CXCL4是人體血小板中含量最多的趨化因子，其與人體中許多生物過程有關，包括發炎反應、趨化性、動脈粥狀硬化、血小板低下、HIV-1感染的抑制和血管新生。CXCL4抗血管新生的活性藉由不同的機制進行調節，包括活化CXCR3受體。大部分的趨化因子具有不同的聚合形式，而不同形態的多聚體能夠控制其下游的生物活性。CXCL4在生理條件下為穩定的四聚體形式。但現在仍無法確定何種形態的CXCL4具有活化CXCR3受體的能力並且CXCL4是如何藉由不同的receptor去執行不同的生理功能。首先，我們發現pH和鹽類的電荷作用力會影響到CXCL4的四聚體形式。再者胺基酸序列中Glu-28對於此四聚體的形成相當重要，二聚體的形成主要是由於第一個β摺疊跟C端的α螺旋。我們也發現CXCL4形成單體時會活化並結合CXCR3A，且受體N端Tyrosine sulfation會影響其與趨化因子的結合能力。最後我們利用CXCL4的變異型，CXCL4L1，針對我們的研究做更進一步的了解。CXCL4L1與CXCL4的差異主要在C端的α螺旋上有三個胺基酸的不同，此三個胺基酸的不同造成CXCL4L1會活化CXCR3A，而native CXCL4則不會。在多聚體結構特性上，CXCL4L1比CXCL4有較高的傾向從多聚體解離成單體，此現象可能是造成CXCL4L1和CXCL4單體會活化CXCR3A的原因。所以，對於此趨化因子家族而言，單體的解離能力似乎是影響CXCR3A結合與活化的重要因子。我們的實驗清楚地說明趨化因子可以藉由改變其聚合態的方式來轉變其生物活性。由於CXCR3受體在人體細胞分佈廣泛，此研究影響了發炎反應、血管新生、癌症發育與腫瘤轉移等生理學上與病理學上許多重要的功能。

CXCL4 is the most abundant chemokine secreted from human platelet and it is critically involved in several biological processes that drive inflammation, chemotaxis, atherosclerosis, heparin-induced thrombocytopenia, inhibition of HIV-1 infection and angiogenesis. CXCL4 has anti-angiogenic properties thought to be mediated by different mechanisms, including CXCR3 receptor activation. Chemokines have distinct oligomerization states that are correlated with their biological functions. CXCL4 exists as a stable tetramer in physiological conditions. It is unclear whether the oligomerization state impacts CXCL4-receptor interactions and how CXCL4 executes different functions in correlation with the receptor. We noticed that the CXCL4 tetramer was sensitive to pH and salt concentration. Furthermore, it was determined that residue Glu-28 was important for tetramer formation, and the first β-strand and the C-terminal helix were critical for dimerization. The CXCL4 monomer acts as the active unit for activating CXCR3A, and N-terminal tyrosine sulfations of the receptor are involved in binding. Noticeably, CXCL4L1, a CXCL4 variant with three-residue difference in the C-terminal helix, could activate CXCR3A. CXCL4L1 but not native CXCL4 showed a higher tendency to directly dissociate into monomers. This indicates that monomeric CXCL4 behaves like CXCL4L1. Thus, in this chemokine family, being in the monomeric state seems critical for CXCR3A binding and activation. Our study clearly indicated chemokines can convert their biological functions by interfering with their oligomerization state. Due to the wide cellular distribution of CXCR3 receptor, this study is relevant in many physiological and pathological situations, such as immunity, angiogenesis, tumor development or metastatic spread.

