The structure and function of platelet integrins

The structure

and function

of platelet

integrins

L.V. Parke Department

of Pharmacology, Center for Thrombosis and Hemostasis, University at Chapel Hill, Chapel Hill, North Carolina, USA Current

Opinion

in Cell Biology

Introduction

The physiologic role of platelets is to prevent blood loss by forming a hemostatic plug at sites of vascular injury. In forming this plug, platelets undergo two basic types of adhesion processes. First, they readily adhere to and spread on the proteins of the subendothelial matrix that become exposed when a blood vessel is injured. From in vitro studies, it appears that platelet adhesion can occur in the absence of platelet activation. Second, platelets aggregate to one another in a process that requires platelet activation by agents such as thrombin, ADP, epinephrine, etc. Under pathologic conditions, the adhesive nature of the platelet causes it to form thrombi that can lead to heart attacks and strokes, which is one reason that the platelet has been so intensely studied. The adhesion and spreading of platelets on the subendothelium and platelet-platelet aggregation require that platelets bind to numerous matrix and plasma proteins. Sign&ant progress has been made over the past year in identifying and characterizing many of the platelet membrane receptors for these proteins. At least five of these receptors are members of the integrin family of adhesive receptors. This article reviews the latest developments in the structure and function of adhesion receptors on human platelets that are members of the integrin family and discusses likely directions for future research.

Platelet

integrins

The integrins are a large family of cell surface glycoprotein receptors that play a fundamental role in cell-cell and cell-matrix interactions. These receptors are widely distributed; they are present on most mammalian cells and are conserved in evolution, with immunologic evidence for their presence in lower organisms such as fungi [l]. The first member of this family to be identiiied and described was the platelet glycoprotein IIb-IIIa complex (GPIIb-IIIa). Since the best studied integrin is GPIIb-IIIa, its structure and function will be discussed as a prototype of the family, and recent developments concerning GPIIb-IIIa and other platelet integrins will be summa-

of North

Carolina

1989, 1:947-952

rized. The integrins discussed in this review will in general be referred to by names commonly used in the literature; their more recent a-p designations are given in Table 1.

Glycoprotein

IllvIlla

(all&J

Glycoprotein IIb-IIIa, like other integrins, consists of two non-covalently linked subunits. The a subunit, i.e. GPIIb, is usually the larger of the two and, in this case, has a molecular weight of 140000 under non-reducing conditions. The p subunit, or GPIIIa, has a molecular weight of 105000. GPIIb is further composed of a heavy (mol.wt = 125000) and light (mol.wt = 25000) disullide-linked chain (Fig. 1; Table 1). The two chains are formed by proteolytic processing of a single polypeptide, with 97% of the cleavage occurring at one site and 3% at an additional site in GPIIb, explaining some of the heterogeneity of these molecules [2]. Some a subunits in the integrin family are not proteolytically processed and remain as a single polypeptide, e.g. the a2 subunit of GPIa-IIa (see Table 1 for summary). Both glycoproteins IIb (Poncz et al, J Biol Gem 1987, 26284768482) and IIIa (Fitzgerald et al, J Biol cbem 1987, 262:39363939) have been cloned and sequenced with modifications in the GPIIIa sequence by Rosa et al (Blood 1988, 72:593-600). Predictions from the primary amino acid sequence are that GPIIb-IIIa contains two membranespanning domains, one near the carboxy-terminus of the light chain of GPIIb and the other near the carboxyterminus of GPIIIa. Each subunit is predicted to have a short cytoplasmic domain: 20 amino acids for GPIIb and 41 amino acids for GPIIIa. GPIIIa and other j3 subunits have four cysteine-rich repeats, which accounts for their slower migration under reducing versus non-reducing gel electrophoresis conditions. The heavy chain of GPIIb has four putative CaZ+-binding domains that have sequence homology with Ca2+ -binding regions in calmodulin. The GPIIb-IIIa complex is maintained in micromolar or greater Ca2+, and it is speculated that these Ca2+-binding domains contribute to this requirement. In support of this, the heavy chain of GPIIb and not the

Abbreviations ECMR-extracehlar

matrix

@ Current

receptor;

CPF-glycoprotein;

Science

Ltd ISSN 0955-0674

VIA-very

late antigen.

