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Endocrine-Related (2000) 7 115–129

PTEN, a unique tumor suppressor

P L M Dahia

Department of Cancer Biology, Dana-Farber Cancer Institute, 44 Binney Street SM1010, Harvard Medical School, Boston, Massachusetts 02215-6084, USA (Requests for offprints should be addressed toPLMDahia; Email: Patricia [email protected])

Introduction al. 1999) with many common features and some phenotypic disparities which have, for the most part, confirmed the For many years, it has been thought that the previous in vitro data available. In addition, four knockouts region 10q22-24 includes one or more that appear to of the PTEN homolog in Caenorhabditis elegans have helped play a role in several human malignancies. PTEN is a new understand, in more detail, the pathways involved in PTEN encoding a dual-specificity downstream signaling (Ogg & Ruvkun 1998, Gil et al. 1999, that was cloned simultaneously by three groups Mihaylova et al. 1999, Rouault et al. 1999), and have set the (Li & Sun 1997, Li et al. 1997, Steck et al. 1997), two of stage for additional studies that will shed light on yet which used a positional cloning approach to identify genes unknown aspects of its regulation. Thus, genetic, biochemical in (Li et al. 1997, Steck et al. 1997). While evidence as well as in vivo disruption have implicated PTEN several protein kinases have been implicated as , in the regulation of multiple cellular functions: growth, and have long been known frequently to extracellular matrix interactions, and devel- antagonize their function, there has been no direct opment interactions. More recently, the crystal structure of demonstration of the role of phosphatases in tumor PTEN has been solved, providing further clues to PTEN development (Myers & Tonks 1997). PTEN characterization substrate specificity, and also helping in the understanding of as a bona fide tumor suppressor gene has confirmed that a some of the biochemical peculiarities of this tumor deficient phosphatase activity can lead to cancer, as detailed suppressor phosphatase. by studies that are described below. All this wealth of data considered, PTEN has had a Since the cloning of PTEN, a considerable number of remarkable appeal to a broad range of research areas and can papers describing its mutational spectrum in a variety of certainly be adopted as an excellent paradigm for the fast human tumors (Cairns et al. 1997, 1998, Dahia et al. 1997, pace of that we are presently witnessing. An Liu et al. 1997, Rasheed et al. 1997, Risinger et al. 1997, update on the current knowledge of the genetic and Tashiro et al. 1997, Teng et al. 1997, Tsou et al. 1997, Wang functional aspects of PTEN highlighted above are the scope et al. 1997), as well as its identification as the susceptibility of this review and will be discussed in more detail in the gene to cancer- syndromes (Liaw et al. 1997, next sections. Marsh et al. 1997a) have been reported in the literature. These studies were followed closely by the initial reports unraveling PTEN’s function in cell signaling. Within a short PTEN cloning, structure and period of time, it has been demonstrated that PTEN indeed functions as a phosphatase (Myers et al. 1997), as initial analysis structural comparisons indicated. However, lipid residues are PTEN’s major, functionally proven substrates (Maehama & Somatic of PTEN Dixon 1999). Tyrosine- or serine/threonine-proteins that The chromosome 10q23 region has long been found to function as targets of PTEN have been more difficult to represent an important target area in several human tumors, establish, although PTEN has been shown to bind to focal such as , prostate and , endo- adhesion kinase (FAK) and to reduce its tyrosine metrial and hematological malignancies (Pfeifer (Tamura et al. 1998). It has also been found et al. 1995, Rasheed et al. 1995, Albarosa et al. 1996, Trybus that PTEN induces growth suppression via arrest et al. 1996, Kobayashi et al. 1997, Bose et al. 1998). It has and/or induction of (Furnari et al. 1998, Myers et also been suspected to be the site for loci associated with al. 1998, Stambolic et al. 1998, Ramaswamy et al. 1999, Sun certain inherited disorders such as hamartomatous syndromes et al. 1999), and inhibits and migration. Three (Nelen et al. 1996, Jacoby et al. 1997, Zigman et al. 1997). PTEN mammalian knockout models have been generated (Di This group of diseases includes (CS), an Cristofano et al. 1998, Stambolic et al. 1998, Podsypanina et autosomal dominant disorder which comprises multiple

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in tissues derived from the three major 10q23 region (Jacoby et al. 1997). JPC is an autosomal embryological layers, and is associated with a higher risk of dominant condition with incomplete penetrance characterized breast and follicular or papillary cancer (25–50% of by hamartomatous polyps of the gastrointestinal tract and affected females, and 10% of affected individuals predisposition to gastrointestinal cancer. respectively), and in a subset of cases, a dysplatic Initial linkage analysis studies have localized CS and ganglyocytoma of the cerebellum, Lhermitte-Duclos disease, JPC to 10q23 (Jacoby et al. 1997, Nelen et al. 1997). In might also develop. Another hamartoma syndrome linked to particular, by studying families with CS with recombination the same chromosomal region is Bannayan-Riley-Ruvalcaba of this area, a critical CS region was delineated which syndrome (BRR). BRR also has an autosomal dominant trait spanned a 1.5 cm area at 10q23.3 (Liaw et al. 1997). and is clinically characterized by , lipomatosis PTEN was identified as a tumor-suppressor gene located and pigmented macules of the glans penis, Hashimoto’s within the CS critical region 10q23.3 and was found to be thyroiditis, and mental and developmental delay (Eng 1997). deleted in a wide variety of human and cell lines, In contrast to CS, an increased risk of malignancy has not including glioblastomas, prostate, breast and kidney cancer previously been documented in a formal manner in BRR. cell lines (Li et al. 1997). This gene has also been designated Besides these two highly related syndromes, other inherited as MMAC1 (for mutated in multiple advanced cancer) due to disorders such as juvenile polyposis coli (JPC) have been its initial apparent association with later-stage tumors (Steck suspected to be linked, in at least a subset of cases, to the et al. 1997) and TEP1 (transforming -β

Figure 1 Frequency of somatic PTEN abnormalities in various tumor types: the mean (±S.D.) results from different reports have been calculated and are depicted as columns (original studies are cited throughout the text). When available, PTEN abnormalities other than mutations, such as transcriptional silencing or low/absent protein expression, are also shown (*; available in only two tumor types, hematological malignancies and breast cancer). Hem. malign, hematological malignancies.

