From www.bloodjournal.org by guest on September 11, 2016. For personal use only.

this study has implications for establishing a diagnosis of LCH and for identifying patients who might benefit from targeted therapies. Although ARAF mutations appear to be uncommon in LCH, they are straightforward to test for and are clinically actionable. As mutations in other genes are identified and functionally validated, it should be feasible to develop a targeted molecular diagnostics panel for LCH that includes BRAF, ARAF, and other driver genes. Finally, this new study raises questions regarding optimal approaches for implementing pathway-directed treatments for LCH. In addition to vemurafenib, the FDA-approved MEK inhibitor trametinib7 is a rational therapeutic strategy for patients with LCH with mutations that aberrantly activate Raf/MEK/ERK signaling (see figure). However, because the long-term risks and benefits of these agents are unknown and other effective treatments exist for many patients with LCH, the optimal indications for administering a tyrosine kinase inhibitor—particularly to children—is an open question. A clinical trial of trametinib that is expected to open later this year in juvenile myelomonocytic leukemia, an aggressive myeloproliferative neoplasm of young children characterized by hyperactive Ras/Raf/MEK/ ERK signaling,9 will generate safety data that might be relevant for pediatric patients with LCH. A recent report demonstrating rapid and dramatic responses of 3 adults with ErdheimChester disease or LCH with BRAFV600E mutations to vemurafenib underscores the clinical potential of targeted therapies for aggressive or refractory disease.10 Whereas patients with melanoma and other advanced cancers with oncogenic BRAF mutations almost invariably relapse after tyrosine kinase inhibitor therapy,2,3,7 durable responses seem more likely in LCH and other neoplasms characterized by extensive differentiation and limited clonal heterogeneity. Treating carefully selected patients and monitoring their clinical and molecular responses are required to test this hypothesis. Conflict-of-interest disclosure: The authors declare no competing financial interests. n

3. Flaherty KT, Puzanov I, Kim KB, et al. Inhibition of mutated, activated BRAF in metastatic melanoma. N Engl J Med. 2010;363(9):809-819. 4. Badalian-Very G, Vergilio JA, Fleming M, Rollins BJ. Pathogenesis of Langerhans cell histiocytosis. Annu Rev Pathol. 2013;8:1-20. 5. Badalian-Very G, Vergilio JA, Degar BA, et al. Recurrent BRAF mutations in Langerhans cell histiocytosis. Blood. 2010;116(11):1919-1923.

BRAF-mutated melanoma. N Engl J Med. 2012;367(2): 107-114. 8. Imielinski M, Greulich H, Kaplan B, et al. Oncogenic and sorafenib-sensitive ARAF mutations in lung adenocarcinoma. J Clin Invest. 2014;124(4):1582-1586. 9. Loh ML. Recent advances in the pathogenesis and treatment of juvenile myelomonocytic leukaemia. Br J Haematol. 2011;152(6):677-687.

6. Berres ML, Lim KP, Peters T, et al. BRAF-V600E expression in precursor versus differentiated dendritic cells defines clinically distinct LCH risk groups. J Exp Med. 2014;211(4):669-683.

10. Haroche J, Cohen-Aubart F, Emile JF, et al. Dramatic efficacy of vemurafenib in both multisystemic and refractory Erdheim-Chester disease and Langerhans cell histiocytosis harboring the BRAF V600E mutation. Blood. 2013;121(9):1495-1500.

7. Flaherty KT, Robert C, Hersey P, et al; METRIC Study Group. Improved survival with MEK inhibition in

© 2014 by The American Society of Hematology

l l l PLATELETS & THROMBOPOIESIS

Comment on Kasirer-Friede et al, page 3156

Platelet aIIbb3 activation: filling in the pieces ----------------------------------------------------------------------------------------------------Joel S. Bennett1

1

UNIVERSITY OF PENNSYLVANIA

In this issue of Blood, Kasirer-Friede and colleagues show that the adhesion and degranulation promoter protein (ADAP) promotes aIIbb3 activation by presenting the cytoplasmic proteins talin and kindlin to the b3 cytoplasmic tail.1

T

he integrin aIIbb3 on circulating platelets is constrained in an inactive conformation by intramolecular interactions involving the

stalk, transmembrane, and cytoplasmic domains of its aIIb and b3 subunits to prevent spontaneous platelet aggregation in the

REFERENCES 1. Nelson DS, Quispel W, Badalian-Very G, et al. Somatic activating ARAF mutations in Langerhand cell histiocytosis. Blood. 2014;123(20):3152-3155. 2. Bollag G, Hirth P, Tsai J, et al. Clinical efficacy of a RAF inhibitor needs broad target blockade in BRAFmutant melanoma. Nature. 2010;467(7315):596-599.