947

948

Cell-to-cell

contad .

Table

7eceptor

1. lntegrin

receptors

Other

%bP3

names

on human

platelets.

a-subunit Non-reduced

fkD) Reduced

Bsubunit fkD) Non-reduced

140

125,

105

GPllb-illa

25

Ligands

von

433

VNR

160

135,~25

105 von

Fibrinogen Fibronectin Willebrand Vitronectin Vitronectin Fibrinogen Willebrand

Function Aggregation,

adhesion

factor

I factor

FNR VLA-5 CPlc-lla ECMR VI

155

130, 25

110-130

Fibronectin

Adhesion

a2Pl

CPlaalla VLA-2 ECMR II

155

170

l-lo-130

Collagen

Adhesion

%P,

VLA-6 GPlc-lla

140

120, 30

110-130’

Laminin

Adhesion

CP, glycoprotein; ‘The g, subunit

VNR, vitronectin is probably the

receptor; FNR, fibronectin same for as, a2 and a, but

receptor; different

light chain forms the Ca2+ -dependent hetenxiimer with GPIIIa [3]. However, other members of the integrin family have similar metal-binding domains but a clear divalent cation requirement for the maintenance of these complexes has not been established. Unlike other integrins thus far studied, the genes for GPIIb and GPIIIa are on the same chromosome, i.e. chromosome 17 (Sosnoski et al, J Clin Invest 1988, 81:19931998), and are physically linked within 260 kb, suggesting that a mechanism of coordinate expression of GPIIb and GPIIIa dependent on the gene proximity maybe operative [4]. Glycoprotein IIb-IIIa mediates platelet aggregation by functioning as a fibrinogen receptor on activated platelets. The receptor activity of GPIIb-IIIa is inducible; on resting platelets GPIIb-IIIa is exposed extracellularly, yet plasma proteins do not bind to it. However, by an unknown mechanism, GPIIb-IIIa becomes a competent receptor on activated platelets. This is in contrast to many members of the integrin family which appear to be constitutively active. Glycoprotein IIb-IIIa also binds to Iibronectin, von Willebrand factor (for additional review see PhiIlips et al, Blood 1988, 71:831-843) and vitronectin (Pytela et al, Science 1986, 231:155%1562; Thiagarajan and Kelly, J Biol C&m 1988, 263:303%3038) in vitro, although fibrinogen is most likely to bind to GPIIb-IIIa in the fluid phase in vivo because it has the higher plasma concentration and competes with the other proteins for binding. Fibrinogen is multivalent, with

VLA, very late antigen; molecular weights have

ECMR, extracellular been reported.

matrix

receptor.

two identical CL,p and y chains. This multivalency is believed to allow fibrinogen to catalyze platelet aggregation by linking adjacent platelets together. All four of these plasma proteins bind to GPIIb-IIIa in a divalent cation-dependent manner. Fibrinogen binding is especially enhanced in the presence of millimolar Ca2+ and Iibronectin binding, in the presence of millimolar Mg2 + . This requirement appears distinct from the micromolar Ca2+, requirement for keeping GPIIb-IIIa in a complex. In addition to its role in platelet aggregation, glycoprotein IIb-IIIa contributes to platelet adhesion and is directly responsible for platelet spreading on the subendothelium (Lawrence and Gralnick, J Lab Clin Med 1987, 109:49%503). Weiss and co-workers [5] recently found that, at high shear rates, platelets adhere to the subendothelium by a GPIIb-ma-mediated mechanism that is independent of fibrinogen. Under these conditions, platelet adhesion was normal with platelets in plasma that lacked fibrinogen. Evidence for the role of GPIIb-IIIa was provided by the ability of anti-GPIIbIIIa monoclonal antibodies and a peptide from the carboxy-terminus of the fibrinogen y chain to inhibit adhesion. Since this peptide also inhibits the ability of other ligands to bind to GPIIb-IIIa, this Iinding suggests’ that subendothelial proteins, such as von Willebrand factor or vitronectin, are responsible for the adhesion observed at high shear rates. Progress is being made in identifying binding sites on GPIIb-IIIa for fibrinogen and other ligands. One ap-

