Cerebral cavernous malformation proteins at a glance · 2014. 2. 12. · JournalofCellScience Cell Science at a Glance Cerebral cavernous malformation proteins at a glance Kyle M.

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Cell Science at a Glance

Cerebral cavernous malformation proteins at a glance

Kyle M. Draheim1, Oriana S. Fisher1, Titus J. Boggon1,2 and David A. Calderwood1,2,3,*

ABSTRACT

Loss-of-function mutations in genes encoding KRIT1 (also known

as CCM1), CCM2 (also known as OSM and malcavernin) or

PDCD10 (also known as CCM3) cause cerebral cavernous

malformations (CCMs). These abnormalities are characterized by

dilated leaky blood vessels, especially in the neurovasculature, that

result in increased risk of stroke, focal neurological defects and

seizures. The three CCM proteins can exist in a trimeric complex,

and each of these essential multi-domain adaptor proteins also

interacts with a range of signaling, cytoskeletal and adaptor

proteins, presumably accounting for their roles in a range of basic

cellular processes including cell adhesion, migration, polarity and

apoptosis. In this Cell Science at a Glance article and the

accompanying poster, we provide an overview of current models

of CCM protein function focusing on how known protein–protein

interactions might contribute to cellular phenotypes and highlighting

gaps in our current understanding.

KEY WORDS: Cerebral cavernous malformations, CCM, Cell

signaling, KRIT1, PCDC10, Rho Signaling, Vascular biology

1Department of Pharmacology, Yale University School of Medicine, New Haven,CT 06520-8066, USA. 2Interdepartmental Program in Vascular Biology andTherapeutics, Yale University School of Medicine, New Haven, CT 06520-8066,USA. 3Department of Cell Biology, Yale University School of Medicine, NewHaven, CT 06520-8066, USA.

*Author for correspondence ([email protected])

� 2014. Published by The Company of Biologists Ltd | Journal of Cell Science (2014) 127, 701–707 doi:10.1242/jcs.138388

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IntroductionCerebral cavernous malformations (CCMs) consist of clusters of

enlarged endothelial channels (‘caverns’) that are arranged back-to-back to form densely packed sinusoids with little or nointervening brain parenchyma (reviewed in Cavalcanti et al.,

2012; Fischer et al., 2013). These lesions lack smooth muscleand elastic tissue, so the vessel walls are thin, leaky and lacksub-endothelial support and an intact basal lamina. Ultra-structural analysis has revealed ruptures in the luminal

endothelium (probably due to damaged intercellular junctions),endothelial detachment from the basal lamina and decreasednumbers of pericytes (Tanriover et al., 2013). CCMs have been

reported in up to 0.5% of the population and, although they areprimarily found within the neurovasculature of the centralnervous system (i.e. brain, spinal cord, retina), where they result

in increased risk for stroke, seizures and focal neurologicaldeficits, they are also seen in the skin and liver (Cavalcanti et al.,2012; Fischer et al., 2013). Currently, the only treatment for

CCM is surgical resection.CCMs are associated with loss-of-function mutations in any

one of the three CCM genes: KRIT1 (Krev interaction trapped 1,also known as CCM1), CCM2 or PDCD10 (programmed cell

death 10, also known as CCM3) (Cavalcanti et al., 2012) (see Box1), suggesting that there is an essential pathway involving allthree proteins (Stahl et al., 2008). This hypothesis is bolstered by

the knowledge that all three proteins can be found in the samecomplex within the cell (see below). However, PDCD10 mightalso act separately from KRIT1 and CCM2, as its mutation often

results in a more severe form of the disease (Denier et al., 2006;Zhu et al., 2011). Despite extensive analysis of CCM-knockoutanimals (see supplementary material Table S1) and recentadvances in identifying binding partners and determining the

structures of KRIT1, CCM2 and PDCD10, there still is limitedknowledge of the mechanisms through which the loss of each ofthese proteins leads to CCM formation. Here, we review the

current information on CCM protein signaling. In light of theCCM disease phenotype, we focus primarily on potential roles invascular cells, but because all three CCM proteins are widely

expressed, they might also have roles outside of the vasculature.

