Integrins

Integrins.

E Ruoslahti

J Clin Invest.

1991;

87(1)

:1-5.

https://doi.org/10.1172/JCI114957

.

Research Article

Perspectives

Integrins

Erkki Ruoslahti

Cancer Research Center, La Jolla Cancer Research Foundation, La Jolla, California 92037

Introduction

Integrins are afamily ofcellsurfaceproteins that mediate cell

adhesion. Adhesion is of fundamental importanceto a cell;it

provides anchorage, cuesformigration, and signals for growth anddifferentiation.There are twoprincipaltypesofcell

adhe-sion: cell-extracellular matrix adhesionand cell-celladhesion.

Integrins appear to be theprimarymediatorsof cell-extracellu-larmatrix adhesion, and they also serve as one ofthe many

families of molecules active in cell-cell adhesion.

The past couple ofyearshave seen a virtual explosion of work done on theintegrins,and this effort has made them the

best understood celladhesion molecules.Anumberof factors contributedto this fast progress. First, the discovery of

inte-grinsbrought together a largenumber of separate observations. The integrin family ofreceptors was discovered in the

mid-1980swhen it was realizedthat a group of chicken adhesion

proteins, the platelet protein gp

H1b/Ila,

a group of lympho-cyte adhesion proteins, the VLAfamilyof cell surface anti-gens,andreceptors for fibronectinandvitronectin all had

re-latedstructuresandactivities.Secondly, the integrin work was preceded by many yearsofdetailed work on the extracellular

matrix proteinsthatintegrinsare the receptors for and, thirdly,

theobvious importance of integrins fora numberofaspects of

biologyandmedicinebrought many new investigators into the

field. The name integrinwascoinedtosignify the presumed roleofthese proteins in integrating the intracellular cytoskele-tonwiththe extracellular matrix.

Aside from their biologicalimportance to fundamental cel-lular processes, the medical importance of theintegrinsis rap-idly being realized as well; integrins have been found to play a rolein platelet aggregation, immune functions, tissue repair, and tumorinvasion, and some diseases are already known to

be caused by mutations in integringenes. Moreover,

knowl-edge of thetargetamino acidsequenceformany integrins, the

Arg-Gly-Asp (RGD)' sequence, can be exploited to design compoundscontrollingcelladhesionfor therapeutic purposes.

This reviewsummarizes some ofthe latest developments in the

field.

Integrin

diversity

Integrinsare a

family

of membrane

glycoproteins consisting

of

two

subunits,

aand

fl.

The

primary

structureofmanyofthese

Receivedforpublication 7 September 1990.

1.Abbreviationsused in this paper: LAD, leukocyte adhesion

defi-ciency;RGD, Arg-Gly-Asp sequence;

_TGF-#l,

transforming growth

factor-#.

subunitshas been deduced fromsequencingofcomplementary

DNA(reviewedinreference 1). This sequence information is

thebasisofthegeneralmodel for the structure and interactions ofintegrins depictedinFig. 1. Theligand-bindingsite of

inte-grins appears to beformedbysequences from both subunits (seereference 2, 3), and theircytoplasmicdomains form

con-nections with the cytoskeleton (see reference 4). Theseproperties endowintegrinswith theabilityto serve as a

linkbetween the cytoskeleton and the extracellular matrix. There are 11 asubunitsand 6

_ft

subunits knownatthis time

thathave been at leastpartially sequencedandtherebyshown tobe distinct(1, 5-7).Theaand

_fl

subunits in various

combina-tionsformatleast 16integrins (Fig. 2).It islikelythatmorewill bediscovered.

It has recently become clear that inadditiontoeach

_[l

being abletoassociatewithmultiple a's, asingleasubunitcan be-comepairedwith more than one

_fl.

Thea,subunit appears to beparticularly versatile;it combines with different

_f

subunits

(see reference

3)

to

comprise

asmanyasfour

integrins.

This

diversity of the integrins providescells with variedcapabilities

torecognize adhesive substrates.

Integrin

expression

in

cells

Thecomplement ofintegrins expressed bydifferent cell types

variesgreatly.Cultured mammalian cell linespossessfromtwo

to 10 different

integrins

(e.g., reference

3).

Some

integrins

are clearlycell

type-specific.

Themost

striking examplesaregpIIb/IIIa,which isexpressed exclusively by megakaryocytes and

platelets (8),

andLFA-1, Mac-1, and, p150/95, whichareexpressed only by leukocytes

(9).

The

a6,f4

integrinis specific forepithelial cellsand tumorsderivedfrom

them(10).