Platelet

CPllla

Parise

GPIIb-IIIa or that the binding of each peptide to separate sites inhibits the binding of the other, possibly through a conformational change in GPIIb-IIIa. Bennett et al [6] found that Arg-Gly-Asp&r and LlO differed in their ability to inhibit the binding of two anti-GPIIbIIIa monoclonal antibodies to activated platelets, suggesting that the two peptides bind to distinct sites on GPIIb-IIIa. Using chemical crosslinking, D’Souza et al [7] reported that an ArgGly-Asp-containing peptide from libronectin becomes crosslinked to a proteolytic fragment of GPIIIa (amino acids 109-171>, suggesting that at least part of the Arg-Gly-Asp binding site is contained here. This region of GPIIIa is highly conserved in other integrins (76%) [6]. This finding was conIirmed by Smith and Cheresh (J Biol cbem 1988, 263:18726-18731), who found that an Arg-Gly-Asp peptide became crosslinked to residues 61-203 of the GPIIIa or p subunit of the vitronectin receptor from placenta.

@subunit) CPllb

integrins

(a-subunit)

The possibility has also been raised that gangliosides may contribute to or modulate the adhesive properties of GPIIb-IIIa on platelets, since a series of gangliosides inhibit the attachment of thrombin-activated platelets to fibrinogen, Iibronectin and von Willebrand factor [8]. However, the role of gangliosides in these events is poorly understood.

COOH

Fig. 1. Structure et al. (Blood

of glycoprotein 1988, 71:831-843)

COOH

(CP) Ilb-illa.

Modified

from

Phillips

preach has been to use the synthetic peptides from the platelet-biding regions of fibrinogen or iibronectin as tools in this identification. One platelet-binding region of fibrinogen consists of a 10-12 amino acid sequence at the carboxyterminus of the fibrinogen y chain (His-His-Leu-Gly-Gly-Ala-Lys-Gin-Ala-G~-~p-V~; Kloczewiak et al, Biochemistry 1984,23:1767-1~4). The 12amino acid sequence, or dodecapeptide, has been termed ‘H12’ and the decapeptide, beginning at leucine (Leu), has been termed ‘LlO’ by Plow et aL, 0 Biol CXwm 1984, 259:5388-5391). The LlO peptide, if attached to an affinity matrix, binds specifically to GPIIb-IIIa from detergent-solubilized platelets (Lam et al, J Biol C&m 1987, 262:947-950). Two additional sites in the fibrinogen u chain, at positions 95-97 and 572-574, contain the Arg-Gly-Asp sequence. This sequence was originally determined to be the active region of the much larger cellbinding domain of fibronectin (Pierschbacher and Ruoslahti, Nature 1984, 30930-33) and was subsequently identified and found to be active in many other matrix and plasma proteins. Surprisingly, Arg-Gly-Asp-containing sequences till elute detergent-solubi GPIIb-IIIa from an LlO affinity column and vice versa, suggesting either that the two peptides compete for the same site on