CCM proteinsKRIT1KRIT1 is the largest of the CCM proteins. It contains an N-terminalNudix domain followed by a stretch of three NPxY/F motifs (Liuet al., 2013), a predicted ankyrin-repeat domain and a C-terminalFERM (band 4.1, ezrin, radixin, moesin) domain (Gingras et al.,

2013; Li et al., 2012) (see poster). No catalytic activity has beenreported for KRIT1, and it is thought to signal through its manybinding partners (see below). KRIT1 is ubiquitously expressed in

early embryogenesis with pronounced endothelial expression inlarge vessels (Guzeloglu-Kayisli et al., 2004). KRIT1 is observedin many different cellular compartments and is actively shuttled

between the cytoplasm and the nucleus (Zawistowski et al., 2005).A polybasic sequence within a KRIT1 Nudix domain loop isresponsible for the targeting of KRIT1 to microtubules, although

the mode of interaction is not yet completely understood (Beraud-Dufour et al., 2007). KRIT1 also localizes to endothelial cellboundaries or cell–cell junctions through its FERM domain(Glading et al., 2007; Zawistowski et al., 2005).

KRIT1 was initially identified in a yeast two-hybrid screenas a Rap1-binding protein (Serebriiskii et al., 1997). Recentcrystallographic analysis revealed a novel mode of interaction

between the KRIT1 FERM domain and the small GTPase Rap1that represents a paradigm for how small G-proteins can bind andrecognize FERM domains (Gingras et al., 2013; Li et al., 2012).

Rap1 binding inhibits the binding of KRIT1 to microtubules(Beraud-Dufour et al., 2007), thereby enabling the relocalizationof KRIT1 and the stabilization of cell–cell junctions (Liu et al.,

2011). The molecular mechanisms by which this occurs are notyet known, but KRIT1 binds Rap1 and microtubules throughdifferent domains, suggesting that there is a conformationalregulation controlling the subcellular localization of KRIT1

(discussed below). The KRIT1 FERM domain can also bind tothe membrane anchor protein heart of glass 1 (HEG1), a proteinessential for KRIT1 junction localization. The functional

importance of these interactions is highlighted in zebrafishstudies, as KRIT1 mutants that are unable to bind either Rap1 orHEG1 do not rescue the KRIT1-null (santa) phenotype, which is

associated with defects in cardiovascular development (Gingraset al., 2012; Liu et al., 2011) (see supplementary material TableS1).

Another important binding partner of KRIT1 is integrin

cytoplasmic domain associated protein-1 (ICAP1), aphosphotyrosine binding (PTB)-domain-containing protein thatnegatively regulates b1 integrin activation (Liu et al., 2013;

Millon-Fremillon et al., 2008). ICAP1 binds KRIT1 in a bidentatemode, recognizing two regions: the highly-conserved RR regionand the first of the three KRIT1 NPxY/F motifs (Liu et al., 2013).

Importantly, ICAP1 uses the same binding site to interact witheither the KRIT1 NPxY motif or the cytoplasmic tail of integrinb1. As ICAP1 cannot inhibit integrin activation when it is bound

to KRIT1, increased integrin activation is observed whenincreasing amounts of KRIT1 are available to bind to ICAP1(Liu et al., 2013). In endothelial cells, KRIT1 also appears tostabilize the ICAP1 protein, so KRIT1 loss leads to decreased

ICAP1 levels and consequently increased b1 integrin activation(Faurobert et al., 2013). Thus, the exact role of KRIT1 inmodulating ICAP1-mediated regulation of integrin activation in

endothelial cells is complex and is still being discerned.KRIT1 has also been linked to several other important

signaling pathways. KRIT1 overexpression leads to increased

expression of HEY1 and DLL4 (indicative of Notch activation)

Box 1. The Genetics of CCM

Familial CCM accounts for only ,20% of cases but tends to bemore severe than sporadic CCM (Cavalcanti et al., 2012), withpatients exhibiting multiple lesions and increased hemorrhagerates. Familial CCM is associated with a heterozygous germlineloss-of-function mutation in KRIT1, CCM2 or PDCD10 (Cavalcantiet al., 2012). Malformation development appears to require a localsecond hit to remove the remaining wild-type copy of the CCMgene (Akers et al., 2009; Dammann et al., 2013; Pagenstecheret al., 2009; Stahl et al., 2008), but other factors in theneurovasculature microenvironment could potentially also have arole in lesion formation (Boulday et al., 2011; Dammann et al.,2013). Local mutations in CCM genes also cause sporadic CCM(Limaye et al., 2009), and there is a growing list of disease-causingmutations, most of which are nonsense mutations leading to a lossof function. In 19 out of 20 of familial cases and two thirds ofsporadic cases with multiple lesions, mutation in at least one of theCCM genes has been identified (Cavalcanti et al., 2012; Riantet al., 2013). In lesions with identified mutations, KRIT1 is mutatedin 65% of cases, CCM2 in 19%, and PDCD10 in 16% (Riant et al.,2013; Stahl et al., 2008).

