Epithelial cell polarity, stem cells and cancer

The development of solid tumours comprises a complex succession of intrinsic and extrinsic events that prompt cells to proliferate out of control and to acquire migra-tory and invasive capabilities1. Whereas several studies have undoubtedly revealed that the loss of epithelial cell polarity is intricately connected to the malignant progres-sion of incipient cancer cells, accumulating evidence has unmasked the crucial roles of regulators of polarity in the early stages of tumorigenesis. Epithelial cell polar-ity is fundamental for tissue function, and it has been conventionally defined as ‘asymmetry’ within cells and epithelial tissues2. This denotes not only a differential positioning of membrane domains and organelles along the apical-basolateral axis, known as apical-basolateral polarity, but also the positioning of cells in the plane of epithelial tissues, known as planar cell polarity3. In this Review, we focus on the mechanisms that regulate the apical-basolateral polarity of epithelial cells, which more clearly contributes to cancer.

Apical-basolateral polarity is characterized by the existence of non-coalescent apical and basolateral plasma membrane domains, which have a differential composition of proteins and lipids. Both simple and stratified epithelia comprise a single layer of divid-ing cells in which the basal domain is attached to the underlying basement membrane. In simple epithelia, such as the intestinal lining, the apical membrane faces the luminal space, and it is distinguished by the presence of microvilli and a primary cilium4. In stratified epithelia, such as the epidermis, the uppermost layers face the external environment and function as a barrier,

although apical and basolateral determinants have been identified in progenitor cells of the basal layer of the epidermis5.

Apical-basolateral polarity contributes to the acqui-sition of cell shape and to the directional transport that characterizes epithelial function, but how is the loss of epithelial polarity associated with tumorigenesis? Cell polarization allows cells to sense and to elicit the proper spatiotemporal responses to cues that arise from neigh-bouring cells and the surrounding microenvironment2. Thus, when polarity is disrupted, cells may become unre-sponsive to growth inhibitory signals and may circumvent differentiation, senescence or apoptosis. Interestingly, sev-eral proteins that regulate apical-basolateral polarity are well-known tumour suppressors and proto-oncoproteins. They also exhibit crosstalk with signalling pathways that control cell growth and proliferation, including WNT, Hedgehog and Hippo pathways, and mTOR-dependent energy metabolism. Additionally, through their connec-tions with the cytoskeleton, their loss is fundamentally associated with alterations in epithelial functions, such as loss of vectorial transport and tissue architecture, and mitotic defects that may lead to the generation of genomic instability. Interestingly, during mitosis, cell polarity mechanisms are also involved in the control of oriented divisions of epithelial stem cells within the tissues. This is particularly important, as evidence suggests that stem cells can be the cell-of-origin for a range of solid tumours.

In this Review, we highlight advances in understand-ing the establishment and maintenance of epithelial cell polarity and the associated implications for cancer. We

1Centro de Biología Molecular Severo Ochoa, Consejo Superior de Investigaciones Científicas, Madrid 28049, Spain.2Epithelial Cell Biology Group, Cancer Cell Biology Programme, Centro Nacional de Investigaciones Oncológicas, Madrid 28019, Spain.e‑mails: [email protected]; [email protected]:10.1038/nrc3169Published online 15 December 2011

Cell polarityA fundamental feature of many types of cells that describes the asymmetrical distribution of its components within a cell.

MicrovilliSmall plasma membrane protrusions on the surface of cells that increase the surface area and facilitate absorption and secretion.

Primary ciliumIn mammalian cells, a specialized protrusion with sensory functions.

Epithelial cell polarity, stem cells and cancerFernando Martin‑Belmonte1 and Mirna Perez‑Moreno2

Abstract | After years of extensive scientific discovery much has been learned about the networks that regulate epithelial homeostasis. Loss of expression or functional activity of cell adhesion and cell polarity proteins (including the PAR, crumbs (CRB) and scribble (SCRIB) complexes) is intricately related to advanced stages of tumour progression and invasiveness. But the key roles of these proteins in crosstalk with the Hippo and liver kinase B1 (LKB1)–AMPK pathways and in epithelial function and proliferation indicate that they may also be associated with the early stages of tumorigenesis. For example, deregulation of adhesion and polarity proteins can cause misoriented cell divisions and increased self-renewal of adult epithelial stem cells. In this Review, we highlight some advances in the understanding of how loss of epithelial cell polarity contributes to tumorigenesis.

R E V I E W S

NATURE REVIEWS | CANCER VOLUME 12 | JANUARY 2012 | 23

© 2012 Macmillan Publishers Limited. All rights reserved

mailto:[email protected]

mailto:[email protected]

Vectorial transportThe transportation of ions or molecules within a polarized cell across an epithelium in a specific direction.

Tight junctionsA type of intercellular junction, which is comprised of several interconnecting integral plasma membrane proteins that are anchored to the cytoskeleton. They link adjacent plasma membranes of neighbouring cells to limit the movement of proteins and lipids between the apical and lateral plasma membrane domains and the intercellular passage of molecules.

Cell cortexA cytoplasmic region under the inner face of the plasma membrane that forms a contractile, mesh-like structure that is rich in contractile actin-myosin filaments (termed cortical actin filaments) and spectrin filaments.

summarize the mechanisms that promote the establish-ment of epithelial cell polarity, and discuss cell polarity proteins and their links with different molecular path-ways that result in the loss of cell polarity and spindle misorientation in epithelia.

The establishment of epithelial cell polarityThe mechanisms that promote the establishment of epi-thelial cell polarity are not completely understood, and can vary between different mammalian epithelial cells and model organisms. In simple epithelia, the formation of polarized cohesive cell layers that form the body compart-ments mainly depends on cell adhesion complexes and epithelial cell polarity complexes. Two intercellular adhe-sion complexes, known as adherens junctions and tight junctions, form part of the apical junctional complex and contribute to the formation and the maintenance of apical and basolateral domains6,7 (BOX 1). This differs in neuro-`epithelial cells, which do not have classical tight junctions, and in stratified epithelia, such as the epidermis, in which tight junctions are maintained in the upper granular layer. At the root of the establishment of the polarity of simple epithelial cells lies a finely tuned interplay between adhe-rens junctions, tight junctions and cell polarity complexes; and in neuroepithelial cells and the epidermis, some parallels of these molecular interactions also take place.

The cell polarity complexes, which were originally identified in model organisms such as yeast, worms and flies, are highly evolutionarily conserved8. Three major

polarity complexes have been identified to date: the PAR polarity complex, comprising PAR3, PAR6, atypi-cal protein kinase C (aPKC) and cell division control protein 42 (CDC42), which promotes the establish-ment of the apical-basal membrane border; the crumbs (CRB) complex, which is formed by the transmembrane protein CRB and the associated cytoplasmic proteins PALS1 (also known as MPP5) and PALS1-associated tight junction protein (PATJ; also known as INADL), which is required to establish the apical membrane; and the scribble homologue (SCRIB)–lethal (2) giant lar-vae homologue (LGL; also known as LLGL)–discs-large homologue (DLG) complex, which defines the basolateral plasma domain9 (FIG. 1).

Protein networks that establish the apical and basolateral domains. Epithelial cells orient their basal surface through the adhesion of integrin receptors to the extra-cellular matrix, and they extend membrane protrusions, known as filopodia, to contact neighbouring cells and so allow the formation of cohesive layers of cells10. At the ini-tial stages of intercellular adhesion, nectin–afadin adhe-sion complexes associate with PAR3 (REFS 11,12) (FIG. 1). This is followed by the recruitment of E-cadherin and junctional adhesion molecule A (JAMA) to the cell cor-tex, which cluster these primordial adhesion sites, known as puncta13,14, into adhesive homophilic interactions. These primordial adhesions contain a mixture of adherens junction and tight junction proteins12,15, and through their anchoring to the actin cytoskeleton and through RHO-GTPase activity, they extend the interface of adhesion along the basolateral domain16,17.

Next, adherens junction-associated proteins are sep-arated from tight junction-associated proteins and the adherens junction belt-like structure and mature tight junctions are formed. This requires the exclusion of PAR3 from primordial adhesions through the localized acti-vation of RAC1 GTPase. Interestingly, PAR3 binds and recruits to these sites T lymphoma invasion and metas-tasis-inducing protein 1 (TIAM1; a RAC1 GTPase gua-nine nucleotide exchange factor (GEF)), which promotes the exclusion of TIAM1 from the subapical sites where it modulates RAC1 activity18,19. aPKC (PKCζ and PKCι in humans) is then recruited to subapical PAR3–PAR6 complexes, and phosphorylates and excludes PAR3 from tight junction sites, allowing tight junction for-mation and the separation of the apical and subapical domains20,21. In Drosophila melanogaster, the separa-tion of the apical domain from the most apical region of the basolateral domain presents many similarities to the polarization of mammalian cells22–25.

The SCRIB–LGL–DLG complex localizes with adhe-rens junctions in mammals and with septate junctions in D. melanogaster and promotes basolateral membrane identity26–28. This complex controls the expansion of the apical domain; LGL competes with PAR3, sequestering the PAR complex from the apical membrane. LGL phos-phorylation by aPKC and the phosphorylation of PAR3 by PAR1 (also known as MARK2) inhibit this interac-tion and promote the separation of the lateral domains and subapical domains29–31. The apical membrane is

At a glance

•Theproteinsthatfinelycontrolepithelialcellpolarityareknownastumoursuppressorsorproto-oncoproteins.Lossofepithelialcellpolarity—throughderegulationoftheseproteins—iscrucialforcancercellinvasionandadvancedtumourprogression.Accumulatingevidenceindicatesthatepithelialcellpolaritycuesandpolarizedcelldivisionscausallycontributetorestrictcarcinomaformation.

•Cellpolarityproteinscrosstalkwithsignallingpathwaysthatregulatecellgrowthandproliferation,includingtheWNTandHippopathways,andliverkinaseB1(LKB1)–mTOR-dependentenergymetabolism.

•ThecomponentsoftheevolutionarilyconservedHippopathwayfunctionasimportanttumoursuppressors.Recentstudieshaveprovidedevidencethatthecellpolarityregulatorslethal(2)giantlarvaehomologue(LGL;alsoknownasLLGL),atypicalproteinkinaseC(aPKC)andcrumbshomologue(CRB),andtheadherensjunctionscomponentsE-cadherin–α-cateninorE-cadherin–β-cateninregulatetheHippopathwayinmammalianandDrosophila melanogasterepithelial cells.

•TheLKB1–AMPK–mTORpathway,whichisamolecularlinkbetweenpolarityandthemetabolicstatusofacell,isessentialintheprocessoftumorigenesis.

•Themaintenanceofmostadultepithelialtissuesreliesonthepresenceofpolarizedstemcells,whichself-renewthroughsymmetriccelldivisions.Duringdifferentiationstemcellsreorienttheirmitoticspindlesanddivideasymmetricallyinordertogeneratethespecializedcellsthatdriveepithelialfunctionandhomeostasis.Thegenesthatcontrolepithelialcellpolarityalsoregulatespindleorientationandthesymmetryofcelldivisionsinstem cells.

•Epithelialtumoursarehighlyheterogeneous,andthecell-of-originthatcaninitiatetumorigenesisisanareaofextensivestudy.Twotheoriesoftumourinitiationhavebeenpostulated;oneproposesthatsometumoursarisefromnormaladultstemorprogenitorcellsthathavegoneawry,andtheotherpostulatesthattheyarisefromdifferentiatedcellsthatacquireself-renewalcapabilities.Evidenceforthestemorprogenitorcell-of-originmodelhasbeenprovidedforsomecarcinomasorspecificsubtypesofcarcinomas;however,thisseemstobeunlikelyorhasnotbeenwelldefinedforseveral others.

R E V I E W S

24 | JANUARY 2012 | VOLUME 12 www.nature.com/reviews/cancer


Nature Reviews | Cancer

Integrins

Nucleus

F-actin

Microtubule

Golgi

Microvilli

Primary cilium

Centrosome

Tight junction

Late

ral

Basa

lA

pica

l

Adherens junction

Nectin

Cadherin

Occludin

ClaudinJAM

Dynein

APC

Actin BP

Actin BP

α-cat

α-cat α-catβ-cat β-cat

α-cat

Spectrin filaments

p120 p120

+

+

+

+

––

–

–

?

Luminal space

Afadin Afadin

ZO2ZO3ZO1

ZO3ZO2ZO1

Basement membrane

Adhesive homophilic interactionsAdhesive interactions in which the interacting molecules are of the same type. Also known as homotypic interactions.

SubapicalA plasma membrane domain that is spatially separated between the apical and basolateral domains. It is thought to coordinate molecular pathways, such as endocytic pathways and sorting machinery between these distinct plasma membrane domains.

Septate junctionsIntercellular junctions of invertebrate epithelia that act as a barrier restricting the diffusion of proteins and lipids between the apical and lateral plasma membrane domains and the intercellular passage of molecules.

characterized by the presence of microvilli, in which there is a meshwork of actin and spectrin filaments linked to the membrane by the ezrin, radixin and moesin (ERM) family of proteins32. The tumour suppressor merlin (also known as NF2) has homology to the ERM family members. Merlin has also been implicated in pro-moting the establishment of adherens junctions and the development of the epidermis33. Additionally, merlin associates with tight junctions through its interaction with angiomotin (AMOT)–PATJ and PALS1, which provides junctional stability34.

The continued expression and functional activity of adherens junctions, tight junctions, merlin and cell polarity complexes are required for polarized cells to remain tightly associated within the epithelium and to coordinate signalling pathways that regulate epithe-lial proliferation. The loss of constituents of adherens junctions and tight junctions, as well as merlin, is asso-ciated with depolarization, loss of differentiated char-acteristics, enhanced epithelial cell proliferation and the acquisition of invasive potential34–39. Furthermore, loss of cell polarity complexes leads to intercellular adhesion breakage, loss of polarized characteristics and hyperproliferation9,40,41. This can occur through various alterations, including overexpression, downregulation, mislocalization, deletion and alternative splicing42. In addition, several key transcription factors that promote

epithelial-to-mesenchymal transition (EMT) directly regu-late the expression of both cell adhesion and cell polarity complex proteins43.