Of interest relative to the Arg-Gly-Asp sequence is the intriguing finding that a monoclonal antibody that recognizes GPIIb-IIIa, PACI, contains a tripeptide mimetic of Arg-Gly-Asp [ 91. PACl binding to GPIIb-IIIa is inhibited fibrinogen and by peptides containing Arg-Gly-Asp. Upon sequencing the complementarity-determining region of this antibody, it was determined that an Arg-?)lr-Asp peptide was present. This region appears to be critical to PACl recognition of GPIIb-IIIa because a synthetic peptide of 21 amino acids from this region that includes ArgTyr-Asp inhibits PACl binding to platelets and platelet aggregation. Thus, antibodies may also use variations of the ‘Arg-Gly-Asp’ motif to recognize integrins. Functions other than adhesion have also been associated with GPIIb-IIIa. Several lines of evidence indicate that GPIIIb-IIIa functions as a passive Ca2+ channel in platelets. First, platelets from patients with Glanzmann’s thrombasthenia, i.e. platelets that are lacking GPIIb-IIIa, have an abnormally low rate of plasma membrane Ca2+ exchange that can be mimicked in normal platelets by dissociating the GPIIb-IIIa complex (Brass, J Biol Cbem 1985, 260:2231-2236). Furthermore, when incorporated into liposomes, the GPIIb-IIIa complex, but not the dissociated subunits, mediates passive Ca2+ transport [lo]. The Ca2+ flux into liposomes is inhibited by the H12 peptide from the carboxy-terminus of the fibrinogen y chain, but not peptides containing the Arg-Gly-Asp&r sequence. This implies that occupancy of specific sites on GPIIb-IIIa controls Ca2+ entry in platelets. Ca2+ entry into platelets was also inhibited by the H12 peptide (Rybak and Renzulli, Circulation 1988,78:11621). In support of this, another group reported that an anti-GPIIbIIIa monoclonal antibody, fibrinogen, or (in apparent conflict to above) an ArgGly-Asp-Ser peptide inhibits Ca2+ flux

949

950

Cell-to-cell

contact

in whole platelets (Sinigaglia et al, Biochem Bioms Res Commun 1988, 154:258-264). Several recent studies suggest that ligand or peptide biiding to GPIIb-IIIa transmits signals across the platelet membrane. FerreU and Martin [ 111 report that thrombin induces three temporal waves of tyrosine phosphorylation ln platelets. The third wave appears to be mediated in part by fibrinogen binding to GPIIb-IIIa, since treatment of platelets with agents that inhibit fibrinogen binding, i.e. with thecalcium chelator EDTA, and the peptides H12 or Arg-Gly-Asp-Ser, also inhibit this phosphorylation. In addition, platelets from patients with Glanzmann’s thrombasthenia, which lack GPIIb-IIIa, exhibit the iirst two waves but not the third wave of phosphorylation. Currently, the link between fibrinogen binding to GPIIb-IIIa and tyrosine phosphorylation is not understood. However, one mechanism by which signal transduction may occur is through a l&and-induced conformational change in GPIIbIIIa Frelinger et al [ 121 obtained evidence for a rapid, reversible conformational change in GPIIb-IIIa on the platelet surface under conditions where an Arg-Gly-Asp-Ser-containing peptide, the peptide LlO from the carboxy-terminus of the fibrinogen y chain, or fibrinogen itself was bound. This conformational change was reported by a monoclonal antibody (PM&l) with an epitope near the carboxy-terminus of GPIIb. The conformational change that these ligands most likely induce is an unfolding of the GPIIb-IIIa complex, as detected by changes in the Stoke’s radius and sedimentation coefficient of purified GPIIb-IIIa in the presence of peptides from the binding domains of fibrinogen and libronectin (Parise et aL, J Biol Cbem 1987, 292:12597-12602). Platelets contain at least four other members of the integrin family. Significant progress has been made within the last year in identifying these and dehning their function.

The vitronectin

receptor

(VNR or a$,)

The vitronectin receptor is the integrin most closely related to GPIIb-IIIa; it has a p subunit with a primary amino acid sequence identical to that of platelet GPIIIa, but an ct subunit distinct from GPIIb (Table 1). In endothelial cells and melanoma cells, where this receptor has been most weU characterized, it appears to bind vitronectin, fibrinogen, and von Wiiebrand factor, but not fibronectin (Char0 et al, J Biol CXwm 1987, 26299359938; Cheresh and Spiro, J Biol Cbem 1987, 262:1770317711). The careful observations of Iam and co-workers [13] identifid a platelet protein that is related, if not identical, to the vitronectin receptor; it reacted with an antibody that recognized the vitronectin receptor a subunit and an antibody that was speciiic for the vitronectin receptor complex. In addition, the amino-terminal sequence of the a subunit was identical to that of the vitronectin receptor a subunit. Curiously, however, this protein complex was not recognized by a monoclonal antibody specific for GPIIIa, raising the possibility that the a subunit of the platelet vitronectin receptor is not identical to GPIIIa. This notion is not without prece-