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(TGFβ)-regulated and epithelial-cell-enriched phosphatase) pathways, each of which requires the acquisition of distinct for its putative signaling effectors (Li & Sun 1997). The molecular defects, and not in a sequential manner, as PTEN gene contains nine exons and encodes a 403 amino previously believed. acid protein that displays high homology in its N-terminal Occasionally, highly divergent mutation frequencies region to dual specificity protein phosphatases and also to have been reported for a given tissue type: this may be tensin, a cytoskeleton protein, and to , a protein explained in some cases by the existence of a PTEN involved in synaptic vesicle transport (Li & Sun 1997, Li et , PTENP1, also known as psiPTEN, PTH2, al. 1997). The initial suggestion that PTEN functions as a PTEN2 (Teng et al. 1997, Dahia et al. 1998, Whang et al. tumor suppressor gene came from studies in which truncating 1998a). This pseudogene carries over 98% homology with or inactivating mutations of PTEN accompanied by deletions PTEN cDNA, and can confound interpretation of the of the wild-type were described with high frequency in mutation data, especially in studies based on analysis of malignant and (Cairns et al. cDNA-based templates. 1997, Liu et al. 1997, Rasheed et al. 1997, Risinger et al. 1997, Wang et al. 1997), with moderate frequency in prostate PTEN mutations cancer (Cairns et al. 1997, Liu et al. 1997, Rasheed et al. 1997, Risinger et al. 1997, Wang et al. 1997) and with a PTEN’s role, however, is not limited to somatic disorders: lower rate in other malignancies such as breast, thyroid, germline mutations of PTEN are the cause of CS and BRR bladder, ovary, small-cell , and hematological cases (Liaw et al. 1997, Marsh et al. 1997a). PTEN malignancies (Dahia et al. 1997, Cairns et al. 1998, mutations have been identified in 13–81% and 57–60% of Gronbaek et al. 1998, Kohno et al. 1998, Ueda et al. 1998, cases of CS and BRR respectively (Lynch et al. 1997, Nelen Yokomizo et al. 1998). Confirmation of the tumor suppressor et al. 1997, Longy et al. 1998, Marsh et al. 1998, 1999, Tsou function of PTEN was provided by studies showing that et al. 1998). PTEN mutations in CS tend to cluster at, but ectopic expression of wild-type, but not mutant PTEN, are not limited to, exon 5, which encodes the phosphatase induces growth suppression of cell lines carrying a mutant signature motif (see Fig. 2). Not only do the two syndromes copy of PTEN (Furnari et al. 1997, Li et al. 1998, Myers et share many clinical features, but the mutations in some al. 1998) (see more on PTEN function below). A large body families are identical, suggesting that these two disorders are of studies is now available reporting PTEN mutation analysis allelic. Recent studies have attempted to establish genotype– in a variety of different human tumors and results from many phenotype correlations in CS and BRR (Marsh et al. 1998a, of such studies are summarized in Fig. 1. Further, a 1999). Genotype-phenotype analyses in CS alone suggest comprehensive review of PTEN mutation spectra in several that the presence of a PTEN mutation may be associated with human tumors has recently been published (Ali et al. 1999). the development of breast (Marsh et al. 1998a). The initial suggestion that PTEN is mutated in later stages or Within the BRR group, an association has been noted more advanced tumor types, such as that seen in between the presence of lipomas and the detection of PTEN glioblastomas, did not stand extensive analysis that included mutation (Marsh et al. 1999). It is possible that the presence other tumor types. It is now known that PTEN abnormalities of a PTEN mutation, whether it be in CS, BRR or BRR/ represent, in fact, an early event in endometrial cancer, in CS families, predisposes to tumor development. However, it which PTEN mutations can be detected as early as in appears from these initial observations that mutation-positive pre-neoplastic lesions (Maxwell et al. 1998, Obata et al. status in BRR predisposes to benign tumors, compared with 1998). In addition, studies attempting to associate PTEN malignant tumors in CS or BRR/CS overlap families. Among mutations with prognostic indicators in have CS, BRR and in BRR/CS overlap families that are PTEN not revealed a clear correlation (McMenamin et al. 1999). mutation-positive, the mutation spectrum appears similar. Surprisingly, two of the most common malignancies Thus, PTEN mutation-positive CS and BRR are possibly associated with CS (see below), breast and non-medullary different presentations of a single syndrome and, hence, both thyroid cancer, have been found to have a low frequency of should receive equal attention with respect to cancer PTEN mutations (Dahia et al. 1997, Chen et al. 1998, surveillance (Marsh et al. 1999). It is possible that other Halachmi et al. 1998, Ueda et al. 1998, Feilotter et al. 1999). modulating factors might be responsible for the observed In particular, non-CS and non-breast cancer associated gene clinical variation between CS and BRR. However, (BRCA)1 or -BRCA2 associated familial breast cancer approximately 20% of CS and 40% of BRR families still syndromes do not have PTEN mutations (FitzGerald et al. remain without an identifiable germline PTEN mutation. 1998, Marsh et al. 1998c). Of interest, we found that thyroid Individuals from these PTEN mutation-negative families carry higher rates of in the cannot at present rely on a diagnostic test for the disease. area spanning the PTEN than malignant thyroid tumors Informative PTEN mutation-negative families have been (Dahia et al. 1997). This finding suggests that benign and shown to be linked to the 10q23 region, where PTEN lies malignant thyroid neoplasms arise through two independent (Nelen et al. 1996, Liaw et al. 1997). While mutations in the

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Figure 2 Distribution of germline PTEN mutations in 30 families with Cowden syndrome, according to mutation type (see key) and localization within the PTEN gene. (Adapted from Marsh et al. 1998a) with permission.

promoter region of PTEN may account for some of these Other mechanisms of PTEN inactivation cases, it is possible that another gene located in this area In addition to structural abnormalities, an increasing amount might be responsible for these PTEN mutation-negative of evidence has now suggested that PTEN may be inactivated cases. A recently identified gene mapping centromeric of by mechanisms other than mutations and/or deletions PTEN in 10q23 (Caffrey et al. 1999, Chi et al. 1999), which (Whang et al. 1998b, Dahia et al. 1999). Epigenetic encodes a histidine phosphatase with the ability to remove mechanisms might be responsible for a number of cases in the 3-phosphate from moieties, was examined for which PTEN expression is downregulated or even totally mutations in PTEN mutation-negative CS and BRR families ablated in the absence of a detectable mutation, as seen in (Dahia et al. 2000). We found no mutations in any of the individuals examined. Therefore, the major genetic defect prostate cancer (Whang et al. 1998b) as well as in our responsible for CS and BRR in cases without detectable observation of a series of and lymphoma cell lines PTEN mutation remains to be established. There has been a (Dahia et al. 1999). In agreement with these findings and in report describing genetic heterogeneity in CS (Tsou et al. favor of the existence of other mechanisms of PTEN 1997b). No candidate gene has emerged to account for these inactivation is the recent report that lack of PTEN expression cases to date. in breast cancer sections is not associated with deletions in a Besides CS and BRR, a definite role for PTEN in other subgroup of samples (Perren et al. 1999). The precise inherited hamartomatous syndromes has not been clearly mechanisms involved in silencing PTEN expression in such confirmed. While an initial report described the detection of malignancies are not known. It has been suggested that PTEN germline PTEN mutations in three cases of juvenile polyposis methylation might occur in a subset of prostate cancers syndrome (JPS) (Olschwang et al. 1998), the study of a (Whang et al. 1998b) but not in another series of prostate larger series of affected families selected by more stringent tumors (Cairns et al. 1997) nor in hematological diagnostic criteria for JPS excluded linkage to an area malignancies (Dahia et al. 1999). spanning 29cM surrounding the PTEN locus (Marsh et al. Our preliminary in vitro studies have suggested that 1997b). Therefore, a definite role for PTEN in JPS -mediated degradation is not enhanced in cell lines pathogenesis still needs to be established. with low or absent PTEN expression in co-existence with

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normal transcript levels and structural defects (P L M Dahia phenotype (discussed below). These results are more in and C Eng, unpublished observations). It remains to be keeping with the findings that heterozygous animals already established whether PTEN degradation is in fact mediated by display hyperplastic and dysplastic phenotypes and with the the ubiquitin-proteasome pathway in vivo and whether any observations that a number of CS tumors do not have component of this cascade is altered under particular detectable LOH (Marsh et al. 1998). This may reflect the fact circumstances in cells with low or absent PTEN protein that a much broader range of tumors lacking a single copy without structural or transcriptional defects. of the gene may suffer from abnormal PTEN function than Other mechanisms involving reduction of PTEN protein was previously anticipated. Table 1 depicts results from expression are still elusive. Analysis of structural motifs of current research that favor or are against a PTEN dosage PTEN might help unravel such mechanisms. It has recently effect. To reconcile these apparent conflicting findings, it been suggested that PTEN’s half-life may be reduced in may eventually be proven that PTEN haploinsufficiency may certain mutated cell lines (Georgescu et al. 1999). PTEN has involve a tissue-dependent or developmentally regulated two PEST homology domains and a PSD-95/Dig/20–1 phenomenon. (PDZ)-binding motif on its C-terminal region. While the Table 1 Features in favor or against a PTEN PEST homology motifs are supposed to assist in protein haploinsufficiency phenotype based on current data. folding, it is expected that their absence would interfere with protein targeting for degradation. The PDZ-binding motif is In favor Against responsible for protein–protein interactions. Studies aimed at CS hamartomas without LOH LOH of hamartomas in CS mapping the functional activity of PTEN constructs lacking Heterozygous mice with LOH seen in somatic tumors these motifs revealed that, not unexpectedly, absence of lymphoid tissue proliferation with PTEN mutations PEST domains results in faster rates of PTEN degradation (an auto-immune like (, endometrial syndrome) carcinoma, prostate (Georgescu et al. 1999). However, no changes in PTEN carcinoma) degradation rates were seen in PDZ-lacking PTEN constructs Heterozygous knockout mice LOH in the majority of tested as compared with the wild-type protein. The tumor develop tumors lymphomas of Pten+/- suppressor function of the mutants lacking either PEST or knockout mice PDZ homology motifs was not altered in the study referred to. More recent work has shown that PTEN associates with a member of a family of membrane-associated scaffold proteins PTEN function known as MAGUKs (membrane-associated guanylate In general terms, PTEN has been shown to modulate, mostly kinases), MAGI-2 (also known as AIP), through PTEN’s PDZ as an inhibitor factor, two seemingly distinct cellular domain, and that this interaction enhances PTEN efficiency in functions: /survival and cell migration/adhesion reducing AKT activity (Wu et al. 2000). It is proposed that through its lipid and activities MAGUKs function as scaffold proteins to assemble respectively. These aspects of PTEN function will be multiprotein signaling complexes and enhance their stability, discussed in detail below. thereby increasing the efficiency of . While many aspects of PTEN structure–function PTEN signaling: substrates and pathways regulation have been unraveled, it still remains to be established how the stability of PTEN protein is regulated in A recent study demonstrated that PTEN acts as a certain tumors without PTEN mutations but with low or phospholipid phosphatase dephosphorylating the phospha-

absent protein levels. tidylinositol 3,4,5 triphosphate (PtdIns(3,4,5)P3 ) and phos-