BLOOD, 15 MAY 2014 x VOLUME 123, NUMBER 20

Talin and kindlin bind to ADAP in resting platelets and are transferred to the b3 cytoplasmic tail after platelet stimulation. Whether intermediary proteins are involved in the associations of ADAP with the platelet membrane and with talin and kindlin is not clear. L, aIIbb3 ligand; GFFKR, conserved membrane proximal region of the aIIb cytoplasmic tail that helps to maintain inactive aIIbb3. Adapted with permission from Figure 8 in O’Toole et al.11 © 1994 Rockefeller University Press. Originally published in Journal of Cell Biology. 124:1047-1059. doi:10.1083jcb.124.6.1047.

3065

From www.bloodjournal.org by guest on September 11, 2016. For personal use only.

circulation.2 However, at sites of vascular trauma, these interactions are disrupted nearly instantaneously, shifting aIIbb3 to its active ligand binding conformation and enabling it to mediate the formation of hemostatic plugs. In an elegant series of experiments, the Shattil and Ginsberg laboratories have shown that binding of the FERM (4.1/ezrin/radixin/moesin) domain of the cytoskeletal protein talin-1 to the b3 cytoplasmic tail is the event that disrupts the aIIbb3 constraints.3 Further, they have proposed that a complex containing the active form of the low-molecular-weight GTP-binding protein Rap-1 and RIAM (Rap1-GTP interacting adapter molecule) delivers talin-1 to b3.4 Thus the complex connects platelet agonist–stimulated Rap-1 activation to aIIbb3 activation. Importantly, the complex also appears to expose the b3-binding site on the talin-1 FERM domain, which is otherwise masked in the absence of talin-1 proteolysis or phosphorylation. These experiments have suggested that talin-1 binding to b3 alone is sufficient, at least in vitro, to cause aIIbb3 activation.5 Thus the observation that mutations in the gene for kindlin-3 in patients with LAD-III (leukocyte adhesion deficiency-III) produce a platelet phenotype resembling Glanzmann thrombasthenia was puzzling.6,7 Talin-1 and kindlin-3 both bind to the b3 cytoplasmic tail, but at separate sites, and in contrast to talin-1, kindlin-3 binding alone does not cause aIIbb3 activation. Thus it is conceivable that kindlin-3 binding “primes” the b3 cytoplasmic tail for talin-1 binding, although this has yet to be shown convincingly. Alternatively, the Ginsberg/ Shattil laboratories have proposed that kindlins may act to increase the affinity of integrins for multivalent ligands like fibrinogen by promoting the clustering of talin-activated integrins.8 In an earlier publication in Blood, KasirerFriede et al reported that agonist-stimulated aIIbb3 activation was decreased in mouse platelets lacking the adapter protein ADAP.9 This implied that ADAP is a component of the signal transduction pathways that activate aIIbb3. ADAP is a 120 or 130 kDa alternatively-spliced adapter protein expressed in hematopoietic cells whose function has been studied extensively in T lymphocytes. In T cells, ADAP is present in a membrane-

3066

associated complex containing SKAP1 (Src kinase-associated phosphoprotein 1, SKAP-55) and RIAM.10 After T-cell receptor activation, activated Rap1 binds to RIAM in the complex and in turn causes activation of b2- and b1-containing integrins. The role of ADAP in the complex appears to be to target SKAP1-RIAM to the membrane and to stabilize the otherwise unstable SKAP1 protein. Platelets do not contain a complex analogous to ADAP-SKAP1-RIAM. However, because ADAP does promote agonist-stimulated aIIbb3 activation, KasirerFriede et al hypothesized that ADAP acts by binding to either talin-1 or kindlin-3, or to both. In this paper, they show that indeed ADAP binds to talin-1 and to kindlins via interactions that do not involve either the SKAP1-homolog SKAP2 present in platelets or RIAM. Talin and kindlin binding to ADAP was detected in unstimulated platelets but decreased after platelet stimulation, suggesting that platelet stimulation induced the transfer of talin and kindlin from ADAP to the b3 tail. By microscopy, they found that ADAP co-localized with talin and kindlin-3 in platelets spread on fibrinogen and with aIIbb3, talin, and kindlin in SFLLRN-stimulated Chinese hamster ovary (CHO) cells. A puzzling feature of kindlin biology has been that although kindlin-2 overexpression in CHO cells supports aIIbb3 activation, kindlin-3 overexpression does not. However, the authors found that co-expressing ADAP with kindlin-3 now allowed kindlin-3 to activate aIIbb3, suggesting that the culprit was the absence of ADAP. Finally, they compared aIIbb3 function in ADAP1/1 and ADAP2/2 mice and found that ADAP deletion impaired agonist-stimulated fibrinogen binding, reduced agonist-stimulated aIIbb3 “avidity modulation,” and reduced the fraction of aIIbb3-bound fibrinogen that becomes irreversibly bound over time. The novel results reported by Kasirer-Friede et al imply that ADAP acts as a scaffold, bringing talin and kindlin to the platelet membrane and presenting them to b3 to promote aIIbb3 activation (see figure). Thus they have supplied yet another piece to