dent, since Cheresh et al. [14] recently reported that the vitronectin receptor a subunit in carcinoma cells can be coupled to a distinct p subunit. Currently, it is not known whether this receptor is inducible in activated platelets or constitutively active, or whether and how this receptor contributes to platelet function.

Fibronectin

receptor

(FNR or c@,)

Platelets have at least two receptors that bind iibronectin, the activation-dependent GPIIbIIIa, and an activationindependent receptor that is probably identical to the iibronectin receptor described by Pytela et al on osteosarcoma cells (Cell 1985, 40:191-198). This receptor has also been termed the very late antigen (VLA)-5 by Hemler and co-workers, who described its presence on human T cells (Takada et al, Nature 1987, 326607609) and the extracellular matrix receptor (ECMR) VI by Carter and co-workers, who identilied it on iibroblasts [ 151. Both groups independently identilied this receptor on platelets [ 15-171 and found that it comigrated on two-dimensional non-reduced-reduced gels with what was previously known as platelet GPIc and GPIIa (Phillips and Poh-Agin, J Biol C&m 1977, 252:2121-2126). Piotrowicz et al, [I61 examined the function of this receptor on platelets. They found that normal platelets and platelets from patients with Glanzmann’s thrombasthenia adhered to fibronectin-coated surfaces in the absence of activation. This binding was inhibited by an Arg-Gly-Asp peptide, an antibody against the p1 subunit [I61 and a monoclonal antibody (PlF8) that is speci6c for either the iibronectin receptor complex or the a5 subunit [ 151. They concluded that this receptor appears to play a major role in the adhesion of non-activated platelets to fibronectincoated surfaces. This notion is supported by the experiments of Nievelstein and Sixma [ 181, who reported that GPIIb-IIIa is not important for platelet adhesion to endothelial ceU matrix proteins under flow conditions, and speculated that GPIc-IIa may contribute to this adhesion.

Glycoprotein

la-lla

(a,&)

GPIa and GPIIa were identified on the platelet surface years ago, but only recently determined to exist as a complex, identical to VIA-2 [ 171 and ECMR II 1191, and to function as a platelet collagen receptor. A monoclonal antibody (PlH5) against the a subunit of a Iibroblast collagen receptor (ECMR II) immunoprecipitates both GPIa and GPIIa, suggesting that the two glycoproteins are noncovalently linked [ 191. PlH5 also inhibits the adhesion of inactivated platelets to type I and III coUagens but not to iibronectin [19]. There appear to be two forms of GPIa on platelets, one that is labeled on the surface of resting platelets and one that is labeled only on activated platelets (Bienz and Clemetson, J Biol Cbem 1989, 264:507-514). Although the two forms have the same molecular mass and isoelectric points, they migrate to distinct positions

Platelet

on two-dimensional non-reduced-reduced gels. It is not known if any functional differences exist between these two forms or if multiple forms exist on other cell types.

(4) (5)

VIA-6 (ctJ3,) VIA-6 was initially identified on platelets by immunoprecipitation with the monoclonal antibody GoH3 against the a subunit [ 171. VLA-6 also migrates on two-dimensional non-reduced-reduced gels with platelet glycoproteins Ic and IIa, as does the fibronectin receptor. Interestingly, the a subunit of VIA-6 ((rg) appears to be less tightly bound to its p subunit than is characteristic of other integrins. VLA-6 was subsequently identilied as a relatively specific platelet laminin receptor; GoH3 blocked platelet adherence to laminin but not to fibrinogen, fibronectin, or collagen [ 201. Platelet adherence to laminin does not require platelet activation, but, like other integrins, does require divalent cations. Iaminin binding is supported by Mg2+, Mn2+, or Co2+ but not Ca2+, Zn2 + , or Cu2 + . This is different from the requirements for GPIIb-IIIa binding to fibrinogen with regard to Ca2 + .