1. Balkwill, F. (2004) Cancer and the chemokine network. Nature reviews. Cancer 4, 540-550
2. Raman, D., Sobolik-Delmaire, T., and Richmond, A. (2011) Chemokines in health and disease. Experimental cell research 317, 575-589
3. Keeley, E. C., Mehrad, B., and Strieter, R. M. (2011) Chemokines as mediators of tumor angiogenesis and neovascularization. Experimental cell research 317, 685-690
4. Mukaida, N., and Baba, T. (2012) Chemokines in tumor development and progression. Experimental cell research 318, 95-102
5. Bikfalvi, A. (2012) Angiogenesis and invasion in cancer. Handbook of clinical neurology 104, 35-43
6. Kiefer, F., and Siekmann, A. F. (2011) The role of chemokines and their receptors in angiogenesis. Cellular and molecular life sciences : CMLS 68, 2811-2830
7. Bikfalvi, A., Moenner, M., Javerzat, S., North, S., and Hagedorn, M. (2011) Inhibition of angiogenesis and the angiogenesis/invasion shift. Biochemical Society transactions 39, 1560-1564
8. Bikfalvi, A. (2009) [Tumoral angiogenesis: models, targets and inhibition]. Journal de la Societe de biologie 203, 167-170
9. Vandercappellen, J., Van Damme, J., and Struyf, S. (2011) The role of the CXC chemokines platelet factor-4 (CXCL4/PF-4) and its variant (CXCL4L1/PF-4var) in inflammation, angiogenesis and cancer. Cytokine & growth factor reviews 22, 1-18
10. Nomiyama, H., Osada, N., and Yoshie, O. (2010) The evolution of mammalian chemokine genes. Cytokine & growth factor reviews 21, 253-262
11. Frederick, M. J., and Clayman, G. L. (2001) Chemokines in cancer. Expert reviews in molecular medicine 3, 1-18
12. Bacon, K., Baggiolini, M., Broxmeyer, H., Horuk, R., Lindley, I., Mantovani, A., Maysushima, K., Murphy, P., Nomiyama, H., Oppenheim, J., Rot, A., Schall, T., Tsang, M., Thorpe, R., Van Damme, J., Wadhwa, M., Yoshie, O., Zlotnik, A., Zoon, K., and Nomenclature, I. W. S. o. C. (2002) Chemokine/chemokine receptor nomenclature. Journal of interferon & cytokine research : the official journal of the International Society for Interferon and Cytokine Research 22, 1067-1068
13. Scholten, D. J., Canals, M., Maussang, D., Roumen, L., Smit, M. J., Wijtmans, M., de Graaf, C., Vischer, H. F., and Leurs, R. (2012) Pharmacological modulation of chemokine receptor function. British journal of pharmacology 165, 1617-1643
14. Balkwill, F. R. (2012) The chemokine system and cancer. The Journal of pathology 226, 148-157
15. Zlotnik, A., and Yoshie, O. (2012) The chemokine superfamily revisited. Immunity 36, 705-716
16. Wang, X., Sharp, J. S., Handel, T. M., and Prestegard, J. H. (2013) Chemokine oligomerization in cell signaling and migration. Progress in molecular biology and translational science 117, 531-578
17. Jansma, A., Handel, T. M., and Hamel, D. J. (2009) Chapter 2. Homo- and hetero-oligomerization of chemokines. Methods in enzymology 461, 31-50
18. Carlson, J., Baxter, S. A., Dreau, D., and Nesmelova, I. V. (2013) The heterodimerization of platelet-derived chemokines. Biochimica et biophysica acta 1834, 158-168
19. Allen, S. J., Crown, S. E., and Handel, T. M. (2007) Chemokine: receptor structure, interactions, and antagonism. Annual review of immunology 25, 787-820
20. Lortat-Jacob, H., Grosdidier, A., and Imberty, A. (2002) Structural diversity of heparan sulfate binding domains in chemokines. Proceedings of the National Academy of Sciences of the United States of America 99, 1229-1234
21. Proudfoot, A. E. (2006) The biological relevance of chemokine-proteoglycan interactions. Biochemical Society transactions 34, 422-426
22. Rot, A. (1993) Neutrophil attractant/activation protein-1 (interleukin-8) induces in vitro neutrophil migration by haptotactic mechanism. European journal of immunology 23, 303-306