Initial observations that hamartomatous tissues from phatidylinositol 3,4 biphosphate (PtdIns(3,4)P2) specifically Cowden patients had high frequencies of loss of at the D3 position of the inositol ring, resulting in heterozygosity (LOH) suggested that homozygous PTEN PtdIns(4,5)P and PtdIns(4)P respectively. These D3 phos- inactivation was required for a tumor suppressor phenotype pholipids are the direct products of the (Marsh et al. 1998b). This was in agreement with findings 3-OH kinase (PI-3 kinase, PI3K), which counteracts PTEN that Pten+/- knockout mice developed lymphomas at a high function (Fig. 3). PI3-kinase triggers signaling through rate where LOH was found at the PTEN locus (Suzuki et al. multiple pathways, many of which are thought to associate 1998). A recent study has now suggested that PTEN has a with cell growth and survival (Liscovitch & Cantley 1994, dosage effect and haploinsufficiency may, in fact, be Vanhaesebroeck et al. 1997). PTEN, working in opposition responsible for impaired PTEN activity (Di Cristofano et al. to PI3-kinase, is therefore associated with cell death or arrest +/- 1999). In this study, heterozygote mutant Pten mice signals. Phospholipid residues such as PtdIns(3,4,5)P3 are showed an abnormal apoptotic response and an autoimmune present in cells upon stimulation by several growth factors,

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Figure 3 PTEN signaling pathway. The diagram summarizes the intracellular signal triggered by growth factors. PI3K is activated by growth factor stimulation. This is followed by phosphorylation of inositol

substrates at position 3, to produce phosphatidyl inositol 3,4,5, triphosphate (PtdIns(3,4,5)P3), which in turn activates serine-threonine AKT and AKT kinases, PDK1 and possibly ILK. Phosphorylated AKT (P-AKT), the active form of this kinase, promotes cell survival through multiple pathways (see text for details). PTEN

antagonizes PI3K, dephosphorylating PtdIns(3,4,5)P3 at position 3. Under these circumstances, AKT is not activated, and cell death signals predominate.

such as platelet-derived growth factor (PDGF), insulin-like AKT pathway and therefore counteracts cell survival growth factor (IGF), and epidermal growth factor (EGF). mechanisms elicited by this signaling. Upon activation by growth factor, proteins containing a The mechanisms of cell survival associated with AKT pleckstrin-homology (PH) domain are recruited to the appear to involve multiple pathways, including growth membrane 3 where they associate with phospholipids. One factors, cytokines, c- overexpression, UV irradiation, and of the PH domain-containing proteins relevant in this matrix detachment (Coffer et al. 1998). One of the known pathway is the serine-threonine kinase, AKT, also known as signals activated by AKT is its phosphorylation of the Bcl-2 PKB or RAC1 (reviewed in Coffer et al. 1998). AKT, in family member, BAD: phosphorylation of BAD results in turn, and as a consequence of lipid binding, alters its suppression of apoptosis (Datta et al. 1997). conformation to allow two of its residues, threonine 308 and AKT has also been reported to counteract the apoptotic serine 473, to be phosphorylated and therefore become response of several cellular factors. Recently, the active. The kinase responsible for phosphorylation of factor NF-kappaB has been implicated in the apoptotic threonine 308 is phosphonositide-dependent kinase 1 response antagonized by the PI3K/AKT pathway (PDK1), an which also contains a PH domain and is (Romashkova & Makarov 1999). In addition, AKT suppresses therefore dependent on lipid binding for its full activity CD-95/Fas-induced apoptosis (Rohn et al. 1998). Another (Alessi et al. 1997). There is some preliminary evidence, possible mechanism by which AKT exerts this function is predominantly from in vitro studies, that a second through the regulation of the activity of FKHRL1, a member lipid-dependent, PH domain-containing enzyme, ILK of the Forkhead family of transcription factors (Brunet et al. (integrin-linked kinase), is responsible for phosphorylation of 1999). In the presence of survival factors, AKT phosphorylates the serine 473 (Delcommenne et al. 1998). Further, a recent FKHRL1, leading to FKHRL1’s retention in the cytoplasm. paper has proposed that the kinase responsible for Ser 473 Survival factor withdrawal leads to FKHRL1 phosphorylation might in fact be PDK1, when it associates , nuclear translocation, and target gene with certain specific proteins, such as PDK1 interacting activation. Within the nucleus, FKHRL1 triggers apoptotic fragment (PIF), as seen by in vitro studies (Balendran et al. signals, possibly by inducing the expression of genes that are

1999). By dephosphorylating D3 residues on PtdIns(3,4,5)P3 critical for cell death, such as the Fas gene. A role for

and PtdIns(3,4)P2, PTEN works in opposition to the PI3K/ PTEN in the regulation of the Fas-induced response has also

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been suggested by studies with the Pten+/- mice mentioned arrest at the and also increases apoptosis induced above (Di Cristofano et al. 1999). In agreement with the by multiple distinct stimuli. The cell-cycle arrest property of involvement of Fas in this pathway, the defect in Fas-mediated PTEN has been detected in some cell types. In the presence apoptosis observed in these Pten+/- mice is mediated by of ectopic expression of wild-type PTEN, glioblastoma and activation of the PI3K/AKT pathway. renal carcinoma cells arrest at G1 (Furnari et al. 1998, PTEN’s involvement in antagonizing the PI3K/AKT Ramaswamy et al. 1999). This effect is enhanced by pathway was confirmed by subsequent studies which showed nocodazole, a G2 blocker, and by low serum conditions that cells lacking wild-type PTEN from gliomas, from patients (Furnari et al. 1998). It has been demonstrated that this with CS or from Pten-deficient mice have elevated levels of function of PTEN is AKT-dependent, and can be rescued by PtdIns(3,4,5)P3. As a result, the activity of AKT was also expression of an activated form of AKT (Li et al. 1998, increased in these cells, indicating that PTEN exerts its Ramaswamy et al. 1999). One of the targets of PTEN in its tumor-suppressor function by negatively regulating the ability to induce cell-cycle arrest has been suggested to be antiapoptotic PI-3 kinase/AKT signaling pathway (Fig. 3). the p27KIP1 cyclin-dependent kinase inhibitor (Li & Sun 1998, Biological evidence that PTEN acts as a tumor suppressor Cheney et al. 1999). It has been shown that PTEN induces came from studies indicating that wild-type PTEN suppresses p27 expression and allows the formation of complexes with the proliferation and the tumor growth of PTEN-deficient . This, in turn, reduces specifically cyclin-dependent glioblastoma cells. In contrast, mutant phosphatase-inactive kinase 2 (CDK2) activity and, consequently, results in low PTEN failed to suppress cell growth. The analysis of mutations phosphorylation of (Rb) protein. Low Rb from tumor specimens or cell lines derived from tumors phosphorylation leads to an arrest in cell-cycle progression. highlighted the importance of the phosphatase domain for the It remains to be established whether the increase in the tumor-suppressor function of PTEN. Indeed, a large proportion formation of p27/cyclinE/CDK2 complexes are truly a direct of these mutations maps to the region encoding the effect of PTEN’s action and which intermediate steps are phosphatase domain (Fig. 2 shows the spectrum of PTEN involved in producing this cell-cycle inhibition signal. mutations in Cowden syndrome). However, several mutations Another potential mechanism of cell-cycle control by and deletions are located distal to the phosphatase core motif, PTEN may be through inhibition of cyclin D1 accumulation. in the C-terminal region of PTEN. From an in vitro functional AKT phosphorylates GSK3 (glycogen synthase kinase 3), analysis of mutations located downstream of the phosphatase leading to its inactivation (Cross et al. 1995). Active GSK3 catalytic domain, in particular those located in exons 7 and 8, phosphorylates cyclin D1, targeting it for degradation (Diehl most of which resulted in premature stop codons, it was shown et al. 1998). AKT, therefore, appears to promote cyclin D1 that these C-terminal mutants inactivated the tumor-suppressor accumulation (Coffer et al. 1998, Cantley & Neel 1999). function of PTEN (Georgescu et al. 1999). The stability and phosphatase activity of these mutants were also affected. Similar results have been identified by independent groups, in PTEN as an apoptotic factor which a functional ‘map’ of PTEN defines an area spanned by Despite the observed cell arrest properties of PTEN,ithas codons 10 through 353 as a minimum requirement for the lipid been suggested that its tumor suppression function also phosphatase and cell-cycle arrest properties (S Ramaswamy & results from PTEN’s ability to increase cellular apoptotic W Sellers, personal communication). These findings suggest rates (Li et al. 1998, Stambolic et al. 1998). However, while that PTEN lipid phosphatase function is dependent on the integrity of a larger portion of the molecule, which extends far AKT is a known promoter of cell survival, an increase in beyond the catalytic motif. The implications of these data are PTEN has not been accompanied by high indexes of far-reaching, as it is expected that mutations scattered apoptosis in all cell types studied, in particular glioblastoma throughout the gene are likely to impair PTEN function. cells, in which the G1 arrest induced by PTEN was Nevertheless, mutations have been reported in glioblastomas extensively documented (Furnari et al. 1998, Ramaswamy et (Rasheed et al. 1997) which are located at the area coding for al. 1999). On the other hand, an increased sensitivity to the final amino acids of the PTEN protein, outside the minimal apoptosis induced by UV, heat shock and radiation was functional region. The relevance of these mutations for PTEN observed upon PTEN overexpression in mice null for PTEN function still remains to be elucidated. In light of the new (Stambolic et al. 1998). In addition, breast lines studies showing the relevance of PTEN’s PDZ domain for showed an increased number of apoptotic cells after PTEN PTEN’s function, it may be found that such mutations expression (Li et al. 1998). Further, it has been suggested alter PTEN stability (Wu et al. 2000). that an apoptotic response in breast cancer cell lines may ensue after initial G1 arrest, in a serial manner (Weng et al. 1999). It is also possible that increased apoptosis is not seen PTEN as an inducer of cell-cycle arrest upon PTEN expression in certain cell lines, due to additional PTEN’s ability to induce growth suppression appears to be molecular defects associated with apoptotic functions in mediated by at least two mechanisms: it promotes cell-cycle these cells.