The polarity complexes in cancerIn humans, SCRIB is a potential tumour suppressor, the expression of which is frequently lost in advanced tumours43 (TABLE 1; see Supplementary information S1 (table)). SCRIB is also frequently mislocalized owing to the loss of E-cadherin, which is required for the locali-zation of SCRIB at the basolateral plasma membrane28. Similarly, loss-of-function mutations in Scrib in eye-antennal discs in D. melanogaster lead to the forma-tion of metastatic tumours through a mechanism that involves the oncoprotein RAS. These tumours also have activated JUN N-terminal kinase (JNK) signalling, and present similar characteristics to human cancers that lack SCRIB, including basement membrane degradation, loss of E-cadherin expression, and migration, invasion and secondary tumour formation44,45. Whereas JNK activity normally promotes the apoptosis of SCRIB-deficient cells, it becomes a driver of cellular overgrowth, tumorigenesis and invasion in the presence of oncogenically activated RAS or Notch signalling46. Recently, in vivo and in vitro findings have shown that the loss of SCRIB inhibits apoptosis and enhances transformation through the MYC proto-oncoprotein in three-dimensional (3D) cultures of

Box 1 | The biology of cellular junctions and their role in cell polarity

Adherensjunctionsandtightjunctionsparticipateintheestablishmentandmaintenanceofapical-basalpolarityinsimpleepithelia.Themajorstructuralcomponentsofadherensjunctionsandtightjunctionsareshowninthefigure.Atthecoreofadherensjunctionsaretwoadhesivecomplexes:cadherin–cateninandnectin–afadincomplexes183.Classicalcadherinsmediatehomophiliccalcium-dependentcell–celladhesionsthroughtheirextracellulardomains,whichlinkcellstoeachother.Thecytoplasmictailsofcadherinsbind,throughdifferentdomains,toβ-catenin(β-cat)andtop120-catenin(p120).Inturn,β-cateninbindstomonomericα-catenin(α-cat)thatindirectlyanchorsthecadherin–catenincomplexestotheactincytoskeletonthroughinteractionswithactin-bindingproteins(actinBPs).Theα-catenindimerpreferentiallybindsactinfilaments184.Additionally,β-cateninbindstothemicrotubulemotordynein168,andp120-cateninconnectscadherin–catenincomplexestomicrotubules174.Thecytoplasmicdomainofnectininteractswithafadin(alsoknownasAF6),whichbindstoactinandRAS-andRAP-familyGTPases(notshown).Thecytoplasmicdomainofnectinalsointeractswithotheractin-bindingproteinssuchasthetightjunctionproteinzonulaoccludens1(ZO1)andα-catenin.Tightjunctionslocalizeapicallytoadherensjunctions,anddistributeattheborderoftheapicalandbasolateraldomain,wheretheyfunctiontorestrictthemovementofproteinsandlipidsbetweenthesemembranedomains.Thesejunctionsalsofunctionasabarrierbydynamicallysealingthespacebetweenneighbouringcells185.Thetightjunctiontransmembraneproteinsoccludin,claudin,tricellulin(notshown)andjunctionaladhesionmolecule(JAM)bindthroughtheircytoplasmicdomaintoseveralintracellularscaffoldingproteins,includingZO1–3,multi-PDZdomainprotein1(MUPP1)andcingulin(notshown)186.

R E V I E W S



http://www.nature.com/nrc/journal/v12/n1/suppinfo/nrc3169.html


Integrins

Nucleus

F-actin

Tight junction

Late

ral

Basa

lA

pica

l

Adherens junction

Nectin

Cadherin

Occludin

ClaudinJAM

Actin BPα-cat α-cat

β-cat β-cat

Spectrin filaments

p120

PAR3PAR3

PAR3

PAR3

PAR3

RAC1

RHOA

PAR3PAR3

PAR1

CRBPALSPATJ

PAR6

PAR6 aPKC

PAR6CDC42 CDC42aPKC

LGL

aPKC

p120

Cadherin

Actin BP

Actin BP

α-cat α-catβ-cat β-cat

p120

Luminal space

Afadin

SCRIBDLG

SCRIBDLG

Afadin

NectinAfadin Afadin

ZO2ZO3ZO1

ZO1

ZO2ZO3ZO1

Basement membrane

CRBPALSPATJ

Intermediate stage

Polarized cell

CRBPALSPATJ

Cell polaritycomplexes in a polarized cell

Initial stage ofcell polarization

ARP2/3

Primordialcell contacts

JAM

WASP

Lateral domainsPlasma membrane domains that face neighbouring cells.

Spectrin filamentsA meshwork of filaments comprised of the cytoskeletal protein spectrin, which has a role in the maintenance of plasma membrane integrity and cytoskeletal structure.

Epithelial-to-mesenchymal transition(EMT). A process through which cells transit from a polarized, epithelial phenotype to a highly motile mesenchymal phenotype. EMT causes pronounced morphological and functional changes in cells, such as loss of epithelial adherens junctions, reorganization of the actin cytoskeleton and loss of apical-basal polarity.

human epithelial cells47. This involves the interaction at cellular junctions of SCRIB with the β-PIX (also known as ARHGEF7)–GIT1 complex, which is a GEF for RAC1, the expression of which is induced by MYC. In turn, RAC1 activation stimulates a JNK–JUN–BCL-2-interacting mediator of cell death (BIM; also known as BCL2L11) sig-nalling pathway. Therefore, reductions in the expression or mislocalization of SCRIB are able to block apoptosis and stimulate tumorigenesis. Finally, SCRIB is down-regulated or mislocalized in human breast tumours and transgenic mouse mammary tumours47. The deregulation of SCRIB in mammary epithelial cells disrupts cell polar-ity, blocks apoptosis and drives the formation of breast tumours when MYC is oncogenically activated47.

Interestingly, the role of SCRIB in tumorigenesis seems to be dependent on cell context: in some cases it induces cell migration through the activation of RAC1, whereas in others it has been associated with restricting invasion and metastasis48,49. The molecu-lar mechanisms that regulate these contrary func-tions are starting to be elucidated. It has been shown that SCRIB restricts HRAS-G12V-mediated invasive-ness of a human breast epithelial cell line through the direct binding of ERK and the consequent inhibition of its phosphorylation and thereby the inhibition of the RAF–MEK–ERK pathway50. Indeed, loss of SCRIB results in increased levels and nuclear translocation of phospho-ERK51.

Figure 1 | Establishment of epithelial cell polarity. Three major polarity complexes participate in the establishment of apical-basal polarity: the crumbs (CRB) complex (shown in red) is required to establish the apical membrane and is formed of the transmembrane protein CRB and the associated cytoplasmic proteins PALS1 and PALS1-associated tight junction protein (PATJ). The PAR complex (shown in blue) promotes the establishment of the apical-lateral membrane border and comprises PAR3, PAR6, atypical protein kinase C (aPKC; PKCζ and PKCι in humans) and cell division control 42 (CDC42). The scribble (SCRIB)–lethal (2) giant larvae homologue (LGL)–discs large homologue (DLG) complex (shown in orange) defines the basolateral plasma membrane domain20,187–191. These three complexes have antagonistic interactions and spatiotemporally regulate epithelial polarization through their interactions with the cytoskeleton and adhesion proteins. At early stages of cell polarization, PAR3 binds to afadin, and these primordial adhesions mature to form the belt-like adhesion junctions and tight junctions that localize at the apical-basal membrane border. This involves aPKC activity and the exclusion of PAR3 from primordial adhesions at intermediate stages of polarization. This is followed by PAR3 exclusion from both the PAR6–aPKC and CRB complexes to establish the apical-lateral border and the apical membrane; whereas, the SCRIB complex defines the basolateral plasma membrane domain by antagonizing the PAR and CRB complexes, which restricts their activity and hence the expansion of the apical domain9,41,188,192. Dashed arrows represent translocations to different subcellular locations. α-cat, α-catenin; actin BP, actin-binding protein; ARP, actin-related protein; β-cat, β-catenin; JAM, junctional adhesion molecule; p120, p120-catenin; WASP, Wiskott–Aldrich syndrome protein; ZO, zonula occludens protein.

R E V I E W S



Table 1 | Alterations of polarity complex proteins in epithelial transformation and human cancer*

Gene (protein) Alterations Cancer type Phenotypes

Crumbs complex

CRB3 (crumbs 3) Downregulated expression Human tumour epithelial-derived cell lines

Cell–cell junctions disrupted and increased metastasis

PAR complex

PARD3 (PAR3) Gene deleted or downregulated expression

Oesophageal squamous cell carcinoma cell lines and primary tumour tissue

Cell–cell junctions disrupted

PARD6A (PAR6α) Overexpressed ER-positive breast cancer cell lines and primary tumour tissue

Hyperproliferation

Overexpressed and phosphorylated Human BRCA1-defective tumour tissues

Lumen filling, cell–cell junctions disrupted and increased metastasis

Overexpressed Stromal cells in non-small-cell lung cancer tissue samples

Associated with good prognosis

PRKCZ (PKCz or aPKC)

Overexpressed Human hepatocellular carcinoma samples

Hyperproliferation

Overexpressed Bladder tumour cell lines and primary tumour tissues

Correlated with invasiveness

Overexpressed and phosphorylated Dysplastic oral epithelial tissue samples, squamous cell carcinoma of the head and neck tissue samples and cell lines

Increased cell proliferation

Overexpressed Pancreatic cancer tissues Invasive and metastatic phenotype

Overexpressed Samples of hyperplastic enlarged lobular units of precancerous breast lesion

Increased cell proliferation

PRKCI (PKCi or aPKC)

Gene amplified and protein overexpressed and mislocalized

Ovarian cancer tissue samples Associated with low survival rate

Overexpressed and phosphorylated Hepatocellular carcinoma tissue samples

Associated with metastasis and invasion

Overexpressed Non-small-cell lung cancer cell lines and primary tumour tissues

Associated with poor prognosis

Overexpressed Primary breast cancer tissue samples Associated with larger tumours, invasion and metastasis

Overexpressed Pancreatic cancer tissue samples Tumour angiogenesis and metastasis

SCRIB complex

SCRIB (scribble) Mislocalized or downregulated High-grade HPV-positive cervical squamous intraepithelial lesions and invasive cervical carcinoma samples

Correlates with invasiveness

Mislocalized or downregulated Neoplastic colon mucosa Loss of tissue architecture

Mislocalized or downregulated Human breast cancer tissue samples Loss of three-dimensional cell polarity and inhibition of apoptosis

DLG1 (DLG) Mislocalized or downregulated High-grade uterine cervical neoplasm samples

Role in cytokinesis, viral trafficking and metastasis pathways

Mislocalized or downregulated Neoplastic colon mucosa samples Loss of tissue architecture

LLGL1 (LGL1) Downregulated Tissue samples from breast, prostate, lung and ovarian tumours

Disruption of cell polarity and tissue architecture, uncontrolled proliferation and growth of neoplastic lesions

Downregulated Colon cancer cell lines and colon cancer primary tissue samples

Associated with advanced stage and lymph node metastases

Aberrantly spliced mRNA and the expression of truncated protein

Hepatocellular carcinoma cell line and primary tissues

Associated with poor differentiation and large tumour size

Mislocalized or downregulated Samples of human gastric epithelial dysplasia and adenocarcinoma

Disruption of tissue morphology

aPKC, atypical protein kinase C; DLG, discs large homologue; ER, oestrogen receptor; HPV, human papilloma virus; LGL, lethal (2) giant larvae homologue; LLGL1, lethal (2) giant larvae homologue 1; PKC, protein kinase C. Table is modified, with permission, from REF. 42 © (2010) Elsevier. *See Supplementary information S1 (table) for a version of this table with references.

R E V I E W S



The other components of the SCRIB complexes, LGL and DLG, are also tumour suppressors that promote the acquisition of invasiveness when their expression is modified. The EMT-associated transcription factor zinc finger E-box-binding homeobox 1 (ZEB1) represses the expression of several polarity proteins, including LGL2, in breast and colorectal cancers, which is associ-ated with a substantial reduction of cell polarity and the induction of metastasis52,53. Similarly, the re-expression of LGL2 in melanoma cells restored cell adhesion and restricted invasion54. A recent study in zebrafish showed that Lgl2 regulates Erbb signalling in epidermal carci-nomas. Loss of Lgl2 induced the activation of Erbb sig-nalling, impaired the formation of hemidesmosomes and induced the mislocalization of E-cadherin, leading to the induction of EMT; pathways involved in pro-liferation were also activated55. Another mechanism associated with LGL loss-of-function in oncogenesis is alternative splicing. Aberrant splice variants of LLGL1 (which encodes LGL1) have been described in hepato-cellular carcinoma56. The majority of these transcripts were found to encode truncated proteins lacking one or more conserved WD-40 repeat motifs and which were therefore unlikely to be functional.

The CRB tumour suppressor was originally identi-fied in D. melanogaster as a regulator of epithelial cell polarity. CRB3, the human orthologue of CRB, also has a fundamental function in the establishment of polarity in mammalian epithelial cells, and in the morphogen-esis of 3D cultures of epithelial cells. CRB3 is a tumour suppressor during the transformation and progression of mammalian epithelial cells57. Further evidence was obtained from observations of immortal embryonic mouse kidney epithelial cells selected in vivo to have acquired tumorigenic capabilities. CRB3 expression was substantially repressed in these cells, which had impaired tight junction formation and loss of apical-basal polarity and contact inhibition. Re-expression of CRB3 restrained cell migration and metastatic potential in vivo57. The molecular mechanisms by which CRB3 exerts its tumour suppressor functions may be related to its interactions with the Hippo pathway (discussed below).