23. Haessler, U., Pisano, M., Wu, M., and Swartz, M. A. (2011) Dendritic cell chemotaxis in 3D under defined chemokine gradients reveals differential response to ligands CCL21 and CCL19. Proceedings of the National Academy of Sciences of the United States of America 108, 5614-5619
24. Weber, M., Hauschild, R., Schwarz, J., Moussion, C., de Vries, I., Legler, D. F., Luther, S. A., Bollenbach, T., and Sixt, M. (2013) Interstitial dendritic cell guidance by haptotactic chemokine gradients. Science 339, 328-332
25. Proudfoot, A. E., Handel, T. M., Johnson, Z., Lau, E. K., LiWang, P., Clark-Lewis, I., Borlat, F., Wells, T. N., and Kosco-Vilbois, M. H. (2003) Glycosaminoglycan binding and oligomerization are essential for the in vivo activity of certain chemokines. Proceedings of the National Academy of Sciences of the United States of America 100, 1885-1890
26. Campanella, G. S., Grimm, J., Manice, L. A., Colvin, R. A., Medoff, B. D., Wojtkiewicz, G. R., Weissleder, R., and Luster, A. D. (2006) Oligomerization of CXCL10 is necessary for endothelial cell presentation and in vivo activity. Journal of immunology 177, 6991-6998
27. Salanga, C. L., and Handel, T. M. (2011) Chemokine oligomerization and interactions with receptors and glycosaminoglycans: the role of structural dynamics in function. Experimental cell research 317, 590-601
28. Laguri, C., Sadir, R., Rueda, P., Baleux, F., Gans, P., Arenzana-Seisdedos, F., and Lortat-Jacob, H. (2007) The novel CXCL12gamma isoform encodes an unstructured cationic domain which regulates bioactivity and interaction with both glycosaminoglycans and CXCR4. PloS one 2, e1110
29. Liehn, E. A., Piccinini, A. M., Koenen, R. R., Soehnlein, O., Adage, T., Fatu, R., Curaj, A., Popescu, A., Zernecke, A., Kungl, A. J., and Weber, C. (2010) A new monocyte chemotactic protein-1/chemokine CC motif ligand-2 competitor limiting neointima formation and myocardial ischemia/reperfusion injury in mice. Journal of the American College of Cardiology 56, 1847-1857
30. Palczewski, K., Kumasaka, T., Hori, T., Behnke, C. A., Motoshima, H., Fox, B. A., Le Trong, I., Teller, D. C., Okada, T., Stenkamp, R. E., Yamamoto, M., and Miyano, M. (2000) Crystal structure of rhodopsin: A G protein-coupled receptor. Science 289, 739-745
31. Stenkamp, R. E., Teller, D. C., and Palczewski, K. (2002) Crystal structure of rhodopsin: a G-protein-coupled receptor. Chembiochem : a European journal of chemical biology 3, 963-967
32. Crasto, C. J. (2010) Hydrophobicity profiles in G protein-coupled receptor transmembrane helical domains. Journal of receptor, ligand and channel research 2010, 123-133
33. Murdoch, C., and Finn, A. (2000) Chemokine receptors and their role in inflammation and infectious diseases. Blood 95, 3032-3043
34. Hoogewerf, A. J., Kuschert, G. S., Proudfoot, A. E., Borlat, F., Clark-Lewis, I., Power, C. A., and Wells, T. N. (1997) Glycosaminoglycans mediate cell surface oligomerization of chemokines. Biochemistry 36, 13570-13578
35. Lau, E. K., Allen, S., Hsu, A. R., and Handel, T. M. (2004) Chemokine-receptor interactions: GPCRs, glycosaminoglycans and viral chemokine binding proteins. Advances in protein chemistry 68, 351-391
36. Lau, E. K., Paavola, C. D., Johnson, Z., Gaudry, J. P., Geretti, E., Borlat, F., Kungl, A. J., Proudfoot, A. E., and Handel, T. M. (2004) Identification of the glycosaminoglycan binding site of the CC chemokine, MCP-1: implications for structure and function in vivo. The Journal of biological chemistry 279, 22294-22305
37. Kuschert, G. S., Coulin, F., Power, C. A., Proudfoot, A. E., Hubbard, R. E., Hoogewerf, A. J., and Wells, T. N. (1999) Glycosaminoglycans interact selectively with chemokines and modulate receptor binding and cellular responses. Biochemistry 38, 12959-12968