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The mechanism of PTEN induction of apoptosis is still necessarily protein-related, PTEN activity. There is currently unclear. It has recently been suggested that Pten+/- no known PTEN mutant that maintains activity towards lipid heterozygous mice have impaired Fas-mediated cell death. substrates and loses protein-substrate function. It will be This function of PTEN appears to be mediated by PI-3 interesting to dissect these multiple functions of PTEN kinase-AKT signaling, as mentioned above. From these further. Other substrates tested, such as paxillin, c-src, and recent studies, a dosage effect of PTEN has been suggested tensin, all associated with adhesion, did not show reduced and Pten+/- mice develop an autoimmune-type syndrome phosphorylation, suggesting that PTEN acts specifically similar to that seen in Fas-deficient mice. These animals towards FAK (Tamura et al. 1998, 1999). develop lymph node , splenomegaly, and Additional insight into PTEN signaling came from work inflammatory infiltrates in multiple organs, in particular the from the same group (Gu et al. 1998) which has shown that kidneys in which immune complexes were observed in the PTEN inhibits the mitogen-activated protein kinase (MAPK) glomeruli, in some cases similar to the segmental pathway, via inhibition of Shc phosphorylation. Shc is an glomerulosclerosis of human or mice autoimmune syndromes SH2-phosphotyrosine-binding adapter protein that associates (Di Cristofano et al. 1999). However, the interrelation tyrosine kinases to Ras signaling by recruiting the growth between the PI3K/PTEN/AKT and the Fas signaling appears factor binding protein 2 (Grb2)-Sos complex to the to involve additional factors, as highlighted by another study plasma membrane in a tyrosine phosphorylation-dependent (Wick et al. 1999), in which ectopic expression of PTEN in manner (Rozakis-Adcock et al. 1992). This effect was not glioblastoma cells lacking functional PTEN sensitizes these observed when a phosphatase-inactive mutant of PTEN was cells to irradiation and Fas-FasL-induced apoptosis via expressed instead. Inhibition of this pathway was rescued, in increased caspase 3 activity. Sensitization of a PTEN-null part, by expression of an upstream molecule in the MAPK glioblastoma cell, U87MG, to CD95L apoptosis by wild-type signaling pathway, MAP or ERK kinase (MEK1). More PTEN is blocked by IGF-I in a PI3K-dependent manner. recent studies from this group revealed that while both FAK Although AKT is a downstream target of PI3K, the and Shc-mediated PTEN inhibition affect cell migration, the protection by IGF-I was not associated with the FAK pathway is associated with a directional migration, reconstitution of AKT phosphorylation in this model. while Shc is related to random migration (Gu et al. 1999). Further insights into the mechanisms of PTEN-induced The differences in the migration pattern are related to the apoptosis are still required to allow for a complete extent of actin cytoskeleton organization, which is more understanding of its functional role. In particular, the abundant in the FAK pathway. These PTEN migration and factor(s) responsible for a cell’s decision to induce cell-cycle adhesion properties have yet to be reproduced by other arrest or apoptosis as a result of PTEN activity remain to groups. be determined. A diagram summarizing currently known and Further studies aiming at characterizing PTEN’s presumed players in PTEN/AKT signaling is shown in Fig. 3. signaling through the FAK pathway revealed that p130Cas,a downstream target of FAK-mediated cell migration, does not associate directly with PTEN, although p130Cas phos- Are phospholipids the only PTEN substrates? phorylation levels are reduced in the presence of PTEN In the search for PTEN protein substrates, it has been shown possibly via inhibition of FAK activity (Tamura et al. 1999). that PTEN expression is associated with reduced cell In addition, while it has been shown that overexpression of migration, spreading and focal adhesion formation, and these FAK and p130Cas is able to rescue PTEN-inhibited cell phenomena were linked to the integrin pathway (Tamura et migration and spreading, only FAK expression reversed al. 1998) (Fig. 4). It has been noted that wild-type PTEN PTEN effects on cell growth. This suggests that certain binds to the same complex as the focal-adhesion kinase functions of PTEN are mediated by pathways that diverge at (FAK) and reduces its phosphorylation. Phosphatase-inactive the level of FAK. In agreement with this notion, a recent PTEN mutants, including C124S and the substrate-trapping report shows that the lipid phosphatase domain of PTEN is mutant, D92A, do not show the same properties (Tamura et not required for PTEN inhibition of migration in glioma cells, al. 1998). Interestingly, a PTEN mutation found in patients suggesting that PTEN exerts functions that are independent with CS, in which glycine is replaced by glutamine in a of AKT inhibition (Maier et al. 1999). residue within the catalytic core motif (G129E), has been shown to behave in a similar manner to wild-type PTEN with Crystal structure solved – new clues into regard to migration, spreading and focal adhesion inhibition PTEN substrate specificity (Tamura et al. 1998). However, this mutant has been shown to lack phospholipid activity (Furnari et al. 1998, Myers et Recently, the PTEN crystal structure has been solved (Lee et al. 1998). This finding gives further hints on PTEN functions al. 1999b). This information provides insights into putative and couples the Cowden syndrome-related and possibly functions of this tumor suppressor, as it helps in the tumor-associated abnormalities to the phospholipid, but not identification of domains other than the catalytic core for

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Figure 4 PTEN function: an integrated model. This diagram attempts to integrate, in a concise form, the intracellular signals regulated by PTEN, according to recent studies. One of the signaling pathways, triggered by growth factors, involves predominantly PTEN’s antagonism to PI-3 kinase activity, which is associated with PTEN dephosphorylation of lipid substrates and relates to control of cell growth and survival. These latter properties appear to be mediated by multiple putative factors whose detailed characterization is still unknown (some of the factors regulated by AKT and presumed to be involved in mediating such functions are shown in the figure). The other, less well studied, pathway of PTEN activity involves (but is not limited to) protein substrates, including FAK and Shc, which mediate cell migration and adhesion functions (see text for details).

PTEN activity, which can be used as the basis for further of critical basic residues at this site results in reduced biochemical studies. The PTEN structure reveals a membrane affinity of PTEN and also impairs its ability to phosphatase domain that is similar to protein phosphatases suppress cell growth. The phosphatase and C2 domains but whose is extended to accommodate the associate extensively, suggesting that the may

phosphoinositide substrate (PtdIns(3,4,5)P3). In addition, a serve to assist in positioning the catalytic domain on the C2 domain was identified in the structure. C2 domains have membrane. This finding is in agreement with the PTEN null been characterized in a growing number of eukaryotic phenotype caused by certain mutations that locate away from signaling proteins that interact with cellular membranes and the catalytic core (see above). In addition, it has repeatedly mediate a broad array of critical intracellular processes, been shown that detection of significant catalytic activity of including membrane trafficking, the generation of PTEN is only obtained when highly acidic substrates are used lipid-second messengers, activation of GTPases, and the (Li & Sun 1997, Myers et al. 1997). The characterization of control of protein phosphorylation (Nalefski & Falke 1996). the structure of PTEN has provided insights into why that These domains bind a variety of different ligands and might be. Two basic residues located in the catalytic motif, substrates, including Ca2+, phospholipids, inositol poly- K125 and K128, as well as H93 outside the catalytic motif, phosphates, and intracellular proteins. The PTEN C2 domain give the pocket a highly basic character which is absent from extends towards the C-terminus of PTEN and binds other phosphatases, such as vaccinia H1-related phosphatase phospholipid membranes in vitro. In consequence, mutation (VHR) and protein tyrosine phosphatase 1B (PTP1B) (Lee et

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al. 1999b). This is in keeping with PTEN’s preference for mutants, in particular G129E, might have suggested. The highly acidic substrates. phosphatase domain appears to be contained in a single In summary, the crystal structure of PTEN has helped in portion of the structure. the understanding of some of the functions attributed to this phosphatase by biochemical studies and has additionally More on PTEN knockout mice provided several clues with regard to PTEN’s unique features. The C2 domain is involved in PTEN’s binding to As mentioned above, three different PTEN knockout mice the membrane, and the lipid phosphatase activity, as were generated. While many features of these animals were suggested by in vitro and in vivo studies, appears to be common to all three mutants, including embryo lethality, critical for PTEN tumor suppressor function. In particular, some of the observed phenotypes varied substantially. One