Theexpression of individual integrinsappearstobe

regu-latedduring development in

Drosophila

( 11) andinvertebrate

species;

agents thataffect growthanddifferentiationcan modu-lateintegrin expression.

Transforming

growth

factor-:

(TGF-fl),

for example,causesa

striking

upregulation

of certain

inte-grins(12). Thepropertemporalexpression of thecorrect

com-plement of integrinsmay makeitpossible for cellstofind their

appropriate

adhesivesubstrates in thebody. Integrin

ligands

and the RGD

sequence

Many integrins bind to extracellular matrix proteins and thereby mediatecell-extracellularmatrixinteractions.Among

the extracellularmatrix

ligands

for integrnnsare

fibronectin,

fibrin(ogen), laminin,

various collagens,

entactin, tenascin,

thrombospondin,

von Willebrand factor, and vitronectin

(1, 13).

Otherintegrins bindtocell membrane proteins

("counter

receptors"),mediatingcell-celladhesion.Theintercellular

ad-hesionproteins

ICAM-l

and ICAM-2 have beenidentifiedas

"counter receptors" for the leukocyte integrinLFA- 1 (also

known as

CDl

la/CD

18 or

_alfi2)

(9),and the counter receptor

J.Clin. Invest.

©The AmericanSociety forClinical Investigation, Inc.

0021-9738/91/01/0001/05 $2.00

CYTO SKELETAL

CONNECTIONS

1. 2.

a5

_P1

FIBRONECTIN

3.

aL q

4

CELL MEMBRANE

P2

CAM-I

Figure1.Integrin-structure, interactions, and the three bindingmodes of variousintegrins.

for thea4,1integrin is VCAM-1(14). Thea4,f1integrin shows

aninteresting dual specificity in that itcanalsobindto fibronec-tin(15). ICAM- 1, ICAM-2, and VCAM-1 aremembers of the

immunoglobulin superfamily, manyofwhichareadhesion

proteins.

Inathird mode of interaction, the major integrin in

plate-lets,gpIIb/IIIa, promotesthe binding ofplateletstoone

an-otherthrough soluble, multivalent mediator molecules.

Fibrin-ogen and von Willebrand factor function as the primary li-gands forgpIlb/lIlainplatelet aggregation, but this integrin also bindstofibronectinand vitronectin(8).These latter

inter-actionsmaybe important for the adhesion of activated

plate-letstothesubendothelial matrix.

The recognition site formanyof theintegrins that bindto

extracellular matrix andplatelet adhesion proteinsisthe

tri-peptide RGD(13). First identifiedin fibronectin,it has since

beenshowntobeacellularrecognitionsequenceinmany

ex-tracellularmatrixandplateletadhesionproteins (Fig. 2). The

conformationof theRGDsiteappearstodetermine which in-tegrinanRGD proteinorRGDpeptidewillbind (13). Short synthetic peptides containing the RGDsequence can be

de-signedtoexhibit varying integrin specificities by restrictingthe

conformationofthepeptide through cyclization.An

RGD-re-latedsequenceinthefibrinogen ysubunitKQAGDmayform

astructurethatresembles RGD,becausepeptides containing

thesesequencesbind essentially interchangeablytoplateletgp HIb/flia(8).

Asequenceentirely different from RGD andKQAGD has

been identifiedasthetargetsequence of the

a4ft

integrin in

fibronectin(15). Thissequenceispresentinoneofthe

alterna-tively splicedsegmentsoffibronectin ( 16). As discussed below, thepeptides reproducing the integrin binding sitesmayprovide anovel classoftherapeuticagents.

Regulation of integrin

activity

and specificity

The main platelet integrin gp Ilb/Ila requires activation to

bind toits ligands. Thisintegrin is presentatthe surface of resting platelets,butnoaggregation results,eventhough

fibrin-ogenand other ligandsareavailable (8, 17). It isnotknown howactivation ofthe plateletsarousesthebindingactivity ofgp Ilb/lIla. Theother platelet integrinsmay not require activa-tion, because unactivated platelets attachtofibronectin,

la-minin, and collagen.