38. Paavola, C. D., Hemmerich, S., Grunberger, D., Polsky, I., Bloom, A., Freedman, R., Mulkins, M., Bhakta, S., McCarley, D., Wiesent, L., Wong, B., Jarnagin, K., and Handel, T. M. (1998) Monomeric monocyte chemoattractant protein-1 (MCP-1) binds and activates the MCP-1 receptor CCR2B. The Journal of biological chemistry 273, 33157-33165
39. Rajarathnam, K., Sykes, B. D., Kay, C. M., Dewald, B., Geiser, T., Baggiolini, M., and Clark-Lewis, I. (1994) Neutrophil activation by monomeric interleukin-8. Science 264, 90-92
40. Qin, L., Kufareva, I., Holden, L. G., Wang, C., Zheng, Y., Zhao, C., Fenalti, G., Wu, H., Han, G. W., Cherezov, V., Abagyan, R., Stevens, R. C., and Handel, T. M. (2015) Structural biology. Crystal structure of the chemokine receptor CXCR4 in complex with a viral chemokine. Science 347, 1117-1122
41. Burg, J. S., Ingram, J. R., Venkatakrishnan, A. J., Jude, K. M., Dukkipati, A., Feinberg, E. N., Angelini, A., Waghray, D., Dror, R. O., Ploegh, H. L., and Garcia, K. C. (2015) Structural biology. Structural basis for chemokine recognition and activation of a viral G protein-coupled receptor. Science 347, 1113-1117
42. Wells, T. N., Power, C. A., Lusti-Narasimhan, M., Hoogewerf, A. J., Cooke, R. M., Chung, C. W., Peitsch, M. C., and Proudfoot, A. E. (1996) Selectivity and antagonism of chemokine receptors. Journal of leukocyte biology 59, 53-60
43. Simpson, L. S., Zhu, J. Z., Widlanski, T. S., and Stone, M. J. (2009) Regulation of chemokine recognition by site-specific tyrosine sulfation of receptor peptides. Chemistry & biology 16, 153-161
44. Ludeman, J. P., and Stone, M. J. (2014) The structural role of receptor tyrosine sulfation in chemokine recognition. British journal of pharmacology 171, 1167-1179
45. Zhu, J. Z., Millard, C. J., Ludeman, J. P., Simpson, L. S., Clayton, D. J., Payne, R. J., Widlanski, T. S., and Stone, M. J. (2011) Tyrosine sulfation influences the chemokine binding selectivity of peptides derived from chemokine receptor CCR3. Biochemistry 50, 1524-1534
46. Colvin, R. A., Campanella, G. S., Manice, L. A., and Luster, A. D. (2006) CXCR3 requires tyrosine sulfation for ligand binding and a second extracellular loop arginine residue for ligand-induced chemotaxis. Molecular and cellular biology 26, 5838-5849
47. Tan, J. H., Ludeman, J. P., Wedderburn, J., Canals, M., Hall, P., Butler, S. J., Taleski, D., Christopoulos, A., Hickey, M. J., Payne, R. J., and Stone, M. J. (2013) Tyrosine sulfation of chemokine receptor CCR2 enhances interactions with both monomeric and dimeric forms of the chemokine monocyte chemoattractant protein-1 (MCP-1). The Journal of biological chemistry 288, 10024-10034
48. Millard, C. J., Ludeman, J. P., Canals, M., Bridgford, J. L., Hinds, M. G., Clayton, D. J., Christopoulos, A., Payne, R. J., and Stone, M. J. (2014) Structural basis of receptor sulfotyrosine recognition by a CC chemokine: the N-terminal region of CCR3 bound to CCL11/eotaxin-1. Structure 22, 1571-1581
49. Karshovska, E., Weber, C., and von Hundelshausen, P. (2013) Platelet chemokines in health and disease. Thrombosis and haemostasis 110, 894-902
50. Zhang, X., Chen, L., Bancroft, D. P., Lai, C. K., and Maione, T. E. (1994) Crystal structure of recombinant human platelet factor 4. Biochemistry 33, 8361-8366
51. Aidoudi, S., and Bikfalvi, A. (2010) Interaction of PF4 (CXCL4) with the vasculature: a role in atherosclerosis and angiogenesis. Thrombosis and haemostasis 104, 941-948
52. Maione, T. E., Gray, G. S., Petro, J., Hunt, A. J., Donner, A. L., Bauer, S. I., Carson, H. F., and Sharpe, R. J. (1990) Inhibition of angiogenesis by recombinant human platelet factor-4 and related peptides. Science 247, 77-79
53. Bikfalvi, A. (2004) Platelet factor 4: an inhibitor of angiogenesis. Seminars in thrombosis and hemostasis 30, 379-385