PtdIns(3,4,5)P3, the only PTEN substrate to carry a phosphate of the models describes deficient development of the three at position D5, is more important to PTEN tumor supressor germinal layers (Di Cristofano et al. 1998), while a second activity than the other D3 substrates without a phosphate at one reports defects on patterning of the cephalic and caudal

this position, PtdIns(3,4)P2 and PtdIns(3)P. This conclusion regions (Suzuki et al. 1998). A third mutant (Podsypanina has been made based on the finding that mutations affecting et al. 1999) displays aberrant lymph node architecture and the histidine at position 93 affect the phosphatase activity hyperplasia, compatible with a B-lymphocyte defect. almost exclusively towards the triphosphate compound, in Lymphomas and non-hamartomatous colonic polyps were comparison with the other two lipid substrates (Lee et al. also observed in this model. The major features of each one 1999b). From the data provided by PTEN structure, it does of the described mutants are presented in Table 2. not appear that PTEN carries an additional phosphatase Differences in the mouse background have been raised as a domain that accounts for a protein-directed activity distinct possible explanation for the differences observed, as well as from the lipid activity, as some of the functional studies with the type of targeting construct used and the time of

Table 2 Comparison between the three PTEN knockout mice models generated to date. Knockout features Suzuki et al. (1998) Di Cristofano et al. (1998) Podsypanina et al. (1999) Mouse background CD1 and intercrosses with 129/Sv and intercrosses with 129svJ and intercrosses with C57BL/6J CJ7ES and C57BL/6 C57BL6/J and B6/129 PTEN exons deleted 3–5 4–5 5 Embryonic day of death E9.5 E7.5 E6.5 Developmental features of Abnormal cephalo–caudal Abnormal differentiation of Abnormal lymph node mutants patterning the three germ layers in architecture Abnormal placentations vitro Lymphoid aggregates Defects in generation of mesodermal lineages Tumorigenic features of T-cell leukemia/lymphomas Dysplastic and inflammatory Follicular and papillary heterozygous mutants Adenomatous liver colonic polyps thyroid tumors Teratocarcinoma Prostate hyperplasia Prostate hyperplasia and Prostate carcinoma Gonadostromal tumors neoplasia Hamartomatous colonic Teratoma Pre-neoplastic multifocal polyps Papillary-like thyroid complex atypical carcinoma hyperplasia of endometrium Myeloid leukemia Lymphoblastic lymphomas Skin hyperkeratosis Poorly differentiated leukemia Liver Hyperplastic polyps of colon with lymphoid infiltrates Lymphoid hyperplasia (females > males) Tumor LOH Lymphomas ND +Lymphomas Other features Increased resistance to Enhanced Increased apoptotic rates apoptosis anchorage-independent (measured by annexin V) in Tumorigenesis accelerated by growth heterozygous mice γ-irradiation Histological abnormalities in bold are commonly found in Cowden syndrome (CS). Other common features of CS that have not been observed in any of the mutants are: breast cancer, multiple hamartomas of skin, breast and thyroid, and macrocephaly. Lymphomas have been described in CS patients; however, it has not been formally established whether they are a true component of the syndrome. ND; not described.

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observation of the heterozygote animals. Of interest, few regulating glucose metabolism and pathways relating to features typical of CS and/or BRR have been described in aging control. these animals. It appears from the knockout data that unknown additional factors possibly help modulate the phenotypical Future directions of PTEN studies features on a tissue-dependent and time of development Despite the large amount of data presently available on basis. Longer observation times of the heterozygotes might PTEN, there are still many open questions with regard to be required for additional features to be identified. It is its regulation and cellular role. The recently solved crystal possible that in the months to come, reports will be available structure of PTEN has already provided several insights into on aging mutant animals that will provide such important the catalytic function of this phosphatase, and has helped us +/- information. Inter-crosses between these Pten knockout to understand in further detail certain aspects of PTEN’s animals with other mutant strains associated with tumors and/ substrate specificity, in comparison with other tyrosine and or developmental abnormalities are also likely to shed light dual-specificity phosphatases (Lee et al. 1999b). Figure 4 onto interactive pathways. The role of PTEN in certain attempts to summarize the current signaling pathways tissues only may be further studied using Cre-Lox systems, associated with PTEN function. However, several issues still whereby tissue-specific PTEN inactivation can be attained. remain unresolved: there are no known upstream regulators of PTEN levels, although there have been some conflicting reports on the potential role of TGFβ on PTEN regulation PTEN homologs – Caenorhabditis (Li & Sun 1997, Lee et al. 1999a). Also, there are no current elegans data on the mechanisms of PTEN activation; it is not known, for example, whether PTEN activity is modulated by In the nematode, C. elegans, an insulin-related pathway post-translational modification such as phosphorylation, and regulates metabolism, development and longevity. Wild-type how PTEN is degraded or inactivated. The putative role of animals enter the developmentally arrested dauer stage in naturally occurring PTEN alternative splice forms is not response to high levels of a secreted pheromone, presently known. Studies determining the actual relevance of accumulating large amounts of fat. In this state, metabolic a PTEN dosage effect and whether haploinsufficiency indeed functions are suppressed and it is associated with a prolonged plays a role in tumorigenesis in humans are expected to be life-span of the worms. Several mutants have been identified available soon. The nature of the factor or factors involved in C. elegans which associate with defective dauer formation in PTEN’s decision towards inducing cell-cycle arrest or (DAF mutants). DAF-2 mutants, which encode a homolog of apoptosis in certain cell types is still elusive. Due to its the mammalian insulin receptor or insulin receptor-like important function as a tumor suppressor, PTEN is a natural (IRL), and AGE-1 mutants, homologs of the catalytic subunit candidate for and it is likely that the next few of mammalian PI-3 kinase, arrest development at the dauer years will provide preliminary analyses attempting to provide stage, and this effect can be rescued by expression of answers to its therapeutic potential in several tumor types. DAF-16, a downstream element in this pathway (Ogg et al. 1997). DAF-16 forkhead is inactivated by AKT-1 and AKT-2 homologs. Recent studies of this pathway Acknowledgements revealed that DAF-18, a gene required for dauer formation, is the PTEN homolog in C. elegans (Ogg & Ruvkun 1998, The author is grateful to Ricardo Aguiar for his helpful Gil et al. 1999, Mihaylova et al. 1999, Rouault et al. 1999). comments and discussions. Thanks are also due to Charis Loss-of-function mutations of DAF-18 prevent entry into the Eng for her critical review of the manuscript. P L M Dahia is dauer stage, and also alter life span, fertility, brood size and the recipient of a Susan G Komen Breast Cancer Foundation metabolism of nondauer animals through inhibition of the Research Fellowship. DAF-2/IRL and AGE-1/PI3K pathways. Thus, similar to that observed in , the PTEN homolog in C. elegans References antagonizes the function of PI-3 kinase (Ogg & Ruvkun 1998, Gil et al. 1999, Mihaylova et al. 1999, Rouault et al. Albarosa R, Colombo BM, Roz L, Magnani I, Pollo B, Cirenei N, 1999). Detailed study of specific DAF-18 mutants has Giani C, Conti AMF, DiDonato S & Finocchiaro G 1996 suggested that certain phenotypes, such as fertility, are less mapping of gliomas suggests the presence of two small sensitive to PtdIns(3,4,5)P levels than dauer formation. The regions for candidate tumor-suppressor genes in a 17-cM 3 interval on chromosome 10q. American Journal of Human existence of distinct phenotypes that arise from different 58 1260–1267. DAF-18 mutants raises the interesting possibility that similar Alessi DR, James SR, Downes CP, Holmes AB, Gaffney PR, diversity may be seen with the mammalian homolog. It is Reese CB & Cohen P 1997 Characterization of a interesting to speculate whether PTEN is involved in 3-phosphoinositide-dependent protein kinase which