Conclusions

0

1. 0

Parise 951

Whether platelet integrins other than GPIIb-IIIa transport Ca2+ or other ions and how this may Sect platelet function. Further definition of integrln structure-function relationships. It is especially important to identify more precisely ligand binding sites on the integrin receptors and receptor binding sites on the ligands. Knowledge of these sites could potentially lead to the development of spectic inhibitors to control the contribution of platelets to thrombotic events.

Annotated reading aa

integrins

references

Of interest Of ouwanding

and recommended

interest

MARCANTONIO EE, HYNES RO: Antibodies to the conserved cytoplasmic domain of the integrin PI subunit react with proteins in vertebrates, invertebrates, and fungi. / CM Bid

1988,

1061765-1772.

Antiserum raised against the cytoplasmic tail of the integrin & subunit is used to immunoprecipitate integrin-like molecules from vertebrates, invertebrates and fungi, demonstrating their wide phylogenetic distribution. LOFI-LJS JC, PLOW EF, JENNINGS LK, GINSBERGMH: Alternative proteolytic processing of platelet membrane glycoprotein Ilb. / Bid &em 1988, 263:1102~11028. In this study, it is determined that proteolytic processing of GPIIb into the heavy chain and light chain occurs primarily at 1 site, but at a second site <3% of the rime. 2.

The /3* and p3 classes of integrins exist on human platelets. The well studied p3 integrin, the GPIIb-IIIa complex, mainly contributes to platelet aggregation, with evidence for a role in platelet adhesion and spreading. The other & integrin, the vitronectin receptor, has an unknown function on platelets. The three p1 integrins recently identified on platelets, i.e. the fibronectin receptor, the GPIa-IIa complex, and VIA-~, all potentially contribute to platelet adhesion to the subendothelium.

Future

directions

With at least tie integrins (and several non-in&grin adhesion receptors) on platelets, future research will likely include the following.

(1) A ‘sorting out’ of how each receptor contributes (2)

(3)

to platelet adhesion or aggregation, and mechanisms that control their interaction with ligands. The role of platelet integrins in signal transduction. It is important to understand the mechanisms by which signal transduction occurs, whether platelet integrins other than GPIIb-IIIa participate in signal transduction, which events in the platelet are modulated by bound adhesive proteins such as fibrinogen, and how these events affect platelet function. The interaction of integrins with the platelet cytoskeleton and how this may regulate both integrin and cytoskeletal activity.

0

IAM SC-T, PLOW EF, GINSBERGMH: glycoprotein Ilb heavy chain forms coprotein IUa that binds arg-gly-asp 73:1513-1518. This paper shows that the GPIIb-IUa complex Arg-Gly-Asp peptides remain when the GPIIb from the complex by partial reduction. 3.

0

4. 0

BRAY PF, BAFSH G, ROSA J-P, Luo

Platelet membrane a complex with glypeptides. B&md 1989, and its ability to bind light chain is removed

XY,

MAGENI.~

E, SHIJMAN

MA Physical linkage of the genes for platelet membrane g$&pcy IIb and Illa. Proc Nut1 Acad Sci USA 1988,

The gen& that encode GPIlb and GPIIIa are located within 260 kb, sug gesting that they may be coordinately regulated. 5. 0

WEI.% HJ, HAWIGER J, RUGGEIU ZM, Tutw~o P, HOFFMANN T: Fibrinogen-independent

VT, THIAGAIL+JAN

platelet adhesion and thrombus formation on subendothelium mediated by glycoprotein IIb-Illa complex at high shear rate. / Ch Invest 1989, 83~288-297. At high shear rates, platelet adhesion to the subendothelium is normal in plasma lacking fibrinogen, suggesting that von Wfiebrand factor or vitronectin may be important. BENNETT JS, SHAWL SJ, POWER JW, GARTNER TK: Interaction of Mwinogen with its platelet receptor. Differential effects of 01and y chain fibrinogen peptides on the glycoprotein Ilb-Lila complex. / Bid Gem 1988, 2631294S12953. In studies comparing the inhibitory effects of Arg-Gly-Asp-W and a pep tide from the carboxy terminus of the fibrinogen r chain, it is concluded that these peptides bid ro spatially distinct sites on GPIIb-IlIa. 6.