54. Aidoudi, S., Bujakowska, K., Kieffer, N., and Bikfalvi, A. (2008) The CXC-chemokine CXCL4 interacts with integrins implicated in angiogenesis. PloS one 3, e2657
55. Lasagni, L., Francalanci, M., Annunziato, F., Lazzeri, E., Giannini, S., Cosmi, L., Sagrinati, C., Mazzinghi, B., Orlando, C., Maggi, E., Marra, F., Romagnani, S., Serio, M., and Romagnani, P. (2003) An alternatively spliced variant of CXCR3 mediates the inhibition of endothelial cell growth induced by IP-10, Mig, and I-TAC, and acts as functional receptor for platelet factor 4. The Journal of experimental medicine 197, 1537-1549
56. Bikfalvi, A. (2004) Recent developments in the inhibition of angiogenesis: examples from studies on platelet factor-4 and the VEGF/VEGFR system. Biochemical pharmacology 68, 1017-1021
57. Rostene, W., Guyon, A., Kular, L., Godefroy, D., Barbieri, F., Bajetto, A., Banisadr, G., Callewaere, C., Conductier, G., Rovere, C., Melik-Parsadaniantz, S., and Florio, T. (2011) Chemokines and chemokine receptors: new actors in neuroendocrine regulations. Frontiers in neuroendocrinology 32, 10-24
58. Koenen, R. R., and Weber, C. (2011) Chemokines: established and novel targets in atherosclerosis. EMBO molecular medicine 3, 713-725
59. Gleissner, C. A., von Hundelshausen, P., and Ley, K. (2008) Platelet chemokines in vascular disease. Arteriosclerosis, thrombosis, and vascular biology 28, 1920-1927
60. Balestrieri, M. L., Balestrieri, A., Mancini, F. P., and Napoli, C. (2008) Understanding the immunoangiostatic CXC chemokine network. Cardiovascular research 78, 250-256
61. Koelink, P. J., Overbeek, S. A., Braber, S., de Kruijf, P., Folkerts, G., Smit, M. J., and Kraneveld, A. D. (2012) Targeting chemokine receptors in chronic inflammatory diseases: an extensive review. Pharmacology & therapeutics 133, 1-18
62. Schwartzkopff, F., Grimm, T. A., Lankford, C. S., Fields, K., Wang, J., Brandt, E., and Clouse, K. A. (2009) Platelet factor 4 (CXCL4) facilitates human macrophage infection with HIV-1 and potentiates virus replication. Innate immunity 15, 368-379
63. Auerbach, D. J., Lin, Y., Miao, H., Cimbro, R., Difiore, M. J., Gianolini, M. E., Furci, L., Biswas, P., Fauci, A. S., and Lusso, P. (2012) Identification of the platelet-derived chemokine CXCL4/PF-4 as a broad-spectrum HIV-1 inhibitor. Proceedings of the National Academy of Sciences of the United States of America 109, 9569-9574
64. Hagedorn, M., Zilberberg, L., Lozano, R. M., Cuevas, P., Canron, X., Redondo-Horcajo, M., Gimenez-Gallego, G., and Bikfalvi, A. (2001) A short peptide domain of platelet factor 4 blocks angiogenic key events induced by FGF-2. FASEB journal : official publication of the Federation of American Societies for Experimental Biology 15, 550-552
65. Petrai, I., Rombouts, K., Lasagni, L., Annunziato, F., Cosmi, L., Romanelli, R. G., Sagrinati, C., Mazzinghi, B., Pinzani, M., Romagnani, S., Romagnani, P., and Marra, F. (2008) Activation of p38(MAPK) mediates the angiostatic effect of the chemokine receptor CXCR3-B. The international journal of biochemistry & cell biology 40, 1764-1774
66. Van Raemdonck, K., Van den Steen, P. E., Liekens, S., Van Damme, J., and Struyf, S. (2015) CXCR3 ligands in disease and therapy. Cytokine & growth factor reviews 26, 311-327
67. de Jong, E. K., de Haas, A. H., Brouwer, N., van Weering, H. R., Hensens, M., Bechmann, I., Pratley, P., Wesseling, E., Boddeke, H. W., and Biber, K. (2008) Expression of CXCL4 in microglia in vitro and in vivo and its possible signaling through CXCR3. Journal of neurochemistry 105, 1726-1736
68. Perollet, C., Han, Z. C., Savona, C., Caen, J. P., and Bikfalvi, A. (1998) Platelet factor 4 modulates fibroblast growth factor 2 (FGF-2) activity and inhibits FGF-2 dimerization. Blood 91, 3289-3299