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phosphorylates and activates protein kinase Balpha. Current Dahia PLM, Aguiar RCT, Alberta J, Kum JB, Caron S, Sill H, Biology 7 261–269. Marsh DJ, Ritz J, Freedman A, Stiles C & Eng C 1999 PTEN is Ali IU, Schriml LM & Dean M 1999 Mutational spectra of PTEN/ inversely correlated with the cell survival factor AKT/PKB and MMAC1 gene: a tumor suppressor with lipid phosphatase is inactivated via multiple mechanisms in haematological activity. Journal of the National Cancer Institute 91 1922–1932. malignancies. Human Molecular Genetics 8 185–193. Balendran A, Casamayor A, Deak M, Paterson A, Gaffney P, Dahia PLM, Gimm O, Chi H, Marsh DJ, Reynolds PR & Eng C Currie R, Downes CP & Alessi DR 1999 PDK1 acquires PDK2 2000 Absence of germline mutations in MINPP1,a activity in the presence of a synthetic peptide derived from the phosphatase-encoding gene centromeric of PTEN, in patients carboxyl terminus of PRK2. Current Biology 9 393–404. with Cowden and Bannayan-Riley-Ruvalcaba syndrome without Bose S, Wang SI, Terry MB, Hibshoosh H & Parsons R 1998 germline PTEN mutations. Journal of Medical Genetics (In Allelic loss of chromosome 10q23 is associated with tumor Press). progression in breast . 17 123–127. Datta SR, Dudek H, Tao X, Masters S, Fu H, Gotoh Y & Brunet A, Bonni A, Zigmond MJ, Lin MZ, Juo P, Hu LS, Greenberg ME 1997 AKT phosphorylation of BAD couples Anderson MJ, Arden KC, Blenis J & Greenberg ME 1999 AKT survival signals to the cell-intrinsic death machinery. Cell 91 promotes cell survival by phosphorylating and inhibiting a 231–241. Forkhead transcription factor. Cell 96 857–868. Delcommenne M, Tan C, Gray V, Rue L, Woodgett J & Dedhar S Caffrey JJ, Hidaka K, Matsuda M, Hirata M & Shears SB 1999 1998 Phosphoinositide-3-OH kinase-dependent regulation of The human and rat forms of multiple inositol polyphosphate glycogen synthase kinase 3 and /AKT by the phosphatase: functional homology with a histidine acid integrin-linked kinase. PNAS 95 11211–11216. phosphatase up-regulated during endochondral ossification. Di Cristofano A, Pesce B, Cordon-Cardo C & Pandolfi PP 1998 FEBS Letters 442 99–104. PTEN is essential for embryonic development and tumour Cairns P, Okami K, Halachmi S, Halachmi N, Esteller M, Herman suppression. Nature Genetics 19 348–355. JG, Jen J, Isaacs WB, Bova GS & Sidransky D 1997 Frequent Di Cristofano A, Kotsi P, Peng YF, Cordon-Cardo C, Elkon KB & inactivation of PTEN/MMAC1 in primary prostate cancer. Pandolfi PP 1999 Impaired fas response and autoimmunity in Cancer Research 57 4997–5000. Pten(+/-) mice. Science 285 2122–2125. Cairns P, Evron E, Okami K, Halachmi N, Esteller M, Herman JG, Diehl JA, Cheng M, Roussel MF & Sherr CJ 1998 Glycogen Bose S, Wang SI, Parsons R & Sidransky D 1998 Point synthase kinase-3beta regulates cyclin D1 proteolysis and mutation and homozygous deletion of PTEN/MMAC1 in subcellular localization. Genes and Development 12 3499–3511. primary bladder cancers. Oncogene 16 3215–3218. Eng C 1997 Cowden syndrome. Journal of Genetic Counseling 6 Cantley LC & Neel BG 1999 New insights into tumor suppression: 181–191. PTEN suppresses tumor formation by restraining the phospho- Feilotter HE, Coulon V, McVeigh JL, Boag AH, Dorion-Bonnet F, inositide 3-kinase/AKT pathway. PNAS 96 4240–4245. Duboue B, Latham WC, Eng C, Mulligan LM & Longy M 1999 Chen J, Lindblom P & Lindblom A 1998 A study of the PTEN/ Analysis of the 10q23 chromosomal region and the PTEN gene MMAC1 gene in 136 breast cancer families. Human Genetics in human sporadic breast carcinoma. British Journal of Cancer 102 124–125. 79 718–723. Cheney IW, Neuteboom ST, Vaillancourt MT, Ramachandra M & FitzGerald MG, Marsh DJ, Wahrer D, Bell D, Caron S, Shannon Bookstein R 1999 Adenovirus-mediated gene transfer of KE, Ishioka C, Isselbacher KJ, Garber JE, Eng C & Haber DA MMAC1/PTEN to glioblastoma cells inhibits entry by 1998 Germline mutations in PTEN are an infrequent cause of the recruitment of p27Kip1 into cyclin E/CDK2 complexes. genetic predisposition to breast cancer. Oncogene 17 727–731. Cancer Research 59 2318–2323. Furnari FB, Lin H, Huang HS & Cavenee WK 1997 Growth Chi H, Tiller GE, Dasouki MJ, Romano PR, Wang J, O’Keefe RJ, suppression of glioma cells by PTEN requires a functional Puzas JE, Rosier RN & Reynolds PR 1999 Multiple inositol phosphatase catalytic domain. PNAS 94 12479–12484. polyphosphate phosphatase: evolution as a distinct group within Furnari FB, Huang HJ & Cavenee WK 1998 The phosphoinositol the histidine phosphatase family and chromosomal localization phosphatase activity of PTEN mediates a serum-sensitive G1 of the human and mouse genes to 10q23 and 19. growth arrest in glioma cells. Cancer Research 58 5002–5008. Genomics 56 324–336. Georgescu MM, Kirsch KH, Akagi T, Shishido T & Hanafusa H Coffer PJ, Jin J & Woodgett JR 1998 Protein kinase B (c-AKT): a 1999 The tumor-suppressor activity of PTEN is regulated by its multifunctional of phosphatidylinositol 3-kinase carboxyl-terminal region. PNAS 96 10182–10187. activation. Biochemical Journal 335 1–13. Gil EB, Malone Link E, Liu LX, Johnson CD & Lees JA 1999 Cross DA, Alessi DR, Cohen P, Andjelkovich M & Hemmings BA Regulation of the insulin-like developmental pathway of 1995 Inhibition of glycogen synthase kinase-3 by insulin Caenorhabditis elegans by a homolog of the PTEN tumor mediated by protein kinase B. Nature 378 785–789. suppressor gene. PNAS 96 2925–2930. Dahia PL, Marsh DJ, Zheng Z, Zedenius J, Komminoth P, Frisk T, Gronbaek K, Zeuthen J, Guldberg P, Ralfkiaer E & Hou-Jensen K Wallin G, Parsons R, Longy M, Larsson C & Eng C 1997 1998 Alterations of the MMAC1/PTEN gene in lymphoid Somatic deletions and mutations in the Cowden disease gene, malignancies (letter). Blood 91 4388–4390. PTEN, in sporadic thyroid tumors. Cancer Research 57 4710– Gu J, Tamura M & Yamada KM 1998 Tumor suppressor PTEN 4713. inhibits integrin- and growth factor-mediated mitogen-activated Dahia PL, FitzGerald MG, Zhang X, Marsh DJ, Zheng Z, Pietsch protein (MAP) kinase signaling pathways. Journal of Cell T, von Deimling A, Haluska FG, Haber DA & Eng C 1998 A Biology 143 1375–1383. highly conserved processed PTEN pseudogene is located on Gu J, Tamura M, Pankov R, Danen EH, Takino T, Matsumoto chromosome band 9p21. Oncogene 16 2403–2406. K & Yamada KM 1999 Shc and FAK differentially regulate cell