0

7. a*

D’Souu

SE, GWBERG

MH,

BURKE TA,

Lw

SC-T,

PLOW EF:

Localization of an arg-gly-asp recognition site within integrin adhesion receptor. Science 1988, 242:91-93.

an

952

Cell-to-cell

contact

In this repon, a consem .+ I of GPIlla that becomes covalently crosslinked fo an Arg-Gly-Asp peptide is identied. This paper is important because the identified region contains a IikeIy binding site for fibrinogen and other Arp-Gly-Asp-containing ligands. 8. 0.

SANTOROSA: Inhibition of platelet adhesion to fibronectin, fibrinogen, and von WiIIebrand factor substrates by complex gangliosides. Bhxi 1989, 73484-489. Complex gangliosides inhibited the attachment of tbrombin-stimulated platelets fo the proteins listed above under conditions where GPUb-IIIa would be the expected receptor. The gangliosides did not inhibit attachment to von Wtiebrand factor mediated by GPIb. a non-integrin adhesion receptor, de?nonstrating the specificity of this inhibition. 9. l

TAUB R, Gouu, RJ, GARSKY VM, CIC~ARONE TM, HOXIE J, FRIEDMAN PA, SHA~I-IL SJ: A monocional antibody against

the platelet fibrinogen receptor contains a sequence that mimics a receptor recognition domain in fibrinogen. / Biol C!wn 1989, 264:25!&265. The authors find that a monoclonal antibody that inhibits fibrinogen binding M) GPIIb-IIla contains a sequence similar to Arg-Gly-Asp in its binding region, suggesting that antibodies, in addition to adhesive proteins, can use this mechanism to recognize integrins. 10. RYBAKME, RENZULULA, BRUNS MJ, CAHALY DP: Platelet glycol e proteins IIb and UIa as a calcium channel in Iiposomes. Bkxd 1988, 72:714-720. When the GPIIb-IIIa complex, but not the dissociated proteins, are incorporated into liposomes, they allow a passive infiux of calcium across the lipid bilayer. suggesting that GPIlb-IIla contributes to calcium flux in platelets. The finding that GPIlb-IIla itself mediates Ca2+ Flux is important in understanding platelet function and suggests that other integrins may have similar roles. 11. FERREUJE JR, m GS: Tyrosine-specific protein phos00 phoryIation is regulated by glycoprotein Ilb-IIIa in platelets. Proc

Natl

had

Sci us4

1989, 86:2234-2238.

In thrombii-stimulated platelets, tyrosine phosphorylarion occurs in 3 temporal waves. The third wave of phosphorylation does not occur In platelets that lack GPIllHIIa or under conditions where fibrinogen binding to GPIIb-IIIa is blocked. This finding is significant because it suggests rhar GPIIb-IIIa regulates the third wave of phosphorylation and possibly other intracelIuIar events. 12. FREUNGER AL III, IAM SC-T, PLOW EF, Shm~ M.& LOFTUS JC, me GINSBERG MH: Occupancy of an adhesive gfycoprotein receptor modulates expression of an antigenic site involved in cell adhesion. J Biol Chem 1988, 263:12397-12402. The epitope for a monoclonal antibody against the GPIlb heavy chain is expressed when fibrinogen, or synthetic peptides from the GPIlb-Illa biding domains of fibrinogen are bound to puriIied GPIIb-IIla or to platelets. This study shows that the conformation of GPIIb-IIIa changes when ligands are bound, suggesting part of a possible mechanism by which occupancy of GPIIb-IIla may a&ct other events in platelet function. La SC-T, Ptow EF, D’Souu SE, CHEIULSH DA, FREUNGER AL III, MH: Isolation and characterization of a platelet membrane protein related to the vitronectin receptor. J Bid them 1989, 2643742-3749. When platelet lysa@sare chromatographed over an Arg-Gly-Asp aBinity column&e platelet titronectin receptor, along with GPIIb-IIla, is retained and specihcally eluted, thus allowing its idenrification. 13. l