69. Billottet, C., Quemener, C., and Bikfalvi, A. (2013) CXCR3, a double-edged sword in tumor progression and angiogenesis. Biochimica et biophysica acta 1836, 287-295
70. Quemener, C., Baud, J., Boye, K., Dubrac, A., Billottet, C., Soulet, F., Darlot, F., Dumartin, L., Sire, M., Grepin, R., Daubon, T., Rayne, F., Wodrich, H., Couvelard, A., Pineau, R., Schilling, M., Castronovo, V., Sue, S. C., Clarke, K., Lomri, A., Khatib, A. M., Hagedorn, M., Prats, H., and Bikfalvi, A. (2016) Dual Roles for CXCL4 Chemokines and CXCR3 in Angiogenesis and Invasion of Pancreatic Cancer. Cancer research
71. Kuo, J. H., Chen, Y. P., Liu, J. S., Dubrac, A., Quemener, C., Prats, H., Bikfalvi, A., Wu, W. G., and Sue, S. C. (2013) Alternative C-terminal helix orientation alters chemokine function: structure of the anti-angiogenic chemokine, CXCL4L1. The Journal of biological chemistry 288, 13522-13533
72. Struyf, S., Burdick, M. D., Peeters, E., Van den Broeck, K., Dillen, C., Proost, P., Van Damme, J., and Strieter, R. M. (2007) Platelet factor-4 variant chemokine CXCL4L1 inhibits melanoma and lung carcinoma growth and metastasis by preventing angiogenesis. Cancer research 67, 5940-5948
73. Struyf, S., Burdick, M. D., Proost, P., Van Damme, J., and Strieter, R. M. (2004) Platelets release CXCL4L1, a nonallelic variant of the chemokine platelet factor-4/CXCL4 and potent inhibitor of angiogenesis. Circulation research 95, 855-857
74. Dubrac, A., Quemener, C., Lacazette, E., Lopez, F., Zanibellato, C., Wu, W. G., Bikfalvi, A., and Prats, H. (2010) Functional divergence between 2 chemokines is conferred by single amino acid change. Blood 116, 4703-4711
75. Mayo, K. H., Roongta, V., Ilyina, E., Milius, R., Barker, S., Quinlan, C., La Rosa, G., and Daly, T. J. (1995) NMR solution structure of the 32-kDa platelet factor 4 ELR-motif N-terminal chimera: a symmetric tetramer. Biochemistry 34, 11399-11409
76. Nesmelova, I. V., Sham, Y., Gao, J., and Mayo, K. H. (2008) CXC and CC chemokines form mixed heterodimers: association free energies from molecular dynamics simulations and experimental correlations. The Journal of biological chemistry 283, 24155-24166
77. Sharma, D., and Rajarathnam, K. (2000) 13C NMR chemical shifts can predict disulfide bond formation. Journal of biomolecular NMR 18, 165-171
78. Ravindran, A., Joseph, P. R., and Rajarathnam, K. (2009) Structural basis for differential binding of the interleukin-8 monomer and dimer to the CXCR1 N-domain: role of coupled interactions and dynamics. Biochemistry 48, 8795-8805
79. Ravindran, A., Sawant, K. V., Sarmiento, J., Navarro, J., and Rajarathnam, K. (2013) Chemokine CXCL1 dimer is a potent agonist for the CXCR2 receptor. The Journal of biological chemistry 288, 12244-12252
80. Joseph, P. R., Poluri, K. M., Gangavarapu, P., Rajagopalan, L., Raghuwanshi, S., Richardson, R. M., Garofalo, R. P., and Rajarathnam, K. (2013) Proline substitution of dimer interface beta-strand residues as a strategy for the design of functional monomeric proteins. Biophysical journal 105, 1491-1501
81. Quay, S. C., and Condie, C. C. (1983) Conformational studies of aqueous melittin: thermodynamic parameters of the monomer-tetramer self-association reaction. Biochemistry 22, 695-700
82. Dyer, D. P., Salanga, C. L., Volkman, B. F., Kawamura, T., and Handel, T. M. (2016) The dependence of chemokine-glycosaminoglycan interactions on chemokine oligomerization. Glycobiology 26, 312-326
83. Liang, W. G., Triandafillou, C. G., Huang, T. Y., Zulueta, M. M., Banerjee, S., Dinner, A. R., Hung, S. C., and Tang, W. J. (2016) Structural basis for oligomerization and glycosaminoglycan binding of CCL5 and CCL3. Proceedings of the National Academy of Sciences of the United States of America 113, 5000-5005