Mutations in members of the PAR complex have not been directly associated with tumour formation or progression. However, they seem to cooperate with oncogenes or tumour suppressors in different epithelial cancers (TABLE 1; see Supplementary information S1 (table)). Hence, some signalling pathways that are usu-ally modified in cancer have been found to regulate cell polarity, apoptosis and growth in a coordinated man-ner40,58. ERBB2, a member of the epidermal growth factor receptor (EGFR) family that is localized in the basolateral membrane, is involved in cell proliferation during normal development in different tissues and wound healing59. However, ERBB2 overexpression, which causes partial mislocalization of ERBB2 to the apical domain, is observed in different epithelial can-cers42. Apically localized ERBB2 binds to PAR6–aPKC and sequesters this complex away from PAR3, which disrupts PAR complex formation at tight junctions,

compromises tight junction integrity and thus disrupts apical-basal polarity60. However, recent studies have shown that the expression of PAR6C (also known as PAR6α) alone induces cell proliferation and the devel-opment of hyperplastic 3D acini in mammary epithelial cells independently of EGF and through the activation of the MEK–ERK signalling pathway, and without affecting apical-basal polarity61. This could be explained by previ-ous studies that have shown that transforming growth factor-β (TGFβ) signals to the PAR complex to con-trol epithelial cell morphology and transformation62,63. TGFβ is a key regulator of EMT in tumour formation64, which promotes invasion and metastasis in advanced carcinomas58.

Accumulating evidence demonstrates that human aPKC (PKCζ and PKCι) are oncoproteins the expres-sion of which is frequently elevated in several human epi thelial cancers, including lung, liver, breast, pancreas, ovarian, prostate and colon, which usually correlates with poor prognosis65. On a mechanistic level, it has been demonstrated that aPKC functions downstream of RAS signalling and upstream of RAC1, in lung and colon carcinogenesis66,67; aPKC in D. melanogaster tumour models signals through the JNK pathway68.

In summary, polarity complexes do exhibit altera-tions in cancer that drive tumorigenesis but they are predominantly associated with tumour progression. However, crosstalk between polarity complexes and other signalling pathways seems to drive tumorigenesis (TABLE 2; see Supplementary information S2 (table)).

The Hippo tumour suppressor pathwayThe components of the Hippo pathway function as important tumour suppressors69,70 that regulate tissue growth by promoting cell cycle exit and apoptosis71,72. Recent data from mammals and D. melanogaster have linked this pathway with cell polarity proteins in the regulation of tissue growth73–76, limiting organ size77 and participating in tissue regeneration78–83 (FIG. 2).

Regarding the link between cell polarity, the Hippo pathway and cancer, recent studies in the developing D. melanogaster eye and imaginal disc epithelia have shown that LGL, aPKC and CRB regulate proliferation and survival by controlling the activity of the Hippo pathway74,75,84,85 (FIG. 2). These findings may explain why alterations in polarity complex proteins are linked to pro-liferation and overgrowth, which has also been observed in mammals. The CRB polarity complex interacts with the transcription factors yes-associated protein 1 (YAP1) and TAZ (also known as WWTR1), which are the mam-malian homologues of Yorkie in the D. melanogaster Hippo pathway. CRB in turn transmits cell density infor-mation, which is transmitted through the localization of CRB to the apical domain upon acquisition of polarized junctions and membrane domains, by promoting YAP1–TAZ phosphorylation and consequently cytoplasmic retention, which suppresses TGFβ signalling. TAZ func-tions as a SMAD nuclear retention factor, and the Hippo pathway links TAZ–YAP1 regulation to cell density, rais-ing the possibility that TGFβ–SMAD signalling might be coupled to cell density-sensing through the Hippo

R E V I E W S




pathway. Disruption of the CRB complex enhances TGFβ signalling and predisposes cells to TGFβ-mediated EMT76, and therefore to late-stage transformation. In addition, the E-cadherin–catenin adhesion complex functions as an upstream regulator of the Hippo signal-ling pathway in mammalian tissues and cells through the control of YAP1–TAZ activity and localization86–88.

In human cancers, the most prevalent mechanism for Hippo pathway inactivation is epigenetic silencing70. In fact, hypermethylation of RASSF (RASSF1A and RASSF2A), the human HPO homologues STK4 (which encodes MST1), STK3 (which encodes MST2) and the human Warts homologues LATS1 and LATS2 (REFS 89–91) has been found in various different epithelial cancers.

Table 2 | The role of cell polarity pathways in epithelial transformation and human cancer*

Gene (protein if different from gene name)

Alteration in cancer

Cancer types Phenotype

Energy metabolism

STK11 (LKB1) Germline mutation Gastrointestinal, pancreatic, ovarian and breast cancers

Germline alterations cause PJS, which is associated with the formation of hamartomatous polyps in the gastrointestinal tract and increased risk of developing cancer

Gene mutated Lung and cervical carcinomas Sporadic tumours

The Hippo pathway

RASSF1 Hypermethylation Epithelial cancers Defects in apoptosis, cell cycle control and microtubule stabilization

STK3 (MST2) and STK4 (MST1)

Reduced gene expression

Colorectal and prostate cancers

Hormone-resistant metastases and poor prognosis

LATS1 and LATS2 Hypermethylation Epithelial cancers Aggressive phenotype

YAP1 Overexpression Epithelial carcinomas Anchorage-independent growth and metastasis

YAP1, WWTR1 (TAZ) and TEAD4

Amplification and overexpression

Liver, lung, pancreatic, oesophageal, oral squamous cell and breast cancer

Proliferation and increased organ size

NF2 (merlin) Loss of merlin expression

Central nervous system and skin tumours

Randomized spindle orientation of basal progenitor cells, reduced tight junction formation and overall defects in epithelial differentiation

Loss of merlin expression

Epithelial and Schwann cell cancers

Hyperproliferation

Protein trafficking

RAB25 Overexpressed Breast and ovarian cancer Associated with aggressive behaviour

Decreased expression Human colon cancer samples Functions as a tumour suppressor for colonic neoplasia

Asymmetric division and/or spindle orientation

CDH1 (E-cadherin) Germline inactivating mutations

Familial gastric cancer Correlates with tumour grade and metastasis

Loss-of-function Epithelial adenomas and carcinomas

Alterations in cell–cell adhesion, tumour grade and metastasis

VHL Loss-of-function or promoter methylation

Sporadic kidney cancers Loss of primary cilia and cell polarity spindle misorientation, spindle checkpoint weakening and chromosomal instability

APC Inactivating mutations or loss-of-function

Gastrointestinal and mammary gland cancers

Deregulation of cell polarity, motility and cell migration; favours symmetric cell divisions. Associated with cancer progression

CTNND1 (p120-catenin) Overexpression Human colon adenocarcinoma cells (HT-29)

Aberrant mitosis and polyploidy

Loss-of-function Skin Mitotic alterations and centrosome abnormalities

CLDN1 (claudin 1) Reduced expression Colorectal cancer Impairs regulation of spindle orientation

Loss-of-function or inactivating mutations

Breast cancer Claudin low subtype is associated with loss of cell adhesion and tumours are enriched for mesenchymal and stem cell features

CTNNB1 (β-catenin) Loss-of-function HCT116 human colon cancer cell lines

Alterations in cell architecture and polarity

Loss-of-function Small colorectal adenomas Associated with loss of cell adhesion

CTNNA1 (a-catenin) Loss-of-function Skin Hyperproliferation, impairs spindle reorientation

APC, adenomatous polyposis coli; LKB1, liver kinase B1; MST, mammalian STE20-like protein kinase; NF2, neurofibromin 2; PJS, Peutz–Jeghers syndrome; STK11, serine/threonine kinase 11; YAP1, yes-associated protein 1. *See Supplementary information S2 (table) for a version of this table with references.

R E V I E W S




Late

ral m

embr

ane

Apical membrane

PAR6 aPKC PAR6 aPKC

LGL DLG

SAV

SDT

Kibra

FAT SCRIB

EX

HPO

CRB

PATJ

FBD

TJ TJ

AJAJ

PAR3 PAR3

MER

MER

RASSF

PDZP

P

P

Nucleus

MATSWTS

YKI

P

14-3-314-3-3YKI

P

14-3-3YKI DM IAP2 Cyclin E

SD

By contrast, the association of mutations in Hippo pathway genes with cancer is less frequent, except for loss-of-function mutations in NF2 (which encodes merlin). Merlin is a FERM domain-containing protein the loss of which results in defective morphogenesis and tumorigenesis in multiple tissues. Mutations in NF2 also cause neurofibromatosis type II, a disease that is characterized by the development of benign Schwann cell-derived tumours92. Merlin is an essen-tial mediator of contact-dependent inhibition of pro-liferation, and is required for the establishment of stable adherens junctions in cultured cells93. Recent data have shown that merlin can directly associate with α-catenin and can connect α-catenin to PAR3, providing a link between adherens junctions and the PAR3 polarity complex33, and therefore representing

a potential new mechanism of crosstalk between cell polarity complexes and the Hippo pathway.

Merlin function is associated with various signal-ling pathways, including RAS, RAC1 and PI3K94. In fact, merlin-organized complexes prevent mitogenic signalling and mediate contact-dependent inhibition of proliferation, although the connections between merlin and these pathways with growth control and oncogenic transformation needs further investiga-tion. It was recently shown that the depletion of Nf2 induced hepatocellular carcinoma and bile duct tumours in mice39,95. Loss-of-function mutations in the core Hippo pathway also produced tumours spe-cifically in these two organs96,97, demonstrating the crosstalk between merlin and the Hippo pathway in tumorigenesis. Additionally, phenotypes in multiple tissues caused by merlin deficiency were largely sup-pressed by heterozygous deletion of Yap1, suggest-ing that YAP1 is a major effector of merlin in growth regulation95. In conclusion, emerging evidence shows that the Hippo pathway is regulated by cell polarity, cell adhesion and cell junction proteins. Hence, cell polarity and cell adhesion proteins seem to modu-late organ size control and regeneration through the Hippo pathway.

The tumour suppressor LKB1 in cell polarityCancer cells have limited access to metabolites for energy production, and the involvement of signalling pathways that regulate energy production is crucial in tumorigenesis, as it is during development. Because the liver kinase B1 (LKB1; also known as STK11) path-way is a molecular link that connects polarity and the metabolic status of a cell, it has been proposed that this crosstalk could be essential in tumorigenesis. LKB1 is a master serine/threonine kinase that phosphorylates AMP-activated protein kinase (AMPK) and at least 13 other related kinases in response to modifications in the intracellular energy levels98. In turn, AMPK phos-phorylates various substrates to control glucose and lipid metabolism. STK11 was identified as a tumour suppressor gene the loss of which is associated with the cancer predisposition disorder Peutz–Jeghers syndrome (PJS)99. STK11 is also commonly mutated (and inactivated) in different sporadic carcinomas, mainly in the lungs100, and has more recently been found in 20% of cervical carcinomas101 (TABLE 2; see Supplementary information S2 (table)). The tumori-genic potential of LKB1 is increased when other onco-genes are expressed, such as KRAS, SRC or MYC102–104, or when tumour suppressors are simultaneously deleted, such as PTEN.

Studies of LKB1 have uncovered novel signalling pathways that link its role in metabolism and growth to cell polarity105 (FIG. 3). In Caenorhabditis elegans and D. melanogaster, the function of PAR4, the orthologue of LKB1, was initially connected to the regulation of cell polarity. PAR4 is required for the first asymmetric divisions during the early stages of embryonic develop-ment and for anterior-posterior axis determination in D. melanogaster oocytes106,107. Subsequently, the Clevers’

Figure 2 | Polarity complex proteins and the Hippo pathway in Drosophila melanogaster. Hippo pathway activation is mediated by Hippo (HPO) binding to Salvador (SAV) to phosphorylate (P) Warts (WTS), which is facilitated by MATS. WTS–MATS then phosphorylate the transcription factor Yorkie (YKI) on specific serine residues70,71, which inactivates it by a mechanism that is thought to involve 14-3-3 binding and translocation of 14-3-3–YKI from the nucleus to the cytoplasm193,194. When the Hippo pathway is inactivated, YKI is dephosphorylated and transported back to the nucleus, where it binds the transcription factor Scalloped (SD), which leads to the upregulation of genes that promote cell proliferation and survival, including Diminutive (DM; the Drosophila melanogaster homologue of MYC), inhibitor of apoptosis 2 (IAP2) and cyclin E195–197. Conversely, the RAS-associated domain family protein (RASSF) inhibits HPO activity by preventing the binding of SAV to HPO198. Interestingly, this function of RASSF seems to be opposite to the well-established role of this protein as a tumour suppressor in humans199. Upstream of HPO, the band 4.1 family proteins Expanded (EX), Merlin (MER), Kibra and the atypical cadherin FAT positively regulate HPO activity and thereby inhibit YKI200. The cell polarity regulators lethal (2) giant larvae (LGL), atypical protein kinase C (aPKC) and crumbs (CRB) regulate proliferation and survival by controlling the activity of the Hippo pathway74,75,84,85. aPKC and LGL function antagonistically to control HPO and RASSF localization and activity. CRB directly binds to EX through its juxtamembrane FBM domain, whereas it binds the other components of the CRB complex Stardust (SDT) and PATJ through the PDZ-binding domain that is present in the cytoplasmic tail201. CRB interacts with YKI to promote YKI phosphorylation and cytoplasmic retention, which thereby inactivates YKI. AJ, adherens junction; SCRIB, scribble; TJ, tight junction.

R E V I E W S




Late

ral m

embr

ane

Apical membrane

aPKC AMPKTJ

AJ

PAR3

PAR3

PAR3

PAR6 LKB1 Ezrin

?

?

?

?

?

