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Blood, 15 July 2004, Vol. 104, No. 2, pp. 299-300. This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Alert me to new issues of the journal Download to citation manager Rights and Permissions Citing Articles Citing Articles via CrossRef Citing Articles via Google Scholar Google Scholar Articles by Bennett, J. S. Search for Related Content PubMed Articles by Bennett, J. S. Related Collections Related Article in Blood Online Social Bookmarking                   What's this?  Previous Article  |  Table of Contents  |  Next Article  HEMOSTASIS, THROMBOSIS, & VASCULAR BIOLOGY Dotting the I of an I-domain Joel S. Bennett UNIVERSITY OF PENNSYLVANIA Schoolmeester and colleagues report the development of a monoclonal antibody that binds specifically to the active conformation of the I-domain of the collagen-binding integrin 21, solving the dilemma of specifying the activation state of this integrin when it mediates platelet adhesion to collagen. The ability of integrins to interact with ligands can be regulated by intracellular signaling. In the most frequently cited example, the platelet fibrinogen receptor, IIb3, only binds soluble ligands on stimulated platelets. Thus, it is not possible to measure an affinity between fibrinogen and IIb3 on resting platelets, but a dissociation constant for fibrinogen of approximately 100 nM is readily measured when platelets are stimulated by adenosine diphosphate (ADP). This change in the affinity of IIb3 for ligands can also be detected using the monoclonal antibody PAC-1, which only binds to the activated conformation of IIb3.1 Similarly, a derivative of PAC-1 named WOW-1 binds exclusively to the active conformation of v3.2 Platelet adhesion to collagen is a complex process involving 2 receptors. One receptor, glycoprotein VI (GPVI), is a member of the immunoglobulin gene superfamily that mediates the initial adhesion of platelets to collagen and, via the associated Fc receptor chain, is responsible for initiating collagen-stimulated platelet signaling. In turn, GPVI-stimulated signaling enables the I-domain of the second receptor, 21, to bind firmly to collagen, stabilizing platelet adhesion to this substrate.3 Subsequent21-initiated signaling serves to amplify the GPVI-initiated signals.4 Heretofore, it has been difficult to directly demonstrate an agonist-induced change in the affinity of 21 for soluble collagen and impossible to demonstrate a change in 21 affinity when platelets adhere to collagen-coated surfaces. Interaction of IAC-1 to platelets deposited on collagen under flow. See the complete figure in the article beginning on page 390.   To remedy these problems, Schoolmeester and colleagues (page 390) produced monoclonal antibodies against a chimeric protein composed of the 2 I-domain and maltose-binding protein. After selecting for antibodies that bound to the I-domain, they identified one, integrin activated conformation-1 (IAC-1), that only bound to platelets stimulated by the GPVI agonist convulxin. But they also found that IAC-1 bound to platelets stimulated by ADP, U46619 [GenBank] , and thrombin. These observations confirm that GPVI-initiated signals shift the 2 I-domain from its inactive to its active conformation and show that the effect of platelet agonists on integrins is not confined to the 3 subfamily. The crystal structure of a 21-residue collagen peptide bound to the 2 I-domain revealed that 3 loops on the upper surface of the I-domain coordinate a metal ion and bind to collagen.5 However, IAC-1 binds to an epitope on the opposite side of the I-domain. This means that platelet stimulation induces a global change in the I-domain conformation. More important from the standpoint of future experimentation, it means that collagen and IAC-1 can bind to the I-domain simultaneously, enabling the antibody to detect activated 2l when platelets adhere to a collagen-coated surface. Platelet adhesion to collagen is an essential facet of platelet function in vivo. IAC-1 has already provided new insights into this process, and it promises to be an exquisite tool for future studies. References Shattil SJ, Hoxie JA, Cunningham M, Brass LF. Changes in the platelet membrane glycoprotein IIb-IIIa complex during platelet activation. J Biol Chem. 1985;260: 11107-11114.[Abstract/Free Full Text] Pampori N, Hato T, Stupack DG, et al. Mechanisms and consequences of affinity modulation of integrin v3 detected with a novel patch-engineered ligand. J Biol Chem. 1999;274: 21609-21616.[Abstract/Free Full Text] Nieswandt B, Brakebusch C, Bergmeier W, et al. Glycoprotein VI but not alpha2beta1 integrin is essential for platelet interaction with collagen. EMBO J. 2001;20: 2120-2130.[CrossRef][Medline] [Order article via Infotrieve] Chen H, Kahn ML. Reciprocal signaling by integrin and nonintegrin receptors during collagen activation of platelets. Mol Cell Biol. 2003;23: 4764-4777.[Abstract/Free Full Text] Emsley J, Knight CG, Farndale RW, Barnes MJ, Liddington RC. Structural basis of collagen recognition by integrin alpha2beta1. Cell. 2000;101: 47-56.[CrossRef][Medline] [Order article via Infotrieve] CiteULike    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this? Related Article in Blood Online: Monoclonal antibody IAC-1 is specific for activated 21 and binds to amino acids 199 to 201 of the integrin 2 I-domain Anne Schoolmeester, Karen Vanhoorelbeke, Shinya Katsutani, Hilde Depraetere, Hendrik B. Feys, Johan M. W. Heemskerk, Marc F. Hoylaerts, and Hans Deckmyn Blood 2004 104: 390-396. [Abstract] [Full Text] [PDF] This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Alert me to new issues of the journal Download to citation manager Rights and Permissions Citing Articles Citing Articles via CrossRef Citing Articles via Google Scholar Google Scholar Articles by Bennett, J. S. Search for Related Content PubMed Articles by Bennett, J. S. Related Collections Related Article in Blood Online Social Bookmarking                   What's this?

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