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and, conversely, silencing of KRIT1 diminishes Notchsignaling (Wustehube et al., 2010). Notch signaling is linked

to phosphoinositol 3-kinase (PI3K)/AKT activation and therepression of ERK1/2. Consistent with this, loss of KRIT1 inendothelial cells leads to increased ERK1/2 phosphorylation,and CCM lesions show increased ERK1/2 phosphorylation

(Wustehube et al., 2010). Loss of KRIT1 also reduces theexpression of the reactive oxygen species (ROS)-scavengingenzyme SOD2 with consequent increases in the steady state

levels of ROS, AKT phosphorylation, and AKT-dependentFOXO1 phosphorylation (and thus inactivation) (Goitre et al.,2010). Another KRIT1 interactor, the long isoform of Nd1

(Nd1-L), an actin cytoskeleton-stabilizing protein, wasrecently identified and data suggests that its co-expressionwith KRIT1 supports SOD2 expression (Guazzi et al., 2012).

Collectively, KRIT1 is involved in multiple pathways thatprevent cell death.

The conformational organization of KRIT1 appears to beimportant for its subcellular localization and consequent

signaling. KRIT1 is thought to harbor intramolecular bindingsites, i.e. its N-terminus can interact with its FERM domain(Beraud-Dufour et al., 2007; Francalanci et al., 2009). It is

therefore possible that KRIT1, like other FERM proteins,adopts both open and closed conformations through a ‘head-to-tail’ interaction. The head–tail interaction probably occurs though

the recognition of the KRIT1 NPxY/F motifs by the FERMdomain, although the specificity of this interaction is still unclear(Faurobert and Albiges-Rizo, 2010). Changes in the conformation

of KRIT1 are thought to regulate its localization; for example,microtubule binding is associated with a presumed ‘closed’conformation (Francalanci et al., 2009) and ICAP1 binding isassociated with a presumed ‘open’ conformation (Beraud-Dufour

et al., 2007). The functional and signaling importance of KRIT1conformational changes has not yet been adequately explored.

CCM2CCM2 is a scaffolding protein with no enzymatic activity and anexpression pattern very similar to that of KRIT1, including in

arterial endothelial cells of multiple tissues (Petit et al., 2006;Seker et al., 2006). It acts as the hub of the CCM complex bysimultaneously binding both KRIT1 and PDCD10, in addition toa number of other signaling proteins (Hilder et al., 2007a). CCM2

contains a predicted PTB domain at its N-terminus (Liquori et al.,2003) and has recently been shown to contain a helical domain atits C-terminus termed the harmonin-homology domain (HHD)

(Fisher et al., 2013) (see poster).CCM2 is found throughout the cell and can shuttle in and out

of the nucleus, probably through its interaction with KRIT1

(Zhang et al., 2007). However, CCM2 binding has also beenimplicated in sequestration of the KRIT1–ICAP1 complex in thecytosol (Faurobert and Albiges-Rizo, 2010). CCM2 localization

to endothelial cell–cell junctions is lost following the loss ofKRIT1 localization to cell-cell junctions, suggesting that it istargeted there by KRIT1. Indeed, when re-expressed in CCM2-knockdown cells, wild-type CCM2 localizes to cell–cell

junctions, but a mutant that cannot bind KRIT1 (CCM2-F217A)does not, implicating an interaction of the CCM2 PTB domainwith one of the KRIT1 NPxY/F motifs in junctional targeting

(Stockton et al., 2010).The CCM2 PTB domain also binds TrkA, a receptor tyrosine

kinase found on nerve cells (Harel et al., 2009). This interaction

induces cell death in neuroblastoma or medulloblastoma,

probably through the recruitment of a complex betweenPCDC10 and STK25, a member of the germinal center kinase

III (GCKIII) group of serine/threonine kinases (Costa et al.,2012). How it induces cell death, and whether this pathwaycontributes to the CCM phenotype, remains unclear.