Theleukocyte integrin LFA-l is also activatable. Interest-ingly, the ligation of theTcellreceptorcausesthe activation of

thisintegrin in lymphocytes (18). Other integrins suchasthe

as5, fibronectin anda6fi laminin receptor,whichare

constitu-tively activated in manytypes ofcells,arealsocontrolled by

activation in leukocytes (19). Itmay beimportant for blood

cellstocontrol theiractivation in thismanner, so that their

03

FIBRINOGEN

?s

r0

a2 coil Coll IV LM

LM

ca'll I

a3

PS

(6

Coll IV

Coll I

LM VN

ROD

FN

ROD

FN

I

Fn alt.

a4

Peyers Patch Addressin

P

FN

a5

aXv

FB

vWF VN OP BSP I \LM

a6

BM

34

aI~

/FB vWF VN FN

(3

(2

ICAM-1 ICAM-2

CMb

FX

FB

a

L

Figure 2. Integrinfamily.The known

subunits, the subunitcombinationsthat form the knownintegrins, andthe known ligands for theseintegrinsareshown.Also shownis theRGD specificityof those integrins that bindtothissequence. The newlyidentified ,6subunit has been tentativelyassignedtothe_a,group because its amino acid sequence ismost

homologous with,B3 (8). FN, fibronectin;

VA", vitronectin; FB, fibrinogen;LM, laminin; vWF, von Willebrand factor;

COLL, collagen;OP, osteopontin;BSP1,

bonesialoprotein 1;

ICAM-J,

ICAM-2,

intercellular adhesionmolecules;FX,factor X; BM, basementmembrane;C3bi,

complement componentC3bi;Fnalt,

fibronectinalternatively spliced domain.

circulation throughthebody isnot

impeded

until

they

become

stimulatedatthesiteofan

injury

orbysomeother

activating

event.

Cells may alsoregulate

integrin

specificity.

The a2X1 inte-grin in platelets isacollagenreceptor, but insomeothercells,it

bindstolaminin and fibronectin in additiontocollagen (20). Elucidation ofthe molecular mechanisms of

integrin

activa-tion is one of the most

important

goals of research on these

proteins.

The

gp

IIb/IIIa

integrin in

platelet function

TheroleofgpIlb/lIla ismostvividly illustratedby thedisease

thatiscausedbyahereditary

deficiency

ofthisreceptor, Glanz-mann'sthrombasthenia(reference 8, 17). Plateletsfrom

indi-vidualswith this trait failto aggregate in response toactivation. This establishesthe roleofgpIIb/IIIaastheprimary mediator of plateletaggregation.

GpIlb/Illa isanattractivetargetfor

therapeutic

manipula-tion of platelet

aggregation.

Monoclonal antibodiesthat

neu-tralize the

activity

ofgpIIb/IIIa

provide

a

possible

inhibitor of

plateletaggregation (21). RGDpeptides, orperhapspeptides containing the related KQAGDVsequence(8), mayofferan

alternative for anti-gp

Ilb/Illa

antibodiesintherapeutic

sup-pression ofthe gp

Ilb/IIla

activity. Cyclization of certain

syn-thetic RGD-containing peptides hasyieldedcompounds that have upto5,000-fold increased affinities forgpIlb/II1a relative

tothelinear peptides witha concurrentdecrease inaffinity for

otherRGD-dependentintegrins (reference22,Pierschbacher,

M.D., personalcommunication). Suchpeptides, therefore, can

serve asefficientandspecific inhibitors ofplateletaggregation. Highly active RGDpeptidesalso exist in nature. Certain snakevenomscontainshortproteinsthathave anactiveRGD sequence inahighlyconserveddisulfideloop and that are very potent inhibitors ofplatelet aggregation (23). These proteins havebeen named

"disintegrins"

to denotetheir ability to

in-hibit gp

Ilb/IIla

and otherintegrins. The disintegrins would appear less suitable as therapeutic agents than the synthetic

peptides,

becausetheylack the

specificity

of the

peptides

de-signedasinhibitorsofgp

Ilb/Illa.

Leukocyte

integrins

Integrins,alongwith othertypesofadhesion

molecules,

play

an

important roleinthefunctionsof the varioustypesof

leuko-cytes; in general theyappeartomediate the attachment that

accompanies the conversion of leukocytes from

circulating

cellstoadherenttissue cells.This

typically happens

inatissue

injury.Leukocytesbindtotheendothelium inan

injured

tissue

as aresultof increasedadhesiveness induced

by

the

injury;

the

lB2 integrins

areactivated in the

leukocytes,

and the endothe-lium expresses increasedamounts of ICAM-1. Mac-I

(aM#2,

CDl lb/CD 18) binds to

fibrinogen

and some other

proteins

notnecessarilypresent onendothelial cells (24). However, this integrinand

p1

50/95

_(a,,#2,

CDl

Ic/CD18) probably havecell surface ligands as well andare important in neutrophil and monocyteadhesionandextravasation.

Themostconcretedemonstration oftheimportantrole the I2

integrins play

in

leukocytes

comesfrom the

hereditary

con-dition known asleukocyte adhesion deficiency or LAD(9). This disease iscausedby the lackofafunctional

_lt2

subunit, the

commonsubunitofLFA- 1, Mac- 1, and p150/95. Thedisease isprimarily characterizedby adefect in leukocyte

extravasa-tion,

resultinginaninability of the patienttofightinfections.