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motility and directionality modulated by PTEN. Journal of Cell BT, Heimdal K, Lubs H, Moller P & King MC 1997 Inherited Biology 146 389–403. mutations in PTEN that are associated with breast cancer, Halachmi N, Halachmi S, Evron E, Cairns P, Okami K, Saji M, Cowden disease, and juvenile polyposis. American Journal of Westra WH, Zeiger MA, Jen J & Sidransky D 1998 Somatic Human Genetics 61 1254–1260. mutations of the PTEN tumor suppressor gene in sporadic McMenamin ME, Soung P, Perera S, Kaplan I, Loda M & Sellers follicular thyroid tumors. Genes, Chromosomes and Cancer 23 WR 1999 Loss of PTEN expression in paraffin-embedded 239–243. primary prostate cancer correlates with high Gleason score and Jacoby RF, Schlack S, Cole CE, Skarbek M, Harris C & Meisner advanced stage. Cancer Research 59 4291–4296. LF 1997 A juvenile polyposis tumor suppressor locus at 10q22 Maehama T & Dixon JE 1999 PTEN: a tumour suppressor that is deleted from nonepithelial cells in the lamina propria. functions as a phospholipid phosphatase. Trends in Cell Biology Gastroenterology 112 1398–1403. 9 125–128. Kobayashi H, Hosoda F, Maseki N, Sakurai M, Imashuku S, Ohki Maier D, Jones G, Li X, Schonthal AH, Gratzl O, Van Meir EG & M & Kaneko Y 1997 Hematologic malignancies with the Merlo A 1999 The PTEN lipid phosphatase domain is not t(10;11) (p13;q21) have the same molecular event and a variety required to inhibit of glioma cells. Cancer Research 59 of morphologic or immunologic phenotypes. Genes, 5479–5482. Chromosomes and Cancer 20 253–259. Marsh DJ, Dahia PL, Zheng Z, Liaw D, Parsons R, Gorlin RJ & Kohno T, Takahashi M, Manda R & Yokota J 1998 Inactivation of Eng C 1997a Germline mutations in PTEN are present in the PTEN/MMAC1/TEP1 gene in human lung cancers. Genes, Bannayan-Zonana syndrome (letter). Nature Genetics 16 333– Chromosomes and Cancer 22 152–156. 334. Lee J, Park BJ, Park JH, Yang MH & Chi SG 1999a TGF-beta1 Marsh DJ, Roth S, Lunetta KL, Hemminki A, Dahia PL, Sistonen inhibition of apoptosis through the transcriptional up-regulation P, Zheng Z, Caron S, van Orsouw NJ, Bodmer WF, Cottrell SE, of Bcl-X(L) in human monocytic leukemia U937 cells. Dunlop MG, Eccles D, Hodgson SV, Jarvinen H, Kellokumpu I, Experimental and Molecular Medicine 31 126–133. Markie D, Neale K, Phillips R, Rozen P, Syngal S, Vijg J, Lee J-O, Yang H, Georgescu M-M, Di Cristofano A, Maehama T, Tomlinson IP, Aaltonen LA & Eng C 1997b Exclusion of PTEN Shi Y, Dixon JE, Pandolfi P & Pavletich NP 1999b Crystal and 10q22-24 as the susceptibility locus for juvenile polyposis structure of the PTEN tumor suppressor: implications for its syndrome. Cancer Research 57 5017–5021. phosphoinositide phosphatase activity and membrane association. Marsh DJ, Coulon V, Lunetta KL, Rocca-Serra P, Dahia PL, Cell 99 323–334. Zheng Z, Liaw D, Caron S, Duboue B, Lin AY, Richardson AL, Li D-M & Sun H 1997 TEP1, encoded by a candidate tumor Bonnetblanc JM, Bressieux JM, Cabarrot-Moreau A, Chompret suppressor locus, is a novel protein tyrosine phosphatase A, Demange L, Eeles RA, Yahanda AM, Fearon ER, Fricker JP, regulated by transforming growth factor B. Cancer Research 57 Gorlin RJ, Hodgson SV, Huson S, Lacombe D, Eng C et al. 2124–2129. 1998a Mutation spectrum and genotype-phenotype analyses in Li DM & Sun H 1998 PTEN/MMAC1/TEP1 suppresses the Cowden disease and Bannayan-Zonana syndrome, two tumorigenicity and induces G1 cell cycle arrest in human hamartoma syndromes with germline PTEN mutation. Human glioblastoma cells. PNAS 95 15406–15411. Molecular Genetics 7 507–515. Li J, Yen C, Liaw D, Podsypanina K, Bose S, Wang S, Puc J, Marsh DJ, Dahia PL, Coulon V, Zheng Z, Dorion-Bonnet F, Call Miliaresis C, Rodgers L, McCombie R, Bigner SH, Giovanella BC, Ittman M, Tycko B, Hibshoosh H, Wigler MH & Parsons R KM, Little R, Lin AY, Eeles RA, Goldstein AM, Hodgson SV, 1997 PTEN, a putative protein tyrosine phosphatase gene Richardson AL, Robinson BG, Weber HC, Longy M & Eng C mutated in human brain, breast and prostate cancer. Science 275 1998b Allelic imbalance, including deletion of PTEN/MMACI, 1943–1947. at the Cowden disease locus on 10q22-23, in hamartomas from Li J, Simpson L, Takahashi M, Miliaresis C, Myers MP, Tonks patients with Cowden syndrome and germline PTEN mutation. N & Parsons R 1998 The PTEN/MMAC1 tumor suppressor Genes, Chromosomes and Cancer 21 61–69. induces cell death that is rescued by the AKT/protein kinase B Marsh DJ, Dahia PLM, Caron S, Kum JB, Frayling IM, Tomlinson oncogene. Cancer Research 58 5667–5672. IPM, Hughes KS, Eeles RA, Hodgson SV, Murday VA, Liaw D, Marsh DJ, Li J, Dahia PL, Wang SI, Zheng Z, Bose S, Houlston R & Eng C 1998c Germline PTEN mutations in Call KM, Tsou HC, Peacocke M, Eng C & Parsons R 1997 Cowden syndrome-like families. Journal of Medical Genetics 35 Germline mutations of the PTEN gene in Cowden disease, an 881–885. inherited breast and thyroid . Nature Genetics Marsh DJ, Kum JB, Lunetta KL, Bennett MJ, Gorlin RJ, Ahmed 16 64–67. SF, Bodurtha J, Crowe C, Curtis MA, Dasouki M, Dunn T, Feit Liscovitch M & Cantley LC 1994 Lipid second messengers. Cell H, Geraghty MT, Graham Jr JM, Hodgson SV, Hunter A, Korf 77 329–334. BR, Manchester D, Miesfeldt S, Murday VA, Nathanson KL, Liu W, James CD, Frederick L, Alderete BE & Jenkins RB 1997 Parisi M, Pober B, Romano C, Tolmie JL, Trembath R, Winter PTEN/MMAC1 mutations and EGFR amplification in glio- RM, Zackai EH, Zori RT, Weng LP, Dahia PL & Eng C 1999 blastomas. Cancer Research 57 5254–5257. PTEN mutation spectrum and genotype–phenotype correlations Longy M, Coulon V, Duboue B, David A, Larregue M, Eng C, in Bannayan-Riley-Ruvalcaba syndrome suggest a single entity Amati P, Kraimps JL, Bottani A, Lacombe D & Bonneau D with Cowden syndrome. Human Molecular Genetics 8 1461– 1998 Mutations of PTEN in patients with Bannayan- 1472. Riley-Ruvalcaba phenotype. Journal of Medical Genetics 35 Maxwell GL, Risinger JI, Gumbs C, Shaw H, Bentley RC, Barrett 886–889. JC, Berchuck A & Futreal PA 1998 Mutation of the PTEN Lynch ED, Ostermeyer EA, Lee MK, Arena JF, Ji H, Dann J, tumor suppressor gene in endometrial . Cancer Swisshelm K, Suchard D, MacLeod PM, Kvinnsland S, Gjertsen Research 58 2500–2503.