CCM2 is also known as osmosensing scaffold for MEKK3

(OSM) because it binds the mitogen-activated protein kinase(MAPK) kinase kinase MEKK3. It is required for hyperosmolar-induced p38 MAPK activation, and upon osmotic shock a

significant portion of CCM2 relocalizes to membrane ruffleswhere CCM2 is thought to scaffold RAC1 and MEKK3 in the p38MAPK cascade (Uhlik et al., 2003; Zawistowski et al., 2005).

CCM2 binds F-actin in in vitro binding assays, suggesting that itorganizes a complex that is capable of linking RAC1-dependentactin reorganization to p38 activity (Hilder et al., 2007b). Recent

work confirms that osmotic stress elicits a response of theCCM2–RAC1 pathway, but indicates that the signaling mightoccur through phospholipase C (PLC)c1 (Zhou et al., 2011).Conversely, another study suggests that CCM2 loss does not

affect the p38 MAPK pathway, but rather affects JNK and MAPKkinase (MKK) signaling (Whitehead et al., 2009). The role ofCCM2 in MAPK pathways is clearly complex, and a better

understanding will require further study.A significant recent advance in the field was the identification

of CCM2-like (CCM2L), a protein with high sequence identity to

CCM2 that is selectively expressed in activated endothelial cells(Zheng et al., 2012). Loss of zebrafish ccm2l phenocopies theccm2-null (valentine) phenotype (supplementary material Table

S1) and can be partially rescued by overexpression of CCM2(Rosen et al., 2013). However, there is not complete overlap infunctions, as CCM2L competes with CCM2 for binding toKRIT1, but not PDCD10, and functionally blocks CCM2-

mediated junctional stability (Zheng et al., 2012). WhetherCCM2L is linked to human disease remains to be determined.

PDCD10PDCD10 is ubiquitously expressed and contains an N-terminaldimerization domain (Kean et al., 2011; Li et al., 2010) and a C-

terminal focal adhesion targeting-homology (FAT-H) domain (Liet al., 2010) (see poster). It binds a variety of proteins includingCCM2 (Hilder et al., 2007a; Voss et al., 2007), the GCKIII serine/threonine kinases (Fidalgo et al., 2010; Xu et al., 2013; Zhang

et al., 2013a), paxillin (through its FAT-H domain) (Li et al.,2011), FAP-1 (also known as PTPN13) (Voss et al., 2007),protocadherin-c (Lin et al., 2010), VEGFR (He et al., 2010),

UNC13D (Zhang et al., 2013b) and striatin (through its FAT-Hdomain) (Goudreault et al., 2009; Kean et al., 2011). It also bindsphosphotidylinositides (Dibble et al., 2010; Ding et al., 2010) (see

poster).The PDCD10 FAT-H domain interacts with CCM2 (Li et al.,

2010), and PDCD10 is the third component in the heterotrimeric

KRIT1–CCM2–PDCD10 complex (Hilder et al., 2007a). Itsfunction in the CCM complex is still being explored, butPDCD10 also has roles outside of this complex. The bestcharacterized of these involves dimerization-domain-mediated

interactions with the GCKIII group of protein kinases, MST4/MASK, STK24/MST3 and STK25/YSK1/SOK1 (Sugden et al.,2013). PDCD10 predominately resides within the striatin

interacting phosphatase and kinase (STRIPAK) complex whereit binds to GCKIII kinases (Fidalgo et al., 2010). PDCD10 alsobinds striatin directly and thus complexes with protein

phosphatase 2 (PP2) and other STRIPAK components

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indirectly, thereby linking GCKIII kinases to phosphatase PP2(Goudreault et al., 2009). Alternatively, PDCD10–GCKIII

localizes to the cis face of the Golgi complex through aGCKIII interaction with GOLGA2, an interaction mutuallyexclusive to the PDCD10–GCKIII interaction with STRIPAK.The PDCD10–GCKIII interaction is important for GCKIII

localization as loss of PDCD10 shifts the binding of GCKIIIkinases from the STRIPAK complex to the Golgi (Kean et al.,2011), leading to a loss of STK25 kinase activity, decreased

protein stability and ultimately Golgi disassembly. Cells depletedof PDCD10 are impaired in repositioning both the Golgi complexand the centrosome towards the leading edge, which impairs cell

migration (Fidalgo et al., 2010). Conversely, overexpression ofPDCD10 or PDCD10–MST4 coexpression leads to increased cellmigration, whereas mutants that are unable to bind one another do

not (Zhang et al., 2013a). As directed migration is essential forblood vessel formation, its perturbation could lead to the vascularmalformations found in CCM. PDCD10 also plays a role inexocytosis, and recent work shows that loss of either PDCD10 or