DVL

ROCK

MRLC2

FIP2P

P

AMPKP P

P

P

P

MST4P

PLKB1

P

Primary cilium or basal body

Liquid flow detection and regulation of cell growth

Cell division and chromosome segregation

PAR1

PPAR1

STRADαMO25

CAMKKβ

PAR3

E-ca

d

PAR3 basolateral exclusion

PAR3

RAB11

Apical brush border formation

Cellular junction formation

Canonical WNT pathway

PCP movements (non-canonical WNT pathway)

Hepatocyte-like polarity

Actomyosin contractility (polarity)

Bile canaliculi formation

Tight junction formation

Assembly of functional E-cadherin adhesion complexes

Centrosome

TSC2

mTORPRAPTOR

Energy metabolism control

LKB1 apicalexclusion

14-3-3P

laboratory showed that on LKB1 activation, single intestinal epithelial cells fully polarize in the absence of junctional cell–cell contacts108, which generated sub-stantial attention on this master kinase in the field of cell polarity in vertebrates. The role of LKB1 in epi-thelial polarity was initially associated with the phos-phorylation of the serine/threonine kinase PAR1 in D. melanogaster, the mammalian homologue of which is formed by a family of four microtubule-associated protein (MAP) and microtubule affinity-regulating kinases (MARK1–4)109.

However, studies have provided evidence that LKB1 might control cell polarity predominantly through AMPK activation, and not through PAR1 (MARK1–4), and this might be the cause of the tumour suppressor function of LKB1 (REFS 110–113). Constitutive activation of AMPK restored many of the defective polarity phenotypes of LKB1-null mutant D. melanogaster epithelial cells110,111. In addition, two studies using calcium-switch experiments (an in vitro assay to analyse cell-junction formation) using monolayers of MDCK epithelial cells showed that AMPK is required for tight junction formation112,113, although it

Figure 3 | LKB1 regulates epithelial cell polarity, cell growth and energy metabolism. The role of liver kinase B1 (LKB1) in epithelial polarity is associated with the phosphorylation (P) of serine and threonine residues in different members of the AMP-associated kinase (AMPK) family98, including the serine/threonine kinase PAR1, AMPK and mammalian STE20-like protein kinase 4 (MST4). PAR1 phosphorylates PAR3 on two conserved serine residues to generate 14-3-3-binding sites. This inhibits the formation of the PAR3–PAR6–atypical protein kinase C (aPKC) complex by blocking PAR3 oligomerization and binding to aPKC. As PAR1 is associated with the basolateral membrane in epithelial cells, this mechanism is required for PAR3 exclusion in the basolateral region. PAR1 inhibits myosin II in a RHO kinase (ROCK)-dependent manner to promote lateral lumen polarity in MDCK cells independently of Ca2+-mediated cell–cell adhesion202. Phosphorylation of RAB11–FIP2 by PAR1 controls cellular junction formation203. PAR1-mediated phosphorylation of Dishevelled (DVL) regulates its membrane localization, but it is not required for canonical WNT signalling. However, PAR1 is essential for the activation of the canonical WNT pathway, possibly through targets other than DVL204. In polarized epithelial cells, E-cadherin (E-cad) regulates AMPK phosphorylation by controlling the localization of the LKB1 complex through binding to STE20-related kinase adaptor-α (STRADα). AMPK is required for tight junction (TJ) formation112,113, although it is possible that this activation of AMPK might be mediated by Ca2+-calmodulin-dependent protein kinase kinase-β (CAMKKβ)114, 115. AMPK regulates bile canalicular formation, TJ formation and polarity maintenance118,119. One of the major growth regulatory pathways controlled by LKB1–AMPK is the mTOR pathway. AMPK directly phosphorylates both tuberous sclerosis 2 (TSC2) and regulatory-associated protein of mTOR (RAPTOR) to inhibit mTOR complex 1 (mTORC1) activity. AMPK indirectly induces phosphorylation of myosin regulatory light chain 2 (MRLC2) to regulate actinomyosin contractility for the establishment of polarity and cell division. AMPK localizes dynamically to the centrosome spindle poles, the central spindle midzone and the midbody, suggesting a potential role in faithful cell division and chromosome segregation during mitosis. LKB1 localizes to the primary cilium and basal body, and fluid flow results in increased AMPK phosphorylation at the basal body and inhibition of the mTOR pathway, which limits cell size. LKB1 also induces apical brush border formation in intestinal cells by phosphorylating MST4, which then activates ezrin. PCP, planar cell polarity.

R E V I E W S



Bile canaliculiThe hepatocyte apical lumen.

CentrosomeSmall organelle that nucleates microtubules, which are important for cell shape, transport, signalling and cell division.

is possible that this activation of AMPK might be medi-ated by Ca2+-calmodulin-dependent protein kinase kinase-β (CAMKKβ; also known as CAMKK2)114,115. This function of LKB1–AMPK might also be associated with E-cadherin-mediated adherens junction forma-tion. In polarized epithelial cells, E-cadherin regulates AMPK phosphorylation by controlling the localization (but not the kinase activity) of LKB1 through the pseudo-kinase STE20-related kinase adaptor-α (STRADα)116. The LKB1 complex therefore seems to function downstream of E-cadherin in tumour suppression (FIG. 3). However, STRADα may not mediate LKB1 function in cell polar-ity. A recent study of the embryonic development of C. elegans has shown that the role of PAR-4 (the ortho-logue of LKB1) in polarity may be independent of the strd‑1 complex (the C. elegans orthologue of STRADα), as PAR-4 in complex with the adaptor protein MOP-25-2 is required for AMPK phosphorylation (aak‑1 and aak‑2 are the C. elegans homologues of the AMPK α-catalytic subunit, and aakb‑1 and aakb‑2 are the β-regulatory sub-unit homologues), whereas the role of PAR-4 in energy metabolism control is dependent on STRD-1 (REF. 117). More recent studies in human hepatic cells have revealed that AMPK and LKB1 also regulate the formation of bile canaliculi in a Ca2+-dependent manner, tight junction formation and polarity maintenance118,119, and that the LKB1–AMPK pathway that mediates hepatocyte polari-zation is activated by bile-acid synthesis120, although whether this influences tumorigenesis in hepatocytes is currently unknown.

Downstream of AMPK is the mTOR pathway. mTOR is a central integrator of nutrient and growth factor sig-nalling that is activated by energy stress; it functions to control cell growth in all eukaryotes and is deregulated in most human cancers121. AMPK phosphorylates both tuberous sclerosis 2 (TSC2; also known as tuberin) and regulatory-associated protein of mTOR (RAPTOR) to inhibit mTOR complex 1 (mTORC1) activity, although it remains possible that additional substrates of AMPK con-tribute to the regulation of mTOR (FIG. 3). Interestingly, mTORC1 is the only signalling pathway downstream of LKB1 characterized so far that seems to be activated in human tumours of the lung with LKB1 mutations and from patients with PJS122,123. However, it seems that the role of LKB1 and AMPK in cell polarity is mediated by the reorganization of the actin cytoskeleton through the motor protein myosin II, at least in D. melano gaster embryos and in a human colorectal carcinoma cell line110. In this case, AMPK induces, possibly indirectly124, the phosphorylation of myosin regulatory light chain 2 (MRLC2; also known as MYL12B) on serine-19 (serine-22 in MLC2 in D. melanogaster)110. Myosin II regulates actinomyosin contractility, which is essential for the establishment of polarity and cell division. LKB1 might also regulate the actin cytoskeleton through an AMPK-independent pathway. Specifically, LKB1 induces apical brush border formation in intestinal cells by activating MST4, which then activates ezrin125 (FIG. 3).

During all of the stages of mitosis (including cytokinesis), phosphorylated (and thus active) AMPK dynamically localizes to the centrosome spindle poles,

the central spindle midzone and the midbody in human cancer-derived epithelial cells, suggesting a potential role of AMPK in faithful chromosome segregation during mitosis126. Additionally, a recent study has established a causal relationship between the primary cilium and cell growth through the regulation of the mTOR signalling pathway. Cilia are signalling platforms that protrude as filiform organelles from the plasma membrane, and they act as sensors to detect external fluid flow. LKB1 local-izes to the cilium and basal body, and fluid flow results in increased AMPK phosphorylation at the basal body and inhibition of the mTOR pathway and thus control of cell size (FIG. 3). In fact, knockdown of LKB1 results in enlarged cells compared with controls, and therefore prevents normal cell size regulation under flow condi-tions in kidney epithelia127, establishing a connection between fluid flow, metabolic control and epithelial tubular size.

In conclusion, LKB1 is an activator of the AMPK–mTOR pathway, which links cell metabolism to growth control. Recent evidence has shown a connection between the LKB1–AMPK–mTOR pathway with cell polarity and epithelial architecture. As the access of tumour cells to the key molecules that are used for energy production can be limiting, this connection might be a primitive tumour suppressor mechanism that has arisen to protect cells from the energetic stress that has increased during evolution to form the modern organism.

Stem cells, spindle orientation and cancerThe maintenance of most adult epithelial tissues relies on the presence of multipotent or unipotent stem cells128. These cells are polarized and self-renew through sym-metric cell divisions; whereas stem cells asymmetrically divide by reorienting mitotic spindles in order to generate differentiated, specialized cells; although this is still under debate for some tissues129. Asymmetric cell divisions have been demonstrated to occur in various epithelial tissues, including the epidermis, gut, mammary glands and lung130–136. In the human and mouse intestine, the align-ment of mitotic spindles during asymmetric division of stem cells has been observed, but further studies will be needed to provide more insight into the distribution of cell fate determinants during this process134 and to rec-oncile the differences that are observed in other models in which symmetric cell divisions seem to take place137,138. Evidence indicates that alterations in normal stem cells or early progenitor cells initiate some types of cancer, including colon, prostate, skin, stomach, breast and lung cancer139, although for other epithelial cancers this has not been well defined. This has given rise to two different theories about the cell-of-origin that initiates tumours. One theory postulates that transformation occurs to normal adult stem cells or progenitor cells, whereas the second model proposes that differentiated cells acquire self-renewal capacity and stem cell-like properties on transformation139,140 (FIG. 4a).

Interestingly, many of the genes that control epithe-lial cell polarity also regulate spindle orientation and the symmetry of stem cell divisions129. Epithelial cell polarity

R E V I E W S




Self-renewalSymmetric cell division

Adult stem or progenitor cell

DifferentiationAsymmetric cell division

Asymmetric cell division Asymmetric cell division

Differentiated epithelial cell

a

b

Mutations

Mutations

Symmetric cell divisions

Tumourprogression

Heterogeneous epithelial tumoursTumour-initiating cells and cells with limited replicative potential

D. melanogaster neuroblast Mammalian epidermal stem or progenitor cell

PAR3–PAR6–aPKC

INSC

GαI, PINS (LGN), MUD (NUMA) andp150glued (DCTN1)

PONS and MIRA

NUMB, BRAT and PROS

Tumour-initiatingcell

NeuroblastsNeuronal precursor cells that generate neurons and glia cells in the brain.

proteins asymmetrically localize within a stem cell and subsequently segregate differentially into the two daugh-ter cells so that one of the two daughters adopts a differ-entiated fate. Therefore, alterations in the expression or functional activity of cell polarity proteins may prompt

stem cells or progenitor cells to divide symmetrically and to evade differentiation, which might endow them with tumorigenic characteristics129,141 (FIG. 4a).

Relationship between the machinery that controls cell polarity and spindle orientation. To understand how the orientation of the mitotic spindle contributes to differ-entiation, it is important to understand the molecular mechanisms that regulate its orientation and their links to the cell cycle. The mechanisms that govern this pro-cess have mostly been identified in neuroblasts (the sen-sory organ progenitor cells (SOPs)) in D. melanogaster and in neuroepithelial cells from C. elegans129. In order to reorient the mitotic spindle, cells first respond to extra-cellular signalling cues, during which Notch signalling has a major role. Additionally, there is an asymmetric distribution of apical and basal determinants during mitosis. In neuroblasts, the apical determinants include the PAR polarity complex, inscuteable (INSC), partner of inscuteable (PINS; also known as RAPS. The mam-malian functional homologue of PINS is LGN (also known as GPSM2)), aurora (AUR; the mammalian functional homologue is aurora kinase A (AURKA)), guanine nucleotide binding protein-αI (GαI), mushroom body defect (MUD; the mammalian functional homo-logue is nuclear mitotic apparatus protein 1 (NUMA1)), the dynein binding protein lissencephaly 1 (LIS1) and p150glued (the mammalian homologue of which is dyn-actin 1 (DCTN1)). The basal determinants include LGL, NUMB, prospero (PROS; the mammalian homologue is prospero-like protein 1 (PROX1)), and the translation inhibitor brain tumour (BRAT), as well as partner of NUMB (PON) and miranda (MIRA; a coiled-coil pro-tein that interacts with both PROS and BRAT) for which mammalian homologues have not yet been described (FIG. 5). During cell division, centrosomes duplicate early in the G1/S phase of the cell cycle and mature during G2, so that at the end of G2 cells contain two centrosomes (which are comprised of four centrioles). Centrosome separation starts early in mitosis, whereby the daughter centriole migrates from the apical to the basal domain to form the bipolar mitotic spindle142. Interestingly, the crosstalk between centrosomal and apical proteins reg-ulates their respective localization143,144. In neuroblasts, this ensures the inheritance of the daughter centrosome by the mother stem cell143. This differs in other cells, such as D. melanogaster male germline stem cells145 and the radial glial progenitor cells in mice146, where the mother centrosome is inherited by the mother stem cell.