Because atleast LFA- 1 isalso involved in various aspects of

immunerecognition, lackofsuch afunction must also play a role in LAD.

Leukocytesalso haveintegrinsother than thoseofthe 62

family.

The

_fB

family of integrins thatincludes fibronectin, laminin,andcollagen receptors was identified as a protein

fam-ily ofunknown functionin lymphocytes. The name VLA for verylateantigenswasgiven to this group of proteins because

theyweregreatlyelevated in lymphocytes that had been

sub-jectedtolong-term stimulation (1). Moreover, lymphocyte

stimulation throughtheCD3 system results in theactivation of the

a5j31

fibronectinreceptor and

a6(,(

laminin receptor within

minutesfrom the activation (19), suggesting that these inte-grins also play arole intheearliestphasesofanimmune re-sponse. Inmacrophages, theligation ofthefibronectinreceptor causesupregulation ofthe complement-binding integrin, Mac-1 (25),suggesting transmission ofasignalintothe cell by the

a5#,

integrin.

An integrin

specific

for

lymphocytes

thathas anunique,

subunitmediates lymphocyte homing. This integrin,

a48p,

rec-ognizes anunknownligand onthehigh endotheliaof lymph

node venules allowing the lymphocytes to enter lymph

nodes(26).

Leukocytes also possess one or more atintegrins; the

_av

subunitcanbecome

associated

withanumber

off

subunitsto

formintegrins with related butdistinct specificities(3, 27).

Their function is not well understood, but interestingly the

avf3

integrin,or a closelyrelated integrin, appears to bind

os-teoclaststoboneby

interacting

withanRGD-containing

pro-tein knownasosteopontin (28).

Theleukocyteintegrins offernewtherapeutic possibilities.

Restoration ofthe 2integrin function bygene therapy may someday becomeatreatmentforLAD (9).On the other hand, at

times

itappears tobe

beneficial

to suppress thefunction of the

_#2

integrins.

Preventing leukocyte migrationto the areasof injury andinflammation after reperfusion oftissues isan exam-pleofonesuch situation (29) in which tissuedamage can be

reduced. Thiscanbeaccomplished byadministering monoclo-nalantibodies against

2.

ThecounterreceptorforLFA- 1,ICAM-1,is alsoareceptor

for

rhinoviruses,

theviruses thatcausethecommoncold. Solu-ble

pieces

of ICAM-1 can

inhibit

viralentryinto cells through competition with the binding of the virus tothe cell surface ICAM-1, suggestinga newtherapy for thecommoncold(9). The leukocyte integrins and

integrins

in other cells canalso serve asentry receptorsfor virusesaswellasforbacteria (30, 31). In thecase of foot-and-mouth

disease,

the entry can be

inhibited with RGD-containing peptides (30).

Thetransactivating

protein,

tat,ofthe human

immunodefi-ciency

virus (HIV) also containsanRGDsequenceandbinds

tocells inan

RGD-dependent

manner

(32).

Thismay be impo-tant,because thetatproteincan enter acellandactivate the

expression of

aresident viralgenomeaswellas act as a

growth

factor

for

Kaposi's

sarcomacells

(33).

If

RGD-dependent

bind-ing playsarole in

these

tat

protein

functions,

inhibitors could

be

readily designed.

Finally,

itmay be

possible

toblock the

binding

of osteoclaststothebonewith

RGD-containing

pep-tides.This

might

prevent bone

resorption

in diseases suchas

osteoporosis.

Integrins in tissue

repair

Thereisincreasing evidencethat the cell movementsthat take place

during

tissue

repair

such aswound

healing

depend on

integrin-mediated interactions.

Keratinocytes freshly

isolated fromnormalskin donotexpressfibronectinreceptors and do not attach to fibronectin-coated dishes. However, if

kept

in

culture

longer,

orifisolated fromwound

tissue,

these cells do expressreceptorsand do attachtofibronectin

(34).

It appears

that

keratinocytes,

when

they

closea

wound,

usethe fibronec-tinreceptorstoattachtoand

migrate

onthe

provisional,

fibro-nectin-containing

wound matrix. This

thinking

agreeswell withtheextensive literaturethatimplicates fibronectin and its

receptorsin various cell

migrations during embryonal

develop-ment (see

references

13 and

16).

Emerging applications

ofthe role ofthe

extracellular

matrix inwoundhealing includetheuse of fibronectin to facilitatethe healingof corneal ulcers (see reference 34) and the use of

syn-theticmaterials thatreproducethefibronectin cell attachment sitesas awoundhealing "glue"(Pierschbacher, M.D.,personal

communication).