STK24 increases neutrophil exocytosis owing to a loss ofinteraction with UNC13D, a regulator of vesicle fusion (Zhanget al., 2013b). This potentially implicates changes in exocytosis in

the defective tubular morphology observed in CCM disease.CCM lesions have poor endothelial integrity. Appropriate

regulation of cell death is essential for maintenance of endothelial

integrity and, as its name suggests, PDCD10 is thought to play arole in cell death. However, a specific role for PDCD10 in cellsurvival is not clear as both pro-survival and pro-apoptopic

effects have been reported. PDCD10 was originally discovered tobe upregulated during granulocyte apoptosis (Wang et al., 1999)and has been linked to several different signaling pathwaysinvolved in survival. For example, PDCD10 complexes and

colocalizes with VEGFR2, and loss of PDCD10 leads todecreased stability of the VEGFR2 protein (He et al., 2010).PDCD10 also appears to induce VEGFR2 endocytosis after

VEGF stimulation (He et al., 2010) and is required for therelocation of MST4 to the cell periphery after oxidative stress,where it phosphorylates and activates ERM proteins, thereby

promoting cell survival (Fidalgo et al., 2012). Conversely,PDCD10 expression has been linked to cell death (Chen et al.,2009; Lin et al., 2010; Zhu et al., 2010) and loss of PCDC10 hasbeen reported to increase survival and proliferation, possibly

through reduced Notch signaling, enhanced VEGF signaling, orincreased ERK activity (Louvi et al., 2011; You et al., 2013; Zhuet al., 2010). How these conflicting observations can be

reconciled remains to be determined.

The role of CCM proteins in RhoA–ROCK signalingThe first indication that RhoA dysregulation might contributeto CCM pathology came from the observation of increasedstress fiber formation (a sign of activated RhoA) after RNA-

interference-mediated knockdown of any of the CCM proteins(Crose et al., 2009; Glading et al., 2007; Stockton et al., 2010;Whitehead et al., 2009; Zheng et al., 2010). Consistent with this,activated (GTP-bound) RhoA is increased in KRIT1-, CCM2- or

PDCD10-deficient endothelial cells. One of the primary effectorsof activated RhoA is the serine/threonine kinase ROCK, whichincreases actomyosin contractility by phosphorylating and

inhibiting the myosin light chain (MLC) phosphatase.Knockdown of KRIT1, CCM2 or PDCD10 increases theamount of phosphorylated MLC, whereas treatment with

ROCK-inhibitors reverses this increase and the stress fiber

accumulation (Borikova et al., 2010; Stockton et al., 2010;Whitehead et al., 2009), confirming a role for RhoA–ROCK in

CCM signaling.How CCM proteins influence RhoA has still not been fully

elucidated, but a recent report implicates b1 integrin signaling(Faurobert et al., 2013). In addition, CCM2 appears to direct the

degradation of RhoA through a CCM2 PTB-domain-mediatedinteraction with the E3 ubiquitin ligase Smad ubiquitin regulatoryfactor 1 (SMURF1) (Crose et al., 2009). Notably, loss of CCM2

leads to increases in RhoA, but not of other SMURF1 substrates,suggesting that CCM2 might selectively promote SMURF1-mediated RhoA degradation. Increased RhoA activity is also seen

when the PDCD10-binding partner STK25 (a GCKIII serine/threonine kinase) is knocked down (Zheng et al., 2010),suggesting that membrane-localized actin-associated complexes

that are regulated by the PDCD10–STK25–ERM pathway mightultimately control RhoA activation. Cells that lack KRIT1,CCM2 or PDCD10 are defective in migration, invasion, three-dimensional tube formation and maintenance of a monolayer

permeability barrier. Each of these functions can be rescued byROCK inhibition (Borikova et al., 2010; Stockton et al., 2010;Whitehead et al., 2009). ROCK inhibitors also rescue

lipopolysaccharide-induced vascular leak in KRIT1- andCCM2-deficient mice (Stockton et al., 2010) and inhibit theformation of vascular lesions in other mouse models (McDonald

et al., 2012). Targeting RhoA–ROCK signaling is therefore onepotential therapeutic strategy for CCM (Li and Whitehead, 2010).