Centrosome separation is regulated by several kinases, including those of the NEK, aurora and polo families142. In D. melanogaster neuroblasts, AUR phos-phorylates PAR6 in the PAR6–aPKC–LGL complex early in prophase. This activates aPKC, which then phosphorylates LGL and excludes it from the com-plex and the apical compartment and targets it to the basal compartment. Next, bazooka (BAZ) associates with the PAR6–aPKC complex and promotes the sub-sequent phosphorylation and exclusion of NUMB and MIRA from an apical to a basal localization147. AUR-mutant embryos have mitotic centrosome and spindle

Figure 4 | Stem cells and polarized cell divisions and cancer. a | Two different theories about the cell-of-origin that initiates tumours have been postulated, one theory proposes that cancers arise from tissue-specific stem or progenitor cells, because stem cells divide during the lifetime of an organism and therefore have the potential to accumulate harmful mutations. The second theory is known as the clonal evolution model, which proposes that tumours arise from normal cells within epithelial tissues that abnormally acquire the capacity to self-renew139,140. Loss of asymmetric cell divisions in stem or progenitor cell compartments, or alterations in differentiated cells, may lead to their proliferation through symmetric cell divisions, which would promote their expansion. These cells sustain tumorigenesis and generate heterogeneous progeny with limited regenerative potential, as is observed in human carcinomas. b | Asymmetric segregation of cell fate determinants regulates the reorientation of the mitotic spindle in Drosophila melanogaster neuroblasts and in mammalian epithelial cells. In D. melanogaster neuroblasts, the PAR3–PAR6–atypical protein kinase C (aPKC) complex segregates apically and recruits the adaptor protein Inscuteable (INSC), which connects this complex to partner of inscuteable (PINS; the functional homologue of mammalian LGN), guanine nucleotide-associated protein-α

I (Gα

I), mushroom body defect (MUD; the functional

homologue of mammalian nuclear mitotic apparatus protein 1 (NUMA)) and p150glued (the mammalian functional homologue of which is dynactin 1 (DCTN1)) to the crescent directing the orientation of the mitotic spindle during asymmetric cell divisions. aPKC promotes the exclusion of partner of numb (PON)–lethal (2) giant larvae homologue (LGL), and NUMB, which, along with Miranda (MIRA), brain tumour (BRAT) and prospero (PROS), localize to the basal crescents. In the mammalian epidermis, it has been shown that the dynamic interaction between some of these complexes is also a key event for the regulation of spindle orientation and cell differentiation.

R E V I E W S




Asymmetric cell division

Symmetric cell divisions

Basement membrane Loss of tumour

suppressors:• APC • p53• VHL• LKB1

Spindle misorientation

Cancer

CentriolesConstituents of the centrosome, which is formed by two pairs of barrel-shaped structures. Involved in the organization of microtubules and mitotic spindles.

defects148. During metaphase and telophase of an asym-metric mitosis BAZ binds to and recruits INSC. This adaptor protein, which is only expressed in cells that divide asymmetrically, recruits PINS, GαI and MUD, which further promotes the exclusion of NUMB, PON, MIRA, BRAT and PROS to the basal crescents (FIG. 5). It is important to mention that the precise function of these proteins is not well understood, but NUMB is involved in endocytosis, is an inhibitor of NOTCH, prevents the self-renewal of neuroblasts and allows differentiation129. MUD binds to PINS, and this complex connects microtubules that radiate from the centrosomes to the cell cortex, known as astral micro-tubules, and generates a pulling force to reorient the mitotic spindle. Additionally, in D. melanogaster neuro-blasts, a parallel pathway that involves the interaction of PINS with DLG and the plus end microtubule-oriented kinesin 73 (KHC73) functions to orient the mitotic spindle asymmetrically149,150.

In mammals, the dynamic interaction between aPKC–PAR6–PAR3 and LGN–DLG–GαI is also a key event for the regulation of spindle orientation and cell differentiation20,151. In 3D cultures of epithelial cells, the distribution of polarity cues to the lateral cortex is required to orient cell divisions symmetrically in the

plane of the basement membrane152. In this regard, CDC42 recruits the PAR6–aPKC complex to the api-cal cortex and activates aPKC21,153,154, which prevents LGN from binding GαI and excludes LGN from the apical cortex151,155,156. During this process, the recruit-ment and activity of CDC42 in the apical cortex is regulated by the CDC42 GEFs TUBA (also known as DNMBP)157 and intersectin 2, which also localize to centrosomes155.

In the mammalian epidermis, the expression of INSC is sufficient to drive asymmetric cell divisions131. LGN colocalizes apically with GαI and NUMA in mitotic basal progenitor cells, where GαI recruits LGN, which in turn recruits NUMA133 (FIG. 4b). Reductions in the levels of these proteins do not alter prolifera-tion or apoptosis, but perturb epithelial differentiation owing to the resulting impairment in Notch signal-ling133, which is a key signalling pathway for epidermal differentiation158.

The role of tumour suppressors in the control of spindle orientation. The formation of tumours and their pro-gression to malignancy involves a progressive cascade of events, typically arising as a consequence of mutations in genes that control cell growth. Some tumour suppres-sors associate directly or indirectly with microtubules to either stabilize or destabilize them, and accumulat-ing evidence indicates that they are also involved in the oriented positioning of the mitotic spindles159,160. These include E-cadherin and adhesion molecules, the WNT signalling protein adenomatous polyposis coli (APC), LKB1, p53 and von Hippel–Lindau tumour suppressor (VHL)161–164 (FIG. 5).

It was recently shown in epithelial cells that adher-ens junctions control the polarized positioning of cen-trosomes in interphase cells165, and that this involves the actin cytoskeleton and CDC42 activity166. Interestingly, this is directly mediated by cadherin-mediated homo-philic cell ligation167. Because the centrosome func-tions as the microtubule organizing centre in most cells, loss of E-cadherin may also lead to alterations in centrosomes, orientation of the mitotic spindle and chromosome segregation. However, it is not yet clear which molecules are responsible for the connection between microtubules and adherens junctions during mitosis. In this regard it was shown, using recombi-nant cadherin ligands, that homophilic ligation of E-cadherin orients mitotic spindles during asymmetric cell division, and blocking E-cadherin function with antibodies, expressing an E-cadherin mutant protein or reducing E-cadherin expression disrupts this polariza-tion and the distribution of APC to the cell cortex167. Consistent with these findings, it has been shown that the loss of α-catenin and β1 integrin in the epidermis impairs correct spindle orientation130. β-catenin binds to dynein168, and it was recently shown that β-catenin colocalizes with NUMB in epicardial cells. β-catenin-null epicardial cells exhibit disruption of adherens junctions and the mislocalization of NUMB, which leads to randomized mitotic spindle orientations in the myocardium169.

Figure 5 | Tumour suppressors and spindle orientation. In addition to the machinery that drives spindle orientation during stem cell divisions, some tumour suppressors have also been associated with regulating the oriented positioning of mitotic spindles159,160, including E-cadherin and adhesion molecules, the WNT signalling protein adenomatous polyposis coli (APC), liver kinase B1 (LKB1), von Hippel–Lindau tumour suppressor (VHL) and p53. Mutations in the corresponding genes may impair the proper organization of microtubules or centrosome function or positioning, which leads to misoriented cell division, alterations in differentiation and the expansion of stem cell compartments.

R E V I E W S



Interestingly, β-catenin is also involved in centrosome separation170. APC binds to β-catenin and to the micro-tubule binding protein end-binding protein 1 (EB1; also known as MAPRE1), which also links kinetochores to kinetochore microtubules, which is important for chro-mosome stability171. In fact, loss of E-cadherin (which binds catenins) in SOPs alters PAR3 distribution and ori-ented cell divisions172. Interestingly, loss of E-cadherin in D. melanogaster germline cells also promotes alterations in centrosome positioning, which is probably mediated by the resulting failure to localize APC173. p120-catenin (also known as catenin-δ1) also binds to microtubules and regulates centrosome divisions in mammalian cells174,175. Further research should provide insight into the possible involvement of adherens junctions and microtubules in the regulation of mitotic spindle orientation.

The mechanisms by which APC regulates spindle orientation are providing additional insight into its roles as a tumour suppressor. APC is mutated in >80% of sporadic colorectal tumours176. Most APC muta-tions that have been found in colorectal cancers encode carboxy-terminal truncations that are required for its interaction with axin and β-catenin, and which there-fore abrogate its function in promoting β-catenin deg-radation, which leads to the constitutive activation of the WNT–β-catenin pathway.

However, APC is also involved in the regulation of the cytoskeleton through its interaction with different microtubule-associated proteins. Interactions between mutated APC and the cytoskeleton have been associ-ated with chromosome instability, and could poten-tially be involved in the deregulation of cell polarity, motility and cell migration, and therefore cancer progression58. Its interactions with the microtubule cytoskeleton, depending on cell context, also seem to be required for correct spindle orientation and cell division. Disrupting APC and its interacting partner, EB1, disorient cell divisions and favour asymmetrical cell division in D. melanogaster epithelial cells177. In mammals, it has been shown that loss of APC in the epidermis and thymus leads to alterations in cell fate and differentiation178. Additionally, APC deletions in intestinal stem cells causally contribute to intestinal adenoma formation179, and APC loss in the gut causes spindle misorientation during cell division and induces pre-tumorigenic lesions in vivo134,180.

The loss of p53 expression or function is one of the most common alterations in carcinomas. It is well established that p53 is involved in cell cycle arrest and apoptosis, and a novel role by which its loss might promote tumorigenesis has recently been described in mammary stem cells135. The loss of p53 in these cells enhances cell proliferation and promotes sym-metric cell divisions that lead to tumorigenesis135. This highlights the relevance of the loss of asymmet-ric cell divisions in tumour initiation. Restoration of p53 expression was sufficient to reduce the replicative potential of mammary cancer stem cells that are defi-cient in p53 and ERBB2, but also reduced the number of symmetric cell divisions, which promoted cell differ-entiation135. The mechanism may involve the regulation

of the Notch signalling pathway by p53 (REFS 135,181). Interestingly, it has been shown that p53 restricts the reprogramming of somatic cells to form induced pluri-potent stem cells (iPS). Loss of p53 allowed the acquisi-tion of stem cell-like properties, and when transplanted in vivo these cells gave rise to teratomas182.

Finally, a new layer of regulation of spindle orien-tation that involves VHL has recently been reported. Germline mutations of VHL are associated with von Hippel–Lindau syndrome, which is characterized by the development of tumours in several organs, mainly owing to the failure of targeting hypoxia-inducible factor 1α (HIF1α) for degradation. However, it has now been shown that VHL regulates spindle orientation by mecha-nisms that are not yet well understood163, but which may involve the deregulation of cell–cell junctions or polarity complexes. In this regard, VHL associates with the PAR polarity complex, and its loss impinges on microtubule stability and cilia formation164.

Overall, increasing evidence suggests that the loss of cell polarity proteins in early stages of carcinogenesis contributes to transformation owing to the loss of con-trol of asymmetric cell divisions. This, in addition to the loss of key tumour suppressor proteins that have been implicated in these processes (such as APC, p53 and VHL), may be responsible for the abnormal self-renewal and expansion of the tumour cells of origin.

Concluding remarks and future perspectivesA wealth of knowledge about the mechanisms that regulate the establishment of cell polarity has unrav-elled their important contributions to preventing the acquisition of tumorigenic characteristics. Given the crucial roles of polarity complexes in the regulation of several pathways that are required for the proper func-tion and proliferation of epithelial tissues, including the establishment of cell adhesion and the regulation of cell growth, apoptosis and metabolism, they can be potential clinical prognostic factors. Future research will shed light on the molecular nature of these inter-actions to possibly influence their assembly through therapeutic interventions. Loss of the finely tuned functions of the polarity complexes may also con-tribute to the early stages of tumorigenesis. The role of polarity complexes in regulating the orientation of centrosomes and mitotic spindles, and thus asymmet-ric cell division, may affect the balance of proliferation and differentiation in epithelial tissues. Furthermore, the crosstalk with tumour suppressors has been shown to contribute to the expansion of the cells of origin of some tumour types, by circumventing growth inhibi-tory signals, differentiation, senescence or apoptosis and the acquisition of migratory and invasive charac-teristics. Understanding the genetic and biochemical mechanisms that control self-renewal and asymmetric division and understanding the role of the stem cell niche in regulating the biological properties of both normal and cancer stem cells are key issues for the future, because preventing these abnormalities may limit the proportion of cells that fuel tumour growth in carcinomas.

R E V I E W S



1. Hanahan, D. & Weinberg, R. A. Hallmarks of cancer: the next generation. Cell 144, 646–674 (2011).

2. St. Johnston, D. & Ahringer, J. Cell polarity in eggs and epithelia: parallels and diversity. Cell 141, 757–774 (2010).

3. Simons, M. & Mlodzik, M. Planar cell polarity signaling: from fly development to human disease. Annu. Rev. Genet. 42, 517–540 (2008).

4. Bornens, M. Organelle positioning and cell polarity. Nature Rev. Mol. Cell Biol. 9, 874–886 (2008).

5. Ray, S. & Lechler, T. Regulation of asymmetric cell division in the epidermis. Cell Div. 6, 12 (2011).

6. Perez-Moreno, M., Jamora, C. & Fuchs, E. Sticky business: orchestrating cellular signals at adherens junctions. Cell 112, 535–548 (2003).

7. Tamura, A. et al. Megaintestine in claudin-15-deficient mice. Gastroenterology 134, 523–534 (2008).

8. Martin-Belmonte, F. & Mostov, K. Regulation of cell polarity during epithelial morphogenesis. Curr. Opin. Cell Biol. 20, 227–234 (2008).

9. Lee, M. & Vasioukhin, V. Cell polarity and cancer--cell and tissue polarity as a non-canonical tumor suppressor. J. Cell Sci. 121, 1141–1150 (2008).

10. Weber, G. F., Bjerke, M. A. & DeSimone, D. W. Integrins and cadherins join forces to form adhesive networks. J. Cell Sci. 124, 1183–1193 (2011).

