (2005). PTEN loss inhibits CHK1 to cause double Harb Symp Quant Biol 59: 265–275. stranded-DNA breaks in cells. Cell Cycle 4: 927–929. Trotman LC, Alimonti A, Scaglioni PP, Koutcher JA, Cordon-Cardo Quelle DE, Ashmun RA, Hannon GJ, Rehberger PA, Trono D, C, Pandolfi PP. (2006). Identification of a tumour suppressor Richter KH et al. (1995). Cloning and characterization of murine network opposing nuclear Akt function. Nature 441: 523–527. p16INK4a and p15INK4b genes. Oncogene 11: 635–645. Trotman LC, Wang X, Alimonti A, Chen Z, Teruya-Feldstein J, Yang Raftopoulou M, Etienne-Manneville S, Self A, Nicholls S, Hall A. H et al. (2007). Ubiquitination regulates PTEN nuclear import and (2004). Regulation of cell migration by the C2 domain of the tumor tumor suppression. Cell 128: 141–156. suppressor PTEN. Science 303: 1179–1181. Vivanco I, Palaskas N, Tran C, Finn SP, Getz G, Kennedy NJ et al. Ramaswamy S, Nakamura N, Vazquez F, Batt DB, Perera S, (2007). Identification of the JNK signaling pathway as a Roberts TM et al. (1999). Regulation of G1 progression by the functional target of the tumor suppressor PTEN. Cancer Cell 11: PTEN tumor suppressor protein is linked to inhibition of the 555–569. phosphatidylinositol 3-kinase/Akt pathway. Proc Natl Acad Sci Vogelstein B, Lane D, Levine AJ. (2000). Surfing the p53 network. USA 96: 2110–2115. Nature 408: 307–310. Saal LH, Gruvberger-Saal SK, Persson C, Lovgren K, Jumppanen M, Waite KA, Eng C. (2002). Protean PTEN: form and function. Am J Staaf J et al. (2008). Recurrent gross mutations of the PTEN tumor Hum Genet 70: 829–844. suppressor gene in breastcancers withdeficientDSB repair. Nat Wang SI, Puc J, Li J, Bruce JN, Cairns P, Sidransky D et al. (1997). Genet 40: 102–107. Somatic mutations of PTEN in glioblastoma multiforme. Cancer Sano T, Lin H, Chen X, Langford LA, Koul D, Bondy ML et al. Res 57: 4183–4186. (1999). Differential expression of MMAC/PTEN in glioblastoma Wang X, Trotman LC, Koppie T, Alimonti A, Chen Z, Gao Z et al. multiforme: relationship to localization and prognosis. Cancer Res (2007). NEDD4-1 is a proto-oncogenic ubiquitin ligase for PTEN. 59: 1820–1824. Cell 128: 129–139. Sansal I, Sellers WR. (2004). The biology and clinical relevance of the Weinberg RA. (1992). The retinoblastoma gene and gene product. PTEN tumor suppressor pathway. J Clin Oncol 22: 2954–2963. Cancer Surv 12: 43–57. Shen WH, Balajee AS, Wang J, Wu H, Eng C, Pandolfi PP et al. Weng L, Brown J, Eng C. (2001a). PTEN induces apoptosis and cell (2007). Essential role for nuclear PTEN in maintaining chromoso- cycle arrest through phosphoinositol-3-kinase/Akt-dependent and mal integrity. Cell 128: 157–170. -independentpathways. Hum Mol Genet 10: 237–242. Shin KH, Kim JM, Rho KS, Park KH, Oh JE, Min BM. (2002). Weng LP, Brown JL, Baker KM, Ostrowski MC, Eng C. (2002). Inactivation of the PTEN gene by mutation, exonic deletion, PTEN blocks insulin-mediated ETS-2 phosphorylation through and loss of transcript in human oral squamous cell carcinomas. MAP kinase, independently of the phosphoinositide 3-kinase Int J Oncol 21: 997–1001. pathway. Hum Mol Genet 11: 1687–1696. Shmueli A, Oren M. (2004). Regulation of p53 by Mdm2: fate is in the Weng LP, Smith WM, Brown JL, Eng C. (2001b). PTEN inhibits numbers. Mol Cell 13: 4–5. insulin-stimulated MEK/MAPK activation and cell growth by

Oncogene PTEN maintains genomic stability through multiple mechanisms Y Yin and WH Shen 5453 blocking IRS-1 phosphorylation and IRS-1/Grb-2/Sos complex Xiong Y, Hannon GJ, Zhang H, Casso D, Kobayashi R, Beach D. formation in a breast cancer model. Hum Mol Genet 10: 605–616. (1993). p21 is a universal inhibitor of cyclin kinases. Nature 366: Whiteman DC, Zhou XP, Cummings MC, Pavey S, Hayward NK, 701–704. Eng C. (2002). Nuclear PTEN expression and clinicopathologic Yin Y, Tainsky MA, Bischoff FZ, Strong LC, Wahl GM. (1992). features in a population-based series of primary cutaneous Wild-type p53 restores cell cycle control and inhibits gene melanoma. Int J Cancer 99: 63–67. ampliﬁcation in cells with mutant p53 alleles. Cell 70: 937–948. Woiwode A, Johnson SA, Zhong S, Zhang C, Roeder RG, Teichmann Yonish-Rouach E, Resnitzky D, Lotem J, Sachs L, Kimchi A, M et al. (2008). PTEN represses RNA polymerase III-dependent Oren M. (1991). Wild-type p53 induces apoptosis of myeloid transcription by targeting the TFIIIB complex. Mol Cell Biol 28: leukaemic cells that is inhibited by interleukin-6. Nature 352: 4204–4214. 345–347.

Supplementary Information accompanies the paper on the Oncogene website (http://www.nature.com/onc)

Oncogene