GINSBERG

14. l e

CHERE~H DA, SMIIH JW, OXPER HM, QU~A V: A novel vitronectin receptor integrin (St A) is responsible for distinct adhesive properties of carcinoma cells. Cell 1989,

57:5%9.

A receptor is identied on carcinoma cells that has an a subunit identical to the vitronecrin receptor a subunit, coupled to a previously undescribed p subunit. This receptor has a different l&and binding specificity than the previously described vitronectin receptor. The fact that an a subunit can combine with different fl subunits was previously unknown, and adds a new dimension of diversity to the expression and function of integrins. 15.

WAYNER

EA, CARTER WG,

Plo~~ow~u

R5, KUNICKI

TJ:

The

function of multiple ext.raceUular matrix receptors in mediating ceU adhesion to extracellular mati preparation of monoclonal antibodies to the fibronectin receptor that speciIicalIy inhibit ceil adhesion to fibronectin and react with platelet glycoproteins Ic-IIa. J Cell Biol 1988, 107:1881-1891. The ECMR VI is found to be identical to the fibronecrin receptor, VIA-5 and platelet GPIc-Ila and fo mediate platelet adhesion to fibronectin. l

16.

PIOTROW~CZ

RS, ORCHEKOW~KI

RP, NUGEM

DJ, YAMADA

KY,

KUNICKITJ: Glycoprotein Ic-IIa functions as as activationindependent fibronectin receptor on human platelets. J Ceil Biol 1988, 156:135+1364. The fibronectin receptor is shown to be present on platelets and has a mobility on 2dimensional gels identical to the GPIc-IIa complex. This receptor mediates the adhesion of unactimted platelets to Iibronectincoated surfaces, thus diaerentiaring it functionally from GPUb-ma. l

17.

HEMIER

ME,

CROUSE C, T.UADA

Y, SONNENBERG

A:

Multiple

very late antigen (VLA) heterodimers on platelets. Evidence for distinct VLA-2 ,VIA-5 (fibronectin receptor), and Vu-6 structures. J Biol &em 1988, 263:766&X65. Through a series of immunoprecipitations and peptide mappings, VIA2, MA-5 (the fibronectin receptor) and a new integrin, ~~4-6, are ident&d on platelets. l

18. l

NIEVELSTEINPFEM, %MA JJ: GIycoprotein IIB-IL4 and RGD(S) are not important for fibronecdn-dependent platelet adhesion under flow conditions. Blood 1988, 72:82-88.

These investigators show that platelet adhesion to fibronectin in matrix proteins derived from endothelial cells is not mediated by Arg-Gly-Asp or GPIIb-IIIa under Bow conditions, suggesting that other sites on fibronectin and other platelet receptors, possibly GPIc-IIa, may be involved. 19. l

KUMCKI TJ, NUGEW DJ, STAAIX SJ, ORCHEKOW~K~ EA, CARTER WG: The human fibroblast class II

RP, WAYNER

extracellular matrix receptor mediates platelet adhesion fo collagen and is identical to the platelet glycoprotein la-IIa complex. J Biol

Gem

1988,

263:4516-i519.

Glycoproteins Ia and IIa are shown to be a heterodimer complex and to function as a platelet collagen receptor. 20.

S~NNENBERG

l

receptor

A, MODDERMAN

PW,

HOGERVORST

on platelets is the integrin

F:

Laminin

VLA-6. Nature 1988,

336487-489.

is identilied as a platelet laminin receptor. The adhesion is supported by Mn2+ and Co2+, but not by the ocher divalent cations tested.

~~4-6