11. Ooshio, T. et al. Cooperative roles of Par-3 and afadin in the formation of adherens and tight junctions. J. Cell Sci. 120, 2352–2365 (2007).

12. Sakisaka, T., Ikeda, W., Ogita, H., Fujita, N. & Takai, Y. The roles of nectins in cell adhesions: cooperation with other cell adhesion molecules and growth factor receptors. Curr. Opin. Cell Biol. 19, 593–602 (2007).

13. Adams, C. L., Chen, Y. T., Smith, S. J. & Nelson, W. J. Mechanisms of epithelial cell-cell adhesion and cell compaction revealed by high-resolution tracking of E-cadherin-green fluorescent protein. J. Cell Biol. 142, 1105–1119 (1998).

14. Vasioukhin, V., Bauer, C., Yin, M. & Fuchs, E. Directed actin polymerization is the driving force for epithelial cell-cell adhesion. Cell 100, 209–219 (2000).

15. Tsukita, S., Katsuno, T., Yamazaki, Y., Umeda, K. & Tamura, A. Roles of ZO-1 and ZO-2 in establishment of the belt-like adherens and tight junctions with paracellular permselective barrier function. Ann. NY Acad. Sci. 1165, 44–52 (2009).

16. Mege, R. M., Gavard, J. & Lambert, M. Regulation of cell-cell junctions by the cytoskeleton. Curr. Opin. Cell Biol. 18, 541–548 (2006).

17. Kovacs, E. M. & Yap, A. S. Cell-cell contact: cooperating clusters of actin and cadherin. Curr. Biol. 18, R667–R669 (2008).

18. Chen, X. & Macara, I. G. Par-3 controls tight junction assembly through the Rac exchange factor Tiam1. Nature Cell Biol. 7, 262–269 (2005).

19. Mertens, A. E., Rygiel, T. P., Olivo, C., van der Kammen, R. & Collard, J. G. The Rac activator Tiam1 controls tight junction biogenesis in keratinocytes through binding to and activation of the Par polarity complex. J. Cell Biol. 170, 1029–1037 (2005).

20. McCaffrey, L. M. & Macara, I. G. Widely conserved signaling pathways in the establishment of cell polarity. Cold Spring Harb. Perspect. Biol. 1, a001370 (2009).

21. Horikoshi, Y. et al. Interaction between PAR-3 and the aPKC-PAR-6 complex is indispensable for apical domain development of epithelial cells. J. Cell Sci. 122, 1595–1606 (2009).

22. Morais-de-Sa, E., Mirouse, V. & St. Johnston, D. aPKC phosphorylation of Bazooka defines the apical/lateral border in Drosophila epithelial cells. Cell 141, 509–523 (2010).This paper describes how the apical-lateral border and the positioning of apical junctional complexes is regulated in epithelial cells in D. melanogaster.

23. Walther, R. F. & Pichaud, F. Crumbs/DaPKC-dependent apical exclusion of Bazooka promotes photoreceptor polarity remodeling. Curr. Biol. 20, 1065–1074 (2010).

24. Sotillos, S., Diaz-Meco, M. T., Caminero, E., Moscat, J. & Campuzano, S. DaPKC-dependent phosphorylation of Crumbs is required for epithelial cell polarity in Drosophila. J. Cell Biol. 166, 549–557 (2004).

25. Krahn, M. P., Buckers, J., Kastrup, L. & Wodarz, A. Formation of a Bazooka-Stardust complex is essential for plasma membrane polarity in epithelia. J. Cell Biol. 190, 751–760 (2010).

26. Bilder, D., Li, M. & Perrimon, N. Cooperative regulation of cell polarity and growth by Drosophila tumor suppressors. Science 289, 113–116 (2000).

27. Laprise, P., Viel, A. & Rivard, N. Human homolog of disc-large is required for adherens junction assembly and differentiation of human intestinal epithelial cells. J. Biol. Chem. 279, 10157–10166 (2004).

28. Navarro, C. et al. Junctional recruitment of mammalian Scribble relies on E-cadherin engagement. Oncogene 24, 4330–4339 (2005).

29. Yamanaka, T. et al. Mammalian Lgl forms a protein complex with PAR-6 and aPKC independently of PAR-3 to regulate epithelial cell polarity. Curr. Bio.13, 734–743 (2003).

30. Benton, R. & St. Johnston, D. Drosophila PAR-1 and 14-3-3 inhibit Bazooka/PAR-3 to establish complementary cortical domains in polarized cells. Cell 115, 691–704 (2003).

31. Yamanaka, T. & Ohno, S. Role of Lgl/Dlg/Scribble in the regulation of epithelial junction, polarity and growth. Front. Biosci. 13, 6693–6707 (2008).

32. Fehon, R. G., McClatchey, A. I. & Bretscher, A. Organizing the cell cortex: the role of ERM proteins. Nature Rev. Mol. Cell Biol. 11, 276–287 (2010).

33. Gladden, A. B., Hebert, A. M., Schneeberger, E. E. & McClatchey, A. I. The NF2 tumor suppressor, Merlin, regulates epidermal development through the establishment of a junctional polarity complex. Dev. Cell 19, 727–739 (2010).

34. Yi, C. et al. A tight junction-associated Merlin-angiomotin complex mediates Merlin’s regulation of mitogenic signaling and tumor suppressive functions. Cancer Cell 19, 527–540 (2011).

35. Birchmeier, W. & Behrens, J. Cadherin expression in carcinomas: role in the formation of cell junctions and the prevention of invasiveness. Biochim. Biophys. Acta 1198, 11–26 (1994).

36. Jeanes, A., Gottardi, C. J. & Yap, A. S. Cadherins and cancer: how does cadherin dysfunction promote tumor progression? Oncogene 27, 6920–6929 (2008).

37. Gonzalez-Mariscal, L., Lechuga, S. & Garay, E. Role of tight junctions in cell proliferation and cancer. Prog. Histochem. Cytochem. 42, 1–57 (2007).

38. Escudero-Esparza, A., Jiang, W. G. & Martin, T. A. The Claudin family and its role in cancer and metastasis. Front. Biosci. 16, 1069–1083 (2011).

39. Benhamouche, S. et al. Nf2/Merlin controls progenitor homeostasis and tumorigenesis in the liver. Genes Dev. 24, 1718–1730 (2010).

40. Tanos, B. & Rodriguez-Boulan, E. The epithelial polarity program: machineries involved and their hijacking by cancer. Oncogene 27, 6939–6957 (2008).

41. Coradini, D., Casarsa, C. & Oriana, S. Epithelial cell polarity and tumorigenesis: new perspectives for cancer detection and treatment. Acta Pharmacol. Sin 32, 552–564 (2011).

42. Huang, L. & Muthuswamy, S. K. Polarity protein alterations in carcinoma: a focus on emerging roles for polarity regulators. Curr. Opin. Genet. Dev. 20, 41–50 (2010).

43. Moreno-Bueno, G., Portillo, F. & Cano, A. Transcriptional regulation of cell polarity in EMT and cancer. Oncogene 27, 6958–6969 (2008).

44. Brumby, A. M. & Richardson, H. E. scribble mutants cooperate with oncogenic Ras or Notch to cause neoplastic overgrowth in Drosophila. EMBO J. 22, 5769–5779 (2003).

45. Pagliarini, R. A. & Xu, T. A genetic screen in Drosophila for metastatic behavior. Science 302, 1227–1231 (2003).

46. Wu, M., Pastor-Pareja, J. C. & Xu, T. Interaction between Ras(V12) and scribbled clones induces tumour growth and invasion. Nature 463, 545–548 (2010).The authors found that the RAS-G12V and inactivating SCRIB– mutations can cause tumours even when the mutations affect different adjacent epithelial cells. The interaction between RASG12V and SCRIB– clones involves activation of JNK signalling and JNK-induced upregulation of cytokines that activate JAK–signal transducer and activator of transcription (STAT) signalling, a compensatory growth mechanism for tissue homeostasis.

47. Zhan, L. et al. Deregulation of scribble promotes mammary tumorigenesis and reveals a role for cell polarity in carcinoma. Cell 135, 865–878 (2008).

48. Etienne-Manneville, S. Polarity proteins in migration and invasion. Oncogene 27, 6970–6980 (2008).

49. Dow, L. E. & Humbert, P. O. Polarity regulators and the control of epithelial architecture, cell migration, and tumorigenesis. Int. Rev. Cytol. 262, 253–302 (2007).

50. Dow, L. E. et al. Loss of human Scribble cooperates with H-Ras to promote cell invasion through deregulation of MAPK signalling. Oncogene 27, 5988–6001 (2008).

51. Nagasaka, K. et al. The cell polarity regulator hScrib controls ERK activation through a KIM site-dependent interaction. Oncogene 29, 5311–5321 (2010).

52. Aigner, K. et al. The transcription factor ZEB1 (deltaEF1) promotes tumour cell dedifferentiation by repressing master regulators of epithelial polarity. Oncogene 26, 6979–6988 (2007).

53. Spaderna, S. et al. The transcriptional repressor ZEB1 promotes metastasis and loss of cell polarity in cancer. Cancer Res. 68, 537–544 (2008).

54. Kuphal, S. et al. Expression of Hugl-1 is strongly reduced in malignant melanoma. Oncogene 25, 103–110 (2006).

55. Reischauer, S., Levesque, M. P., Nusslein-Volhard, C. & Sonawane, M. Lgl2 executes its function as a tumor suppressor by regulating ErbB signaling in the zebrafish epidermis. PLoS Genet. 5, e1000720 (2009).

56. Lu, X. et al. Aberrant splicing of Hugl-1 is associated with hepatocellular carcinoma progression. Clin. Cancer Res. 15, 3287–3296 (2009).

57. Karp, C. M. et al. Role of the polarity determinant crumbs in suppressing mammalian epithelial tumor progression. Cancer Res. 68, 4105–4115 (2008).This study identified a mammalian orthologue of the D. melanogaster polarity regulator CRB, CRB3, the loss of expression of which promoted tumour progression.

58. Wodarz, A. & Nathke, I. Cell polarity in development and cancer. Nature Cell Biol. 9, 1016–1024 (2007).

59. Hynes, N. E. & MacDonald, G. ErbB receptors and signaling pathways in cancer. Curr. Opin. Cell Biol. 21, 177–184 (2009).

60. Aranda, V. et al. Par6-aPKC uncouples ErbB2 induced disruption of polarized epithelial organization from proliferation control. Nature Cell Biol. 8, 1235–1245 (2006).

61. Nolan, M. E. et al. The polarity protein Par6 induces cell proliferation and is overexpressed in breast cancer. Cancer Res. 68, 8201–8209 (2008).

62. Ozdamar, B. et al. Regulation of the polarity protein Par6 by TGFbeta receptors controls epithelial cell plasticity. Science 307, 1603–1609 (2005).

63. Townsend, T. A., Wrana, J. L., Davis, G. E. & Barnett, J. V. Transforming growth factor-beta-stimulated endocardial cell transformation is dependent on Par6c regulation of RhoA. J. Biol. Chem. 283, 13834–13841 (2008).

64. Singh, A. & Settleman, J. EMT, cancer stem cells and drug resistance: an emerging axis of evil in the war on cancer. Oncogene 29, 4741–4751 (2010).

65. Murray, N. R., Kalari, K. R. & Fields, A. P. Protein kinase Ciota expression and oncogenic signaling mechanisms in cancer. J. Cell Physiol. 226, 879–887 (2011).

66. Murray, N. R. et al. Protein kinase Ciota is required for Ras transformation and colon carcinogenesis in vivo. J. Cell Biol. 164, 797–802 (2004).

67. Regala, R. P. et al. Atypical protein kinase C iota is an oncogene in human non-small cell lung cancer. Cancer Res. 65, 8905–8911 (2005).

68. Brumby, A. M. et al. Identification of novel Ras-cooperating oncogenes in Drosophila melanogaster: a RhoGEF/Rho-family/JNK pathway is a central driver of tumorigenesis. Genetics 188, 105–125 (2011).

69. Genevet, A. & Tapon, N. The Hippo pathway and apico-basal cell polarity. Biochem. J. 436, 213–224 (2011).

70. Pan, D. The hippo signaling pathway in development and cancer. Dev. Cell 19, 491–505 (2010).

71. Harvey, K. & Tapon, N. The Salvador-Warts-Hippo pathway - an emerging tumour-suppressor network. Nature Rev. Cancer 7, 182–191 (2007).

72. Parsons, L. M., Grzeschik, N. A., Allott, M. L. & Richardson, H. E. Lgl/aPKC and Crb regulate the Salvador/Warts/Hippo pathway. Fly (Austin) 4, 288–293 (2010).

73. Zhao, M., Szafranski, P., Hall, C. A. & Goode, S. Basolateral junctions utilize warts signaling to control epithelial-mesenchymal transition and proliferation crucial for migration and invasion of Drosophila ovarian epithelial cells. Genetics 178, 1947–1971 (2008).

74. Grzeschik, N. A., Parsons, L. M., Allott, M. L., Harvey, K. F. & Richardson, H. E. Lgl, aPKC, and Crumbs regulate the Salvador/Warts/Hippo pathway through two distinct mechanisms. Curr. Biol. 20, 573–581 (2010).

R E V I E W S



75. Robinson, B. S., Huang, J., Hong, Y. & Moberg, K. H. Crumbs regulates Salvador/Warts/Hippo signaling in Drosophila via the FERM-domain protein Expanded. Curr. Biol. 20, 582–590 (2010).

76. Varelas, X. et al. The Crumbs complex couples cell density sensing to Hippo-dependent control of the TGF-β-SMAD pathway. Dev. Cell 19, 831–844 (2010).The authors demonstrate that the CRB polarity complex interacts with the Hippo pathway transcriptional regulators TAZ–YAP1, which relays cell density information by promoting TAZ–YAP1 phosphorylation and its cytoplasmic retention and thereby suppresses TGFβ signalling. Disruption of the CRB complex enhanced TGFβ signalling and predisposed cells to TGFβ-mediated EMT.