the puzzle of aIIbb3 activation and a new direction for future research. In addition, their results raise a number of interesting questions. What is the relative contribution of the ADAP- and RIAM-associated pathways to aIIbb3 activation? How do agonists stimulate the ADAP-associated pathway? How does ADAP associate with the platelet membrane, and are intermediary proteins involved? Finally, do talin and kindlin bind directly to ADAP or are as yet unidentified proteins analogous to SKAP1 and RIAM required? This platelet biologist eagerly looks forward to the answers to these questions. Conflict-of-interest disclosure: The author declares no competing financial interests. n REFERENCES 1. Kasirer-Friede A, Kang J, Kahner B, Ye F, Ginsberg MH, Shattil SJ. ADAP interactions with talin and kindlin promote platelet integrin aIIbb3 activation and stable fibrinogen binding. Blood. 2014;123(20):3156-3165. 2. Bennett JS. Structure and function of the platelet integrin alphaIIbbeta3. J Clin Invest. 2005;115(12): 3363-3369. 3. Han J, Lim CJ, Watanabe N, et al. Reconstructing and deconstructing agonist-induced activation of integrin alphaIIbbeta3. Curr Biol. 2006;16(18):1796-1806. 4. Lee HS, Lim CJ, Puzon-McLaughlin W, Shattil SJ, Ginsberg MH. RIAM activates integrins by linking talin to ras GTPase membrane-targeting sequences. J Biol Chem. 2009;284(8):5119-5127. 5. Ye F, Hu G, Taylor D, et al. Recreation of the terminal events in physiological integrin activation. J Cell Biol. 2010; 188(1):157-173. 6. Moser M, Legate KR, Zent R, F¨assler R. The tail of integrins, talin, and kindlins. Science. 2009;324(5929): 895-899. 7. Malinin NL, Zhang L, Choi J, et al. A point mutation in KINDLIN3 ablates activation of three integrin subfamilies in humans. Nat Med. 2009;15(3): 313-318. 8. Ye F, Petrich BG, Anekal P, et al. The mechanism of kindlin-mediated activation of integrin aIIbb3. Curr Biol. 2013;23(22):2288-2295. 9. Kasirer-Friede A, Moran B, Nagrampa-Orje J, et al. ADAP is required for normal alphaIIbbeta3 activation by VWF/GP Ib-IX-V and other agonists. Blood. 2007;109(3): 1018-1025. 10. M´enasch´e G, Kliche S, Chen EJ, Stradal TE, Schraven B, Koretzky G. RIAM links the ADAP/ SKAP-55 signaling module to Rap1, facilitating T-cellreceptor-mediated integrin activation. Mol Cell Biol. 2007; 27(11):4070-4081. 11. O’Toole TE, Katagiri Y, Faull RJ, et al. Integrin cytoplasmic domains mediate inside-out signal transduction. J Cell Biol. 1994;124(6):1047-1059. © 2014 by The American Society of Hematology

BLOOD, 15 MAY 2014 x VOLUME 123, NUMBER 20

From www.bloodjournal.org by guest on September 11, 2016. For personal use only.

2014 123: 3065-3067 doi:10.1182/blood-2014-02-557355

Platelet αIIbβ3 activation: filling in the pieces Joel S. Bennett

Updated information and services can be found at: http://www.bloodjournal.org/content/123/20/3065.full.html Articles on similar topics can be found in the following Blood collections Free Research Articles (4041 articles) Information about reproducing this article in parts or in its entirety may be found online at: http://www.bloodjournal.org/site/misc/rights.xhtml#repub_requests Information about ordering reprints may be found online at: http://www.bloodjournal.org/site/misc/rights.xhtml#reprints Information about subscriptions and ASH membership may be found online at: http://www.bloodjournal.org/site/subscriptions/index.xhtml

Blood (print ISSN 0006-4971, online ISSN 1528-0020), is published weekly by the American Society of Hematology, 2021 L St, NW, Suite 900, Washington DC 20036. Copyright 2011 by The American Society of Hematology; all rights reserved.

Platelet αIIbβ3 activation: filling in the pieces.

In this issue of Blood, Kasirer-Friede and colleagues show that the adhesion and degranulation promoter protein (ADAP) promotes αIIbβ3 activation by p...
576KB Sizes 3 Downloads 3 Views