Vascular polarity and permeability in CCMsIn animal models, KRIT1, CCM2 and PDCD10 are essential forcardiovascular development (supplementary material Table S1).Loss of either KRIT1 or PDCD10 leads to an induction of

angiogenesis by impaired Delta–Notch signaling, and PDCD10might be essential for venous endothelial cell differentiation(Wustehube et al., 2010; You et al., 2013; Zheng et al.,

2010). Additionally, KRIT1 deficiency disrupts the junctionallocalization of the TIAM–PAR3–PKCf polarity complex(Lampugnani et al., 2010), impairing directed migration and

vascular lumen formation. PDCD10 is also important forendothelial cell polarization in directed migration through itseffects on Golgi positioning (Fidalgo et al., 2010), further linkingPDCD10 with vascular development.

The leaky vasculature in CCM lesions is explained by theirweak and disordered cell–cell junctions (see poster). Theimportance of KRIT1 in cell–cell junctions is underscored

by its association with the junctional proteins VE-cadherin,a-catenin, b-catenin, AF6 (afadin, also known as MLLT4)and p120-catenin (Glading et al., 2007). Loss of KRIT1 reduces

b-catenin and VE-cadherin at cell–cell junctions, leading toincreased nuclear b-catenin and upregulation of its transcriptionaltargets (Glading and Ginsberg, 2010). Activation of Rap1 (which

stabilizes KRIT1 in junctions) inhibits b-catenin transcription in aKRIT1-dependent manner. Interestingly, loss of CCM2 also leadsto loss of KRIT1 from cell–cell junctions and theirdestabilization, a phenotype that is rescued by a mutant CCM2

that cannot bind KRIT1 (Schneider et al., 2011; Stockton et al.,2010). Recent data suggest that extracellular matrix (ECM)remodeling could also contribute to CCM pathology (Faurobert

et al., 2013). As noted above, loss of KRIT1, CCM2 or PDCD10increases RhoA-dependent contractility; this results in abnormalremodeling of ECM, altered integrin signaling and further

increases in cell tension, which increases destabilization of

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cell–cell junctions. This may explain the abnormal ECM patternsseen in CCM lesions.

Exciting new data show that the loss of KRIT1 or PDCD10 leadsto increased expression of mesenchymal markers in endothelialcells. This endothelial–mesenchymal transition (EndMT) is due toincreased BMP6–SMAD signaling (Maddaluno et al., 2013), a

pathway that is also upregulated in patient samples with KRIT1 orCCM2 mutations. EndMT is characterized by loss of polarity,increases in cell proliferation or migration and changes in cell–cell

junctions, characteristics frequently seen in CCM lesions,suggesting that EndMT might contribute to CCM initiation andprogression. It will be interesting to see whether BMP6 or TGFbinhibitors will be effective therapeutics for patients with CCM.

ConclusionsDespite significant progress in determining the genetics of CCM andthe functions of CCM proteins, many questions remain. The relevantsignaling pathways in which CCM proteins participate in endothelialcells have not been fully established and the significance of the

trimeric CCM protein complex remains controversial. Because ofthe disease phenotype, much work has focused on the roles of CCMproteins in the vasculature, but CCM proteins are widely expressed

and why their loss results predominantly in a neurovasculaturephenotype requires further study. It is clear, however, that furtherstudies are necessary before we can link the known functions of

CCM proteins to CCM formation.

AcknowledgementsSpace limitations preclude detailed discussion of all reported CCM interactionsand we apologize to colleagues whose work is not cited.

Competing interestsThe authors declare no competing interests.

FundingK.M.D. is supported by an American Cancer Society post-doctoral fellowship.O.S.F. is funded by a National Science Foundation Graduate ResearchFellowship. T.J.B. and D.A.C. are funded by the National Institutes of Health.Deposited in PMC for release after 12 months.

Supplementary materialSupplementary material available online athttp://jcs.biologists.org/lookup/suppl/doi:10.1242/jcs.138388/-/DC1

Cell science at a glanceA high-resolution version of the poster is available for downloadingin the online version of this article at jcs.biologists.org. Individualposter panels are available as JPEG files athttp://jcs.biologists.org/lookup/suppl/doi:10.1242/jcs.138388/-/DC2.

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