77. Zhao, B., Tumaneng, K. & Guan, K. L. The Hippo pathway in organ size control, tissue regeneration and stem cell self-renewal. Nature Cell Biol. 13, 877–883 (2011).

78. Ashton, G. H. et al. Focal adhesion kinase is required for intestinal regeneration and tumorigenesis downstream of Wnt/c-Myc signaling. Dev. Cell 19, 259–269 (2010).

79. Cai, J. et al. The Hippo signaling pathway restricts the oncogenic potential of an intestinal regeneration program. Genes Dev. 24, 2383–2388 (2010).

80. Karpowicz, P., Perez, J. & Perrimon, N. The Hippo tumor suppressor pathway regulates intestinal stem cell regeneration. Development 137, 4135–4145 (2010).

81. Shao, J. & Sheng, H. Amphiregulin promotes intestinal epithelial regeneration: roles of intestinal subepithelial myofibroblasts. Endocrinology 151, 3728–3737 (2010).

82. Shaw, R. L. et al. The Hippo pathway regulates intestinal stem cell proliferation during Drosophila adult midgut regeneration. Development 137, 4147–4158 (2010).

83. Staley, B. K. & Irvine, K. D. Warts and Yorkie mediate intestinal regeneration by influencing stem cell proliferation. Curr. Bio. 20, 1580–1587 (2010).

84. Ling, C. et al. The apical transmembrane protein Crumbs functions as a tumor suppressor that regulates Hippo signaling by binding to Expanded. Proc. Natl Acad. Sci. USA 107, 10532–10537 (2010).

85. Chen, C. L. et al. The apical-basal cell polarity determinant Crumbs regulates Hippo signaling in Drosophila. Proc. Natl Acad. Sci. USA 107, 15810–15815 (2010).References 74, 75, 84 and 85 provide evidence that the cell polarity regulators LGL, aPKC and CRB regulate proliferation and survival in the developing eye and imaginal disc epithelia by controlling activity of the Hippo tumour suppressor pathway in D. melanogaster.

86. Kim, N. G., Koh, E., Chen, X. & Gumbiner, B. M. E-cadherin mediates contact inhibition of proliferation through Hippo signaling-pathway components. Proc. Natl Acad. Sci. USA 108, 11930–11935 (2011).

87. Heallen, T. et al. Hippo pathway inhibits Wnt signaling to restrain cardiomyocyte proliferation and heart size. Science 332, 458–461 (2011).

88. Schlegelmilch, K. et al. Yap1 acts downstream of alpha-catenin to control epidermal proliferation. Cell 144, 782–795 (2011).

89. Richter, A. M., Pfeifer, G. P. & Dammann, R. H. The RASSF proteins in cancer; from epigenetic silencing to functional characterization. Biochim. Biophys. Acta 1796, 114–128 (2009).

90. Seidel, C. et al. Frequent hypermethylation of MST1 and MST2 in soft tissue sarcoma. Mol. Carcinog. 46, 865–871 (2007).

91. Takahashi, Y. et al. Down-regulation of LATS1 and LATS2 mRNA expression by promoter hypermethylation and its association with biologically aggressive phenotype in human breast cancers. Clin. Cancer Res. 11, 1380–1385 (2005).

92. Okada, T., You, L. & Giancotti, F. G. Shedding light on Merlin’s wizardry. Trends Cell Biol. 17, 222–229 (2007).

93. Morrison, H. et al. The NF2 tumor suppressor gene product, merlin, mediates contact inhibition of growth through interactions with CD44. Genes Dev. 15, 968–980 (2001).

94. McClatchey, A. I. & Giovannini, M. Membrane organization and tumorigenesis-the NF2 tumor suppressor, Merlin. Genes Dev. 19, 2265–2277 (2005).

95. Zhang, N. et al. The Merlin/NF2 tumor suppressor functions through the YAP oncoprotein to regulate tissue homeostasis in mammals. Dev. Cell 19, 27–38 (2010).

96. Lee, K. P. et al. The Hippo-Salvador pathway restrains hepatic oval cell proliferation, liver size, and liver tumorigenesis. Proc. Natl Acad. Sci. USA 107, 8248–8253 (2010).

97. Lu, L. et al. Hippo signaling is a potent in vivo growth and tumor suppressor pathway in the mammalian liver. Proc. Natl Acad. Sci. USA 107, 1437–1442 (2010).References 39, 95, 96 and 97 link merlin to mammalian Hippo signalling and implicate YAP1 activation as a mediator of phenotypes that occur in patients with neurofibromatosis type II.

98. Williams, T. & Brenman, J. E. LKB1 and AMPK in cell polarity and division. Trends Cell Biol. 18, 193–198 (2008).

99. Hemminki, A. et al. A serine/threonine kinase gene defective in Peutz-Jeghers syndrome. Nature 391, 184–187 (1998).

100. Sanchez-Cespedes, M. A role for LKB1 gene in human cancer beyond the Peutz-Jeghers syndrome. Oncogene 26, 7825–7832 (2007).

101. Wingo, S. N. et al. Somatic LKB1 mutations promote cervical cancer progression. PLoS One 4, e5137 (2009).

102. Mizrachy-Schwartz, S., Cohen, N., Klein, S., Kravchenko-Balasha, N. & Levitzki, A. Up-regulation of AMP-activated protein kinase in cancer cell lines is mediated through c-Src activation. J. Biol. Chem. 286, 15268–15277 (2011).

103. Morton, J. P. et al. LKB1 haploinsufficiency cooperates with Kras to promote pancreatic cancer through suppression of p21-dependent growth arrest. Gastroenterology 139, 586–597 (2010).

104. Partanen, J. I., Nieminen, A. I., Makela, T. P. & Klefstrom, J. Suppression of oncogenic properties of c-Myc by LKB1-controlled epithelial organization. Proc. Natl Acad. Sci. USA 104, 14694–14699 (2007).

105. Shackelford, D. B. & Shaw, R. J. The LKB1-AMPK pathway: metabolism and growth control in tumour suppression. Nature Rev. Cancer 9, 563–575 (2009).

106. Kemphues, K. J., Priess, J. R., Morton, D. G. & Cheng, N. S. Identification of genes required for cytoplasmic localization in early, C. elegans embryos. Cell 52, 311–320 (1988).

107. Martin, S. G. & St. Johnston, D. A role for Drosophila LKB1 in anterior-posterior axis formation and epithelial polarity. Nature 421, 379–384 (2003).

108. Baas, A. F. et al. Complete polarization of single intestinal epithelial cells upon activation of LKB1 by STRAD. Cell 116, 457–466 (2004).

109. Mirouse, V. & Billaud, M. The LKB1/AMPK polarity pathway. FEBS Lett. 585, 981–985 (2011).

110. Lee, J. H. et al. Energy-dependent regulation of cell structure by AMP-activated protein kinase. Nature 447, 1017–1020 (2007).

111. Mirouse, V., Swick, L. L., Kazgan, N., St. Johnston, D. & Brenman, J. E. LKB1 and AMPK maintain epithelial cell polarity under energetic stress. J. Cell Biol. 177, 387–392 (2007).

112. Zhang, L., Li, J., Young, L. H. & Caplan, M. J. AMP-activated protein kinase regulates the assembly of epithelial tight junctions. Proc. Natl Acad. Sci. USA 103, 17272–17277 (2006).

113. Zheng, B. & Cantley, L. C. Regulation of epithelial tight junction assembly and disassembly by AMP-activated protein kinase. Proc. Natl Acad. Sci. USA 104, 819–822 (2007).References 110–113 provide substantial evidence that LKB1, through AMPK activation and not PAR1 activation, controls cell polarity in D. melanogaster and tight junction formation in human cells. The role of LKB1 and AMPK in cell polarity is mediated by the reorganization of the actin cytoskeleton through the motor protein myosin II.

114. Hurley, R. L. et al. The Ca2+/calmodulin-dependent protein kinase kinases are AMP-activated protein kinase kinases. J. Biol. Chem. 280, 29060–29066 (2005).

115. Woods, A. et al. Ca2+/calmodulin-dependent protein kinase kinase-beta acts upstream of AMP-activated protein kinase in mammalian cells. Cell. Metab. 2, 21–33 (2005).

116. Sebbagh, M., Santoni, M. J., Hall, B., Borg, J. P. & Schwartz, M. A. Regulation of LKB1/STRAD localization and function by E-cadherin. Curr. Biol. 19, 37–42 (2009).

117. Narbonne, P., Hyenne, V., Li, S., Labbe, J. C. & Roy, R. Differential requirements for STRAD in LKB1-dependent functions in C. elegans. Development 137, 661–670 (2010).

118. Tang, X. X. et al. Lymphocytes accelerate epithelial tight junction assembly: role of AMP-activated protein kinase (AMPK). PLoS ONE 5, e12343 (2010).

119. Fu, D., Wakabayashi, Y., Ido, Y., Lippincott-Schwartz, J. & Arias, I. M. Regulation of bile canalicular network formation and maintenance by AMP-activated protein kinase and LKB1. J. Cell Sci. 123, 3294–3302 (2010).

120. Fu, D., Wakabayashi, Y., Lippincott-Schwartz, J. & Arias, I. M. Bile acid stimulates hepatocyte polarization through a cAMP-Epac-MEK-LKB1-AMPK pathway. Proc. Natl Acad. Sci. USA 108, 1403–1408 (2011).

121. Guertin, D. A. & Sabatini, D. M. Defining the role of mTOR in cancer. Cancer Cell 12, 9–22 (2007).

122. Carretero, J. et al. Dysfunctional AMPK activity, signalling through mTOR and survival in response to energetic stress in LKB1-deficient lung cancer. Oncogene 26, 1616–1625 (2007).

123. Shackelford, D. B. et al. mTOR and HIF-1alpha-mediated tumor metabolism in an LKB1 mouse model of Peutz-Jeghers syndrome. Proc. Natl Acad. Sci. USA 106, 11137–11142 (2009).

124. Bultot, L. et al. Myosin light chains are not a physiological substrate of AMPK in the control of cell structure changes. FEBS Lett. 583, 25–28 (2009).

125. ten Klooster, J. P. et al. Mst4 and Ezrin induce brush borders downstream of the Lkb1/Strad/Mo25 polarization complex. Dev. Cell 16, 551–562 (2009).

126. Vazquez-Martin, A., Oliveras-Ferraros, C. & Menendez, J. A. The active form of the metabolic sensor: AMP-activated protein kinase (AMPK) directly binds the mitotic apparatus and travels from centrosomes to the spindle midzone during mitosis and cytokinesis. Cell Cycle 8, 2385–2398 (2009).

127. Boehlke, C. et al. Primary cilia regulate mTORC1 activity and cell size through Lkb1. Nature Cell Biol. 12, 1115–1122 (2010).This paper shows that LKB1 localizes in the cilium and flow results in increased AMPK phosphorylation at the basal body. The authors demonstrate that the cilium regulates mTOR signalling and cell size, and identify the cilium-basal body compartment as a spatially restricted activation site for LKB1–AMPK–mTOR signalling.

128. Blanpain, C., Horsley, V. & Fuchs, E. Epithelial stem cells: turning over new leaves. Cell 128, 445–458 (2007).

129. Knoblich, J. A. Asymmetric cell division: recent developments and their implications for tumour biology. Nature Rev. Mol. Cell Biol. 11, 849–860 (2010).

130. Lechler, T. & Fuchs, E. Asymmetric cell divisions promote stratification and differentiation of mammalian skin. Nature 437, 275–280 (2005).This is the first paper to show that asymmetric cell divisions occur in mammalian epidermal stem cells.

131. Poulson, N. D. & Lechler, T. Robust control of mitotic spindle orientation in the developing epidermis. J. Cell Biol. 191, 915–922 (2010).

132. Luxenburg, C., Pasolli, H. A., Williams, S. E. & Fuchs, E. Developmental roles for Srf, cortical cytoskeleton and cell shape in epidermal spindle orientation. Nature Cell Biol. 13, 203–214 (2011).

133. Williams, S. E., Beronja, S., Pasolli, H. A. & Fuchs, E. Asymmetric cell divisions promote Notch-dependent epidermal differentiation. Nature 470, 353–358 (2011).

134. Quyn, A. J. et al. Spindle orientation bias in gut epithelial stem cell compartments is lost in precancerous tissue. Cell Stem Cell 6, 175–181 (2010).

135. Cicalese, A. et al. The tumor suppressor p53 regulates polarity of self-renewing divisions in mammary stem cells. Cell 138, 1083–1095 (2009).

136. El-Hashash, A. H. et al. Eya1 controls cell polarity, spindle orientation, cell fate and Notch signaling in distal embryonic lung epithelium. Development 138, 1395–1407 (2011).

137. Snippert, H. J. et al. Intestinal crypt homeostasis results from neutral competition between symmetrically dividing Lgr5 stem cells. Cell 143, 134–144 (2010).

138. Lopez-Garcia, C., Klein, A. M., Simons, B. D. & Winton, D. J. Intestinal stem cell replacement follows a pattern of neutral drift. Science 330, 822–825 (2010).

139. Visvader, J. E. Cells of origin in cancer. Nature 469, 314–322 (2011).

140. Clevers, H. The cancer stem cell: premises, promises and challenges. Nature Med. 17, 313–319 (2011).

141. Florian, M. C. & Geiger, H. Concise review: polarity in stem cells, disease, and aging. Stem Cells 28, 1623–1629 (2010).

R E V I E W S



142. Lukasiewicz, K. B. & Lingle, W. L. Aurora A, centrosome structure, and the centrosome cycle. Environ. Mol. Mutagen. 50, 602–619 (2009).

143. Januschke, J., Llamazares, S., Reina, J. & Gonzalez, C. Drosophila neuroblasts retain the daughter centrosome. Nature Commun. 2, 243 (2011).