84. Farzan, M., Babcock, G. J., Vasilieva, N., Wright, P. L., Kiprilov, E., Mirzabekov, T., and Choe, H. (2002) The role of post-translational modifications of the CXCR4 amino terminus in stromal-derived factor 1 alpha association and HIV-1 entry. The Journal of biological chemistry 277, 29484-29489
85. Farzan, M., Mirzabekov, T., Kolchinsky, P., Wyatt, R., Cayabyab, M., Gerard, N. P., Gerard, C., Sodroski, J., and Choe, H. (1999) Tyrosine sulfation of the amino terminus of CCR5 facilitates HIV-1 entry. Cell 96, 667-676
86. Fong, A. M., Alam, S. M., Imai, T., Haribabu, B., and Patel, D. D. (2002) CX3CR1 tyrosine sulfation enhances fractalkine-induced cell adhesion. The Journal of biological chemistry 277, 19418-19423
87. Gutierrez, J., Kremer, L., Zaballos, A., Goya, I., Martinez, A. C., and Marquez, G. (2004) Analysis of post-translational CCR8 modifications and their influence on receptor activity. The Journal of biological chemistry 279, 14726-14733
88. Preobrazhensky, A. A., Dragan, S., Kawano, T., Gavrilin, M. A., Gulina, I. V., Chakravarty, L., and Kolattukudy, P. E. (2000) Monocyte chemotactic protein-1 receptor CCR2B is a glycoprotein that has tyrosine sulfation in a conserved extracellular N-terminal region. Journal of immunology 165, 5295-5303
89. Duma, L., Haussinger, D., Rogowski, M., Lusso, P., and Grzesiek, S. (2007) Recognition of RANTES by extracellular parts of the CCR5 receptor. Journal of molecular biology 365, 1063-1075
90. Veldkamp, C. T., Seibert, C., Peterson, F. C., De la Cruz, N. B., Haugner, J. C., 3rd, Basnet, H., Sakmar, T. P., and Volkman, B. F. (2008) Structural basis of CXCR4 sulfotyrosine recognition by the chemokine SDF-1/CXCL12. Science signaling 1, ra4
91. Skelton, N. J., Quan, C., Reilly, D., and Lowman, H. (1999) Structure of a CXC chemokine-receptor fragment in complex with interleukin-8. Structure 7, 157-168
92. Lu, B., Humbles, A., Bota, D., Gerard, C., Moser, B., Soler, D., Luster, A. D., and Gerard, N. P. (1999) Structure and function of the murine chemokine receptor CXCR3. European journal of immunology 29, 3804-3812
93. Wang, X., Li, X., Schmidt, D. B., Foley, J. J., Barone, F. C., Ames, R. S., and Sarau, H. M. (2000) Identification and molecular characterization of rat CXCR3: receptor expression and interferon-inducible protein-10 binding are increased in focal stroke. Molecular pharmacology 57, 1190-1198
94. Kufareva, I., Salanga, C. L., and Handel, T. M. (2015) Chemokine and chemokine receptor structure and interactions: implications for therapeutic strategies. Immunology and cell biology 93, 372-383
95. Romagnani, P., Annunziato, F., Lasagni, L., Lazzeri, E., Beltrame, C., Francalanci, M., Uguccioni, M., Galli, G., Cosmi, L., Maurenzig, L., Baggiolini, M., Maggi, E., Romagnani, S., and Serio, M. (2001) Cell cycle-dependent expression of CXC chemokine receptor 3 by endothelial cells mediates angiostatic activity. The Journal of clinical investigation 107, 53-63
96. Eishingdrelo, H., and Kongsamut, S. (2013) Minireview: Targeting GPCR Activated ERK Pathways for Drug Discovery. Current chemical genomics and translational medicine 7, 9-15
97. Struyf, S., Salogni, L., Burdick, M. D., Vandercappellen, J., Gouwy, M., Noppen, S., Proost, P., Opdenakker, G., Parmentier, M., Gerard, C., Sozzani, S., Strieter, R. M., and Van Damme, J. (2011) Angiostatic and chemotactic activities of the CXC chemokine CXCL4L1 (platelet factor-4 variant) are mediated by CXCR3. Blood 117, 480-488
98. Vandercappellen, J., Liekens, S., Bronckaers, A., Noppen, S., Ronsse, I., Dillen, C., Belleri, M., Mitola, S., Proost, P., Presta, M., Struyf, S., and Van Damme, J. (2010) The COOH-terminal peptide of platelet factor-4 variant (CXCL4L1/PF-4var47-70) strongly inhibits angiogenesis and suppresses B16 melanoma growth in vivo. Molecular cancer research : MCR 8, 322-334