Aparticularly intriguing effect ofan extracellularmatrix

molecule is theability of laminintopromotetheoutgrowthof

cellularprocesses,neurites, byneurons(13). The lackof

regen-erativecapacity in the adult centralnervoussystemcould be,at leastinpart,duetothefact that laminin isnotexpressed inthe

fully developedbrain, although it ispresentin the developing fetal brain. Indeed, implantation ofalaminin-containing

de-vice intoabrainlesioncanimprovetherestoration ofanatomi-cal connectionsacross the lesion(35) suggesting therapeutic potentialinthisapproach.

Integrins in cancer

Normal cellsdepositfibronectin,laminin,collagens, and other

extracellular matrixcomponents aroundthemselvesas a net-workofinsolubleprotein. They can then attach to this matrix

through their cell surface integrins. For reasonsthatare only

partially understood,mosttumorigeniccells, atleast inculture,

fail todeposit such a matrix or do so to alesserdegreethan

normalcells. Itisknown that the"classical"fibronectin recep-tor,the

_ashyl

integrin is needed forthematrixdeposition and its expression is oftenreducedintumorcells(36). Moreover,

in-creasing

the

expression

ofthe

as3, integrin

by gene transfer increasesthedeposition offibronectinbytumorigenicChinese

hamsterovarycells (37). However,atleastoneotherfactor,a

"matrix assemblyreceptor",is also neededfor fibronectin

de-position

(38). Thisfactor, which is absent in matrix-deficient cells, hasnotbeencharacterizedyet, but willobviouslybe an

important object of future studies.

Aconsequenceof the lack of matrix deposition isthat the tumorcellshave anaddeddegreeof freedom; their mobility is 'notlimited by adhesiontotheirown matrix.Theimportance ofthis constraint innormalcellularbehavior issuggested by

the

_a5fl,

genetransfer experiment mentioned above. The cells

expressing high

levelsofthis integrin from the transfectedgenes notonlydepositmorefibronectin matrix,buthavebecome less

migratory

than the control cells,growlesswell insoftagar,and, unlike the parental

cells,

failtoformtumorsin nude mice (37). Theexpression

of

fibronectinreceptorsandtheassembly ofa

fibronectin matrix may therefore be very closely associated with the expression of the

tumorigenic

phenotype.

There is anotheraspectoffibronectin in

malignancy,

how-ever. It appearsthat fibronectin (and extracellular matrix in general) play a dual role

malignancy:

as discussed

above,

a tumorcellshould lackitsownextracellularmatrixtobe ableto

proliferate

fastand

migrate

optimally.

However, such a cell needssome matrix adhesiontobe abletoderivetraction for migration from the matrices ofother cells.This is suggested by

thefact thattumorcellsand other

migratory

cells

preferentially

migrate

onsurfacescoatedwith

adhesive

extracellular matrix

proteins.

Moreover, the RGD

peptides

caninhibit

migration

of

tumorcells

through

tissue in invasionassays

(reviewed

in

refer-ence

39).

The RGD

peptides

canalsoaffecttumorcells in vivo.

Sev-erallaboratorieshave

published experiments

inwhich dissemi-nation of

intravenously

injectedtumorcells in mousetissues

peptide(reviewed inreference 39). The loss of adhesion result-ingfrom thepeptidetreatment maydeny the cells anchorage andtractionfor growth andmigration.Alternatively, the RGD peptides may becapableof inducing the receptors to deliver a growthinhibitorysignal into the cell;anindicationof the abil-ityof the peptides to delivera signal is that they have been shown toinduce expression ofproteases infibroblast cultures and that at high doses they also stop theproliferation of cells (39, 40). Thus, thepeptides may be receptor agonists with re-gard tosignaling, in additiontobeing inhibitors of adhesion.

These observations suggest new modesof cancer therapy; thepeptides already at hand allow modulation of invasiveness and metastasis through control ofintegrins. Research along theselines could prove extremely rewarding in that it would target invasion and metastasis rather than the properties of cancer cellstargeted bymoretraditional therapies.

Acknowledaments

Ithank Drs.WayneBorder, EvaEngvall,FilippoGiancotti, Lucia Lan-guino, David Mann,andMichaelPierschbacher forcomments on the manuscript.

Thewritingof this review and author'soriginal workaresupported by grants CA 42507, CA 28896, and Cancer Center support grant CA 30199 from the National Cancer Institute and grant HL 26838 from the National Heart, Lung, and BloodInstitute,Departmentof Health and HumanServices.

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