144. Rebollo, E. et al. Functionally unequal centrosomes drive spindle orientation in asymmetrically dividing Drosophila neural stem cells. Dev. Cell 12, 467–474 (2007).

145. Yamashita, Y. M., Mahowald, A. P., Perlin, J. R. & Fuller, M. T. Asymmetric inheritance of mother versus daughter centrosome in stem cell division. Science 315, 518–521 (2007).

146. Wang, X. et al. Asymmetric centrosome inheritance maintains neural progenitors in the neocortex. Nature 461, 947–955 (2009).

147. Wirtz-Peitz, F., Nishimura, T. & Knoblich, J. A. Linking cell cycle to asymmetric division: Aurora-A phosphorylates the Par complex to regulate Numb localization. Cell 135, 161–173 (2008).This paper links the cell cycle machinery and the regulation of asymmetric cell divisions in D. melanogaster neuroblasts.

148. Glover, D. M., Leibowitz, M. H., McLean, D. A. & Parry, H. Mutations in aurora prevent centrosome separation leading to the formation of monopolar spindles. Cell 81, 95–105 (1995).

149. Siegrist, S. E. & Doe, C. Q. Microtubule-induced Pins/Galphai cortical polarity in Drosophila neuroblasts. Cell 123, 1323–1335 (2005).

150. Januschke, J. & Gonzalez, C. The interphase microtubule aster is a determinant of asymmetric division orientation in Drosophila neuroblasts. J. Cell Biol. 188, 693–706 (2010).

151. Zheng, Z. et al. LGN regulates mitotic spindle orientation during epithelial morphogenesis. J. Cell Biol. 189, 275–288 (2010).This study provides evidence that LGN, together with GαI and NUMA, regulates the orientation of mitotic spindles in mammalian cells.

152. Bryant, D. M. & Mostov, K. E. From cells to organs: building polarized tissue. Nature Rev. Mol. Cell Biol. 9, 887–901 (2008).

153. Jaffe, A. B., Kaji, N., Durgan, J. & Hall, A. Cdc42 controls spindle orientation to position the apical surface during epithelial morphogenesis. J. Cell Biol. 183, 625–633 (2008).

154. Durgan, J., Kaji, N., Jin, D. & Hall, A. Par6B and atypical PKC regulate mitotic spindle orientation during epithelial morphogenesis. J. Biol. Chem. 286, 12461–12474 (2011).

155. Rodriguez-Fraticelli, A. E. et al. The Cdc42 GEF Intersectin 2 controls mitotic spindle orientation to form the lumen during epithelial morphogenesis. J. Cell Biol. 189, 725–738 (2010).

156. Hao, Y. et al. Par3 controls epithelial spindle orientation by aPKC-mediated phosphorylation of apical pins. Curr. Biol. 20, 1809–1818 (2010).

157. Qin, Y., Meisen, W. H., Hao, Y. & Macara, I. G. Tuba, a Cdc42 GEF, is required for polarized spindle orientation during epithelial cyst formation. J. Cell Biol. 189, 661–669 (2010).

158. Blanpain, C., Lowry, W. E., Pasolli, H. A. & Fuchs, E. Canonical notch signaling functions as a commitment switch in the epidermal lineage. Genes Dev. 20, 3022–3035 (2006).

159. Gonzalez, C. Spindle orientation, asymmetric division and tumour suppression in Drosophila stem cells. Nature Rev. Genet. 8, 462–472 (2007).

160. Hernandez, P. & Tirnauer, J. S. Tumor suppressor interactions with microtubules: keeping cell polarity and cell division on track. Dis. Model Mech. 3, 304–315 (2010).

161. Szczepanska, K. & Maleszewski, M. LKB1/PAR4 protein is asymmetrically localized in mouse oocytes and associates with meiotic spindle. Gene Expr. Patterns 6, 86–93 (2005).

162. Bonaccorsi, S. et al. The Drosophila Lkb1 kinase is required for spindle formation and asymmetric neuroblast division. Development 134, 2183–2193 (2007).

163. Thoma, C. R. et al. VHL loss causes spindle misorientation and chromosome instability. Nature Cell Biol. 11, 994–1001 (2009).

164. Pease, J. C. & Tirnauer, J. S. Mitotic spindle misorientation in cancer--out of alignment and into the fire. J. Cell Sci. 124, 1007–1016 (2011).

165. Dupin, I., Camand, E. & Etienne-Manneville, S. Classical cadherins control nucleus and centrosome position and cell polarity. J. Cell Biol. 185, 779–786 (2009).

166. Desai, R. A., Gao, L., Raghavan, S., Liu, W. F. & Chen, C. S. Cell polarity triggered by cell-cell adhesion via E-cadherin. J. Cell Sci. 122, 905–911 (2009).

167. den Elzen, N., Buttery, C. V., Maddugoda, M. P., Ren, G. & Yap, A. S. Cadherin adhesion receptors orient the mitotic spindle during symmetric cell division in mammalian epithelia. Mol. Biol. Cell 20, 3740–3750 (2009).

168. Ligon, L. A., Karki, S., Tokito, M. & Holzbaur, E. L. Dynein binds to beta-catenin and may tether microtubules at adherens junctions. Nature Cell Biol. 3, 913–917 (2001).

169. Wu, M. et al. Epicardial spindle orientation controls cell entry into the myocardium. Dev. Cell 19, 114–125 (2010).

170. Bahmanyar, S. et al. beta-Catenin is a Nek2 substrate involved in centrosome separation. Genes Dev. 22, 91–105 (2008).

171. Caldwell, C. M. & Kaplan, K. B. The role of APC in mitosis and in chromosome instability. Adv. Exp. Med. Biol. 656, 51–64 (2009).

172. Le Borgne, R., Bellaiche, Y. & Schweisguth, F. Drosophila E-cadherin regulates the orientation of asymmetric cell division in the sensory organ lineage. Curr. Biol. 12, 95–104 (2002).

173. Inaba, M., Yuan, H., Salzmann, V., Fuller, M. T. & Yamashita, Y. M. E-cadherin is required for centrosome and spindle orientation in Drosophila male germline stem cells. PLoS ONE 5, e12473 (2010).This study shows that E-cadherin-mediated adhesion not only maintains the stem cell niche but also controls mitotic spindle orientation in germline stem cells.

174. Meng, W. & Takeichi, M. Adherens junction: molecular architecture and regulation. Cold Spring Harb. Perspect. Biol. 1, a002899 (2009).

175. Chartier, N. T. et al. Cyclin-dependent kinase 2/cyclin E complex is involved in p120 catenin (p120ctn)-dependent cell growth control: a new role for p120ctn in cancer. Cancer Res. 67, 9781–9790 (2007).

176. Dikovskaya, D. et al. Loss of APC induces polyploidy as a result of a combination of defects in mitosis and apoptosis. J. Cell Biol. 176, 183–195 (2007).

177. Lu, B., Roegiers, F., Jan, L. Y. & Jan, Y. N. Adherens junctions inhibit asymmetric division in the Drosophila epithelium. Nature 409, 522–525 (2001).

178. Kuraguchi, M. et al. Adenomatous polyposis coli (APC) is required for normal development of skin and thymus. PLoS Genet. 2, e146 (2006).

179. Barker, N. et al. Crypt stem cells as the cells-of-origin of intestinal cancer. Nature 457, 608–611 (2009).

180. Fleming, E. S., Temchin, M., Wu, Q., Maggio-Price, L. & Tirnauer, J. S. Spindle misorientation in tumors from APC(min/+) mice. Mol. Carcinog. 48, 592–598 (2009).

181. Tao, L. et al. Repression of mammary stem/progenitor cells by p53 is mediated by Notch and separable from apoptotic activity. Stem Cells 29, 119–127 (2010).

182. Marion, R. M. et al. A p53-mediated DNA damage response limits reprogramming to ensure iPS cell genomic integrity. Nature 460, 1149–1153 (2009).

183. Niessen, C. M. & Gottardi, C. J. Molecular components of the adherens junction. Biochim. Biophys. Acta 1778, 562–571 (2008).

184. Drees, F., Pokutta, S., Yamada, S., Nelson, W. J. & Weis, W. I. Alpha-catenin is a molecular switch that binds E-cadherin-beta-catenin and regulates actin-filament assembly. Cell 123, 903–915 (2005).

185. Cereijido, M., Contreras, R. G., Shoshani, L., Flores-Benitez, D. & Larre, I. Tight junction and polarity interaction in the transporting epithelial phenotype. Biochim. Biophys. Acta 1778, 770–793 (2008).

186. Tsukita, S., Yamazaki, Y., Katsuno, T., Tamura, A. & Tsukita, S. Tight junction-based epithelial microenvironment and cell proliferation. Oncogene 27, 6930–6938 (2008).

187. Assemat, E., Bazellieres, E., Pallesi-Pocachard, E., Le Bivic, A. & Massey-Harroche, D. Polarity complex proteins. Biochim. Biophys. Acta 1778, 614–630 (2008).

188. Bulgakova, N. A. & Knust, E. The Crumbs complex: from epithelial-cell polarity to retinal degeneration. J. Cell Sci. 122, 2587–2596 (2009).

189. Suzuki, A. & Ohno, S. The PAR-aPKC system: lessons in polarity. J. Cell Sci. 119, 979–987 (2006).

190. Goldstein, B. & Macara, I. G. The PAR proteins: fundamental players in animal cell polarization. Dev. Cell 13, 609–622 (2007).

191. Humbert, P. O. et al. Control of tumourigenesis by the Scribble/Dlg/Lgl polarity module. Oncogene 27, 6888–6907 (2008).

192. Feigin, M. E. & Muthuswamy, S. K. Polarity proteins regulate mammalian cell-cell junctions and cancer pathogenesis. Curr. Opin. Cell Biol. 21, 694–700 (2009).

193. Dong, J. et al. Elucidation of a universal size-control mechanism in Drosophila and mammals. Cell 130, 1120–1133 (2007).

194. Zhao, B. et al. Inactivation of YAP oncoprotein by the Hippo pathway is involved in cell contact inhibition and tissue growth control. Genes Dev. 21, 2747–2761 (2007).

195. Goulev, Y. et al. SCALLOPED interacts with YORKIE, the nuclear effector of the hippo tumor-suppressor pathway in Drosophila. Curr. Biol. 18, 435–441 (2008).

196. Wu, S., Liu, Y., Zheng, Y., Dong, J. & Pan, D. The TEAD/TEF family protein Scalloped mediates transcriptional output of the Hippo growth-regulatory pathway. Dev. Cell 14, 388–398 (2008).

197. Zhang, L. et al. The TEAD/TEF family of transcription factor Scalloped mediates Hippo signaling in organ size control. Dev. Cell 14, 377–387 (2008).

198. Polesello, C., Huelsmann, S., Brown, N. H. & Tapon, N. The Drosophila RASSF homolog antagonizes the hippo pathway. Curr. Biol. 16, 2459–2465 (2006).

199. Donninger, H., Vos, M. D. & Clark, G. J. The RASSF1A tumor suppressor. J. Cell Sci. 120, 3163–3172 (2007).

200. Hamaratoglu, F. et al. The tumour-suppressor genes NF2/Merlin and Expanded act through Hippo signalling to regulate cell proliferation and apoptosis. Nature Cell Biol. 8, 27–36 (2006).

201. Rashbass, P. & Skaer, H. Cell polarity: Nailing Crumbs to the scaffold. Curr. Biol. 10, R234–R236 (2000).

202. Cohen, D., Tian, Y. & Musch, A. Par1b promotes hepatic-type lumen polarity in Madin Darby canine kidney cells via myosin II- and E-cadherin-dependent signaling. Mol. Biol. Cell 18, 2203–2215 (2007).

203. Ducharme, N. A. et al. MARK2/EMK1/Par-1Balpha phosphorylation of Rab11-family interacting protein 2 is necessary for the timely establishment of polarity in Madin-Darby canine kidney cells. Mol. Biol. Cell 17, 3625–3637 (2006).

204. Ossipova, O., Dhawan, S., Sokol, S. & Green, J. B. Distinct PAR-1 proteins function in different branches of Wnt signaling during vertebrate development. Dev. Cell 8, 829–841 (2005).

AcknowledgementsThe authors thank C. M. Ruiz-Jarabo for comments on the manuscript and all members of the Martín-Belmonte and Perez-Moreno laboratories for their insightful discussions and support. Research by F.M-B. is supported by grants from the Human Frontiers Science Program (HFSP-CDA 00011/2009), Marie Curie (IRG-209382), MICINN (BFU2008-01916) and CONSOLIDER (CSD2009-00016). An institutional Grant from Fundación Ramón Areces to CBMSO is also acknowledged. M.P-M.’s group is supported by grants from the Spanish Ministry of Science and Innovation (BFU2009-11885), the Association for International Cancer Research AICR-UK (10-0746), and from the CNIO (BC1102).

Competing interests statementThe authors declare no competing financial interests.

FURTHER INFORMATIONFernando Martin-Belmonte’s homepage: http://www.martinbelmontelab.net/fmb/Home.htmlMirna Perez-Moreno’s homepage: http://www.cnio.es/es/grupos/plantillas/presentacion.asp?grupo=50006585

SUPPLEMENTARY INFORMATIONSee online article: S1 (table) | S2 (table)

ALL LINKS ARE ACTIVE IN THE ONLINE PDF

R E V I E W S



http://www.martinbelmontelab.net/fmb/Home.html

http://www.martinbelmontelab.net/fmb/Home.html

http://www.cnio.es/es/grupos/plantillas/presentacion.asp?grupo=50006585

http://www.cnio.es/es/grupos/plantillas/presentacion.asp?grupo=50006585



Epithelial cell polarity, stem cells and cancer

Documents

Transcript of Epithelial cell polarity, stem cells and cancer