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Mihaylova VT, Borland CZ, Manjarrez L, Stern MJ & Sun H 1999 Rasheed BKA, McLendon RE, Friedman HS, Friedman AH, Fuchs The PTEN tumor suppressor homolog in Caenorhabditis elegans HE, Bigner DD & Bigner SH 1995 Chromosome 10 deletion regulates longevity and dauer formation in an insulin mapping in human gliomas: a common deletion region in 10q25. receptor-like signaling pathway. PNAS 96 7427–7432. Oncogene 10 2243–2246. Myers MP & Tonks NK 1997 PTEN: sometimes taking it off can Rasheed BK, Stenzel TT, McLendon RE, Parsons R, Friedman be better than putting it on. American Journal of Human AH, Friedman HS, Bigner DD & Bigner SH 1997 PTEN gene Genetics 61 1234–1238. mutations are seen in high-grade but not in low-grade gliomas. Myers MP, Stolarov J, Eng C, Li J, Wang SI, Wigler MH, Parsons Cancer Research 57 4187–4190. R & Tonks NK 1997 PTEN, the tumor suppressor from human Risinger JI, Hayes AK, Berchuck A & Barrett JC 1997 PTEN/ chromosome 10q23, is a dual specificity phosphatase. PNAS 94 MMAC1 mutations in endometrial cancers. Cancer Research 57 9052–9057. 4736–4738. Myers MP, Pass I, Batty IH, Van der Kaay J, Stolarov JP, Rohn JL, Hueber AO, McCarthy NJ, Lyon D, Navarro P, Hemmings BA, Wigler MH, Downes CP & Tonks NK 1998 The Burgering BM & Evan GI 1998 The opposing roles of the AKT lipid phosphatase activity of PTEN is critical for its tumor and c-Myc signalling pathways in survival from CD95-mediated supressor function. PNAS 95 13513–13518. apoptosis. Oncogene 17 2811–2818. Nalefski EA & Falke JJ 1996 The C2 domain calcium-binding Romashkova JA & Makarov SS 1999 NF-kappaB is a target of motif: structural and functional diversity. Protein Science 5 AKT in anti-apoptotic PDGF signalling. Nature 401 86–90. 2375–2390. Rouault JP, Kuwabara PE, Sinilnikova OM, Duret L, Thierry-Mieg Nelen MR, Padberg GW, Peeters EAJ, Lin AY, van den Helm B, D & Billaud M 1999 Regulation of dauer larva development in Frants RR, Coulon V, Goldstein AM, van Reen MMM, Easton Caenorhabditis elegans by daf-18, a homologue of the tumour DF, Eeles RA, Hodgson S, Mulvihill JJ, Murday VA, Tucker suppressor PTEN. Current Biology 9 329–332. MA, Mariman ECM, Starink TM, Ponder BAJ, Ropers HH, Rozakis-Adcock M, McGlade J, Mbamalu G, Pelicci G, Daly R, Li Kremer H, Longy M & Eng C 1996 Localization of the gene for W, Batzer A, Thomas S, Brugge J, Pelicci PG et al. 1992 Cowden disease to 10q22-23. Nature Genetics 13 114–116. Association of the Shc and Grb2/Sem5 SH2-containing proteins Nelen MR, van Staveren WC, Peeters EA, Hassel MB, Gorlin RJ, is implicated in activation of the Ras pathway by tyrosine kinases. Nature 360 689–692. Hamm H, Lindboe CF, Fryns JP, Sijmons RH, Woods DG, Stambolic V, Suzuki A, de la Pompa JL, Brothers GM, Mirtsos C, Mariman EC, Padberg GW & Kremer H 1997 Germline Sasaki T, Ruland J, Penniger JM, Siderovski DP & Mak TW mutations in the PTEN/MMAC1 gene in patients with Cowden 1998 Negative regulation of PKB/AKT-dependent cell survival disease. Human Molecular Genetics 6 1383–1387. by the tumor suppressor PTEN. Cell 95 29–39. Obata K, Morland SJ, Watson RH, Hitchcock A, Chenevix-Trench Steck PA, Pershouse MA, Jasser SA, Yung WKA, Lin H, Ligon G, Thomas EJ & Campbell IG 1998 Frequent PTEN/MMAC AH, Langford LA, Baumgard ML, Hattier T, Davis T, Frye C, mutations in endometrioid but not serous or mucinous epithelial Hu R, Swedlund B, Teng DHF & Tavtigian SV 1997 ovarian tumors. Cancer Research 58 2095–2097. Identification of a candidate tumour suppressor gene, MMAC1, Ogg S & Ruvkun G 1998 The C. elegans PTEN homolog, at chromosome 10q23.3 that is mutated in multiple advanced DAF-18, acts in the insulin receptor-like metabolic signaling cancers. Nature Genetics 15 356–362. pathway. Molecular Cell 2 887–893. Sun H, Lesche R, Li DM, Liliental J, Zhang H, Gao J, Gavrilova Ogg S, Paradis S, Gottlieb S, Patterson GI, Lee L, Tissenbaum N, Mueller B, LiuX&WuH1999 PTEN modulates cell cycle HA & Ruvkun G 1997 The Fork head transcription factor progression and cell survival by regulating phosphatidylinositol DAF-16 transduces insulin-like metabolic and longevity signals 3,4,5,-trisphosphate and AKT/protein kinase B signaling in C. elegans. Nature 389 994–999. pathway. PNAS 96 6199–6204. Olschwang S, Serova-Sinilnikova OM, Lenoir GM & Thomas G Suzuki A, de la Pompa JL, Stambolic V, Elia AJ, Sasaki T, del 1998 PTEN germ-line mutations in juvenile polyposis coli Barco Barrantes I, Ho A, Wakeham A, Itie A, Khoo W, (letter). Nature Genetics 18 12–14. Fukumoto M & Mak TW 1998 High cancer susceptibility and Perren A, Weng L-P, Boag A, Ziebold U, Thakore K, Dahia P, embryonic lethality associated with mutation of the PTEN tumor Komminoth P, Lees J, Mulligan L, Mutter G & Eng C 1999 suppressor gene in mice. Current Biology 8 1169–1178. Immunohistochemical evidence of loss of PTEN expression in Tamura M, Gu J, Matsumoto K, Aota S, Parsons R & Yamada primary ductal adenocarcinomas of the breast. American Journal KM 1998 Inhibition of cell migration, spreading, and focal of Pathology 155 1253–1260. adhesions by tumor suppressor PTEN. Science 280 1614–1617. Pfeifer SL, Herzog TJ, Tribune DJ, Mutch DG, Gersell DJ & Tamura M, Gu J, Takino T & Yamada KM 1999 Tumor Goodfellow PJ 1995 Allelic loss of sequence from the long arm suppressor PTEN inhibition of cell invasion, migration, and of chromosome 10 and replication errors in endometrial cancers. growth: differential involvement of focal adhesion kinase and Cancer Research 55 1922–1926. p130Cas. Cancer Research 59 442–449. Podsypanina K, Ellenson LH, Nemes A, Gu J, Tamura M, Yamada Tashiro H, Blazes MS, Wu R, Cho KR, Bose S, Wang SI, Li J, KM, Cordon-Cardo C, Catoretti G, Fisher PE & Parsons R 1999 Parsons R & Ellenson LH 1997 Mutations in PTEN are frequent Mutation of PTEN/Mmac1 in mice causes neoplasia in multiple in endometrial carcinoma but rare in other common organ systems. PNAS 96 1563–1568. gynecological malignancies. Cancer Research 57 3935–3940. Ramaswamy S, Nakamura N, Vazquez F, Batt DB, Perera S, Teng DH, Hu R, Lin H, Davis T, Iliev D, Frye C, Swedlund B, Roberts TM & Sellers WR 1999 Regulation of G1 progression Hansen KL, Vinson VL, Gumpper KL, Ellis L, El-Naggar A, by the PTEN tumor suppressor protein is linked to inhibition of Frazier M, Jasser S, Langford LA, Lee J, Mills GB, Pershouse the phosphatidylinositol 3-kinase/AKT pathway. PNAS 96 2110– MA, Pollack RE, Tornos C, Troncoso P, Yung WK, Fujii G, 2115. Berson A, Steck PA et al. 1997 MMAC1/PTEN mutations in

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Endocrine-Related Cancer (2000) 7 115–129

specimens and tumor cell lines. Cancer Research Weng L-P, Smith WM, Dahia PLM, Ziebold U, Gil E, Lees JA & 57 5221–5225. Eng C 1999 PTEN suppresses breast cancer cell growth by Trybus TM, Burgess AC, Wojno KJ, Glover TW & Macoska JA phosphatase activity-dependent G1 arrest followed by cell death. 1996 Distinct areas of allelic loss on chromosomal regions 10p Cancer Research 59 5808–5814. and 10q in human prostate cancer. Cancer Research 56 2263– Whang YE, Wu X & Sawyers CL 1998a Identification of a 2267. pseudogene that can masquerade as a mutant allele of the PTEN/ Tsou HC, Teng DH, Ping XL, Brancolini V, Davis T, Hu R, Xie MMAC1 tumor suppressor gene. Journal of the National Cancer XX, Gruener AC, Schrager CA, Christiano AM, Eng C, Steck P, Institute 90 859–861. Ott J, Tavtigian SV & Peacocke M 1997 The role of MMAC1 Whang YE, Wu X, Suzuki H, Reiter RE, Tran C, Vessella RL, mutations in early-onset breast cancer: causative in association Said JW, Isaacs WB & Sawyers CL 1998b Inactivation of the with Cowden syndrome and excluded in BRCA1-negative cases. tumor suppressor PTEN/MMAC1 in advanced human prostate American Journal of Human Genetics 61 1036–1043. cancer through loss of expression. PNAS 95 5246–5250. Wick W, Furnari FB, Naumann U, Cavenee WK & Weller M 1999 Tsou HC, Ping XL, Xie XX, Gruener AC, Zhang H, Nini R, PTEN gene transfer in human malignant glioma: sensitization to Swisshelm K, Sybert V, Diamond TM, Sutphen R & Peacocke irradiation and CD95L-induced apoptosis. Oncogene 18 3936– M 1998 The genetic basis of Cowden’s syndrome: three novel 3943. mutations in PTEN/MMAC1/TEP1. Human Genetics 102 467– Wu X, Hepner K, Castelino-Prabhu S, Do D, Kaye MB, Yuan XJ, 473. Wood J, Ross C, Sawyers CL & Whang YE 2000 Evidence for Ueda K, Nishijima M, Inui H, Watatani M, Yayoi E, Okamura J, regulation of the PTEN tumor suppressor by a membrane- Yasutomi M, Nakamura Y & Miyoshi Y 1998 Infrequent localized multi-PDZ domain containing scaffold protein mutations in the PTEN/MMAC1 gene among primary breast MAGI-2. PNAS 97 4233–4238. cancers. Japanese Journal of Cancer Research 89 17–21. Yokomizo A, Tindall DJ, Hartmann L, Jenkins RB, Smith DI & Vanhaesebroeck B, Leevers SJ, Panayotou G & Waterfield MD Liu W 1998 Mutation analysis of the putative tumor suppressor 1997 Phosphoinositide 3-kinases: a conserved family of signal PTEN/MMAC1 in human . International Journal transducers. Trends in Biochemical Sciences 22 267–272. of 13 101–105. Wang SI, Puc J, Li J, Bruce JN, Cairns P, Sidransky D & Parsons Zigman AF, Lavine JE, Jones MC, Boland CR & Carethers JM R 1997 Somatic mutations of PTEN in glioblastoma multiforme. 1997 Localization of the Bannayan-Riley-Ruvalcaba syndrome Cancer Research 57 4183–4186. gene to chromosome 10q23. Gastroenterology 113 1433–1437.

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