99. Rajarathnam, K., Prado, G. N., Fernando, H., Clark-Lewis, I., and Navarro, J. (2006) Probing receptor binding activity of interleukin-8 dimer using a disulfide trap. Biochemistry 45, 7882-7888
100. Leong, S. R., Lowman, H. B., Liu, J., Shire, S., Deforge, L. E., Gillece-Castro, B. L., McDowell, R., and Hebert, C. A. (1997) IL-8 single-chain homodimers and heterodimers: interactions with chemokine receptors CXCR1, CXCR2, and DARC. Protein science : a publication of the Protein Society 6, 609-617
101. Jin, H., Shen, X., Baggett, B. R., Kong, X., and LiWang, P. J. (2007) The human CC chemokine MIP-1beta dimer is not competent to bind to the CCR5 receptor. The Journal of biological chemistry 282, 27976-27983
102. Tan, J. H., Canals, M., Ludeman, J. P., Wedderburn, J., Boston, C., Butler, S. J., Carrick, A. M., Parody, T. R., Taleski, D., Christopoulos, A., Payne, R. J., and Stone, M. J. (2012) Design and receptor interactions of obligate dimeric mutant of chemokine monocyte chemoattractant protein-1 (MCP-1). The Journal of biological chemistry 287, 14692-14702
103. Nasser, M. W., Raghuwanshi, S. K., Grant, D. J., Jala, V. R., Rajarathnam, K., and Richardson, R. M. (2009) Differential activation and regulation of CXCR1 and CXCR2 by CXCL8 monomer and dimer. Journal of immunology 183, 3425-3432
104. Fernando, H., Chin, C., Rosgen, J., and Rajarathnam, K. (2004) Dimer dissociation is essential for interleukin-8 (IL-8) binding to CXCR1 receptor. The Journal of biological chemistry 279, 36175-36178
105. Joseph, P. R., and Rajarathnam, K. (2015) Solution NMR characterization of WT CXCL8 monomer and dimer binding to CXCR1 N-terminal domain. Protein science : a publication of the Protein Society 24, 81-92
106. Drury, L. J., Ziarek, J. J., Gravel, S., Veldkamp, C. T., Takekoshi, T., Hwang, S. T., Heveker, N., Volkman, B. F., and Dwinell, M. B. (2011) Monomeric and dimeric CXCL12 inhibit metastasis through distinct CXCR4 interactions and signaling pathways. Proceedings of the National Academy of Sciences of the United States of America 108, 17655-17660
107. Delaglio, F., Grzesiek, S., Vuister, G. W., Zhu, G., Pfeifer, J., and Bax, A. (1995) NMRPipe: a multidimensional spectral processing system based on UNIX pipes. Journal of biomolecular NMR 6, 277-293
108. Schwarzinger, S., Kroon, G. J., Foss, T. R., Chung, J., Wright, P. E., and Dyson, H. J. (2001) Sequence-dependent correction of random coil NMR chemical shifts. Journal of the American Chemical Society 123, 2970-2978
109. Mizuguchi, T., Uchimura, H., Kataoka, H., Akaji, K., Kiso, Y., and Saito, K. (2012) Intact-cell-based surface plasmon resonance measurements for ligand affinity evaluation of a membrane receptor. Analytical biochemistry 420, 185-187
110. Alves, I. D., Delaroche, D., Mouillac, B., Salamon, Z., Tollin, G., Hruby, V. J., Lavielle, S., and Sagan, S. (2006) The two NK-1 binding sites correspond to distinct, independent, and non-interconvertible receptor conformational states as confirmed by plasmon-waveguide resonance spectroscopy. Biochemistry 45, 5309-5318
111. Salamon, Z., Fitch, J., Cai, M., Tumati, S., Navratilova, E., and Tollin, G. (2009) Plasmon-waveguide resonance studies of ligand binding to integral proteins in membrane fragments derived from bacterial and mammalian cells. Analytical biochemistry 387, 95-101
112. Harte, E., Maalouli, N., Shalabney, A., Texier, E., Berthelot, K., Lecomte, S., and Alves, I. D. (2014) Probing the kinetics of lipid membrane formation and the interaction of a nontoxic and a toxic amyloid with plasmon waveguide resonance. Chemical communications 50, 4168-4171