臺灣博碩士論文加值系統

中 文 摘 要
細胞的吸附作用調控著很多重要的生理功能, 包括胚胎的發育, 細胞的游移, 生長, 分化, 以及免疫細胞的活化, 血小板的凝集, 腫瘤細胞的轉移. Integrins為媒介細胞與細胞以及細胞和細胞外間質交互作用最主要的分子, 其雙向訊息傳導的功能整合了細胞外的訊息以及細胞內的生理狀態. Disintegrin家族為一群由蛇毒中分離出來的蛋白所組成, 其成員在結構上的特性為小型多胜鏈, 富含cysteine以及一段RGD或KGD序列. Disintegrin已被證實能阻斷fibrinogen與血小板上integrin的結合以抑制血小板的凝集. Rhodostomin(又稱為kistrin, 簡稱RHO)是從Agkistrodon rhodostoma(又稱Calloselasma rhodostoma, 馬來亞蝮蛇)蛇毒中所純化出來的disintegrin, 包含68個氨基酸, 12個cysteine, 6對雙硫鍵, 以及RGD motif在氨基酸位置49-51. 關於rhodostomin的生物功能已經有很多的研究報告, 尤其是將rhodostomin的RGD序列換為LDV所造成之rhodostomin-LDV嵌合體具有類似fibronectin之LDV片段的結合活性及專一性. 但是, rhodostomin分子骨架以及RGD鄰近序列對於rhodostomin活性的重要性仍然未明. 本實驗利用插入突變以及取代突變的方式, 探討鄰近序列與RGD相對位置的重要性以及分子骨架所扮演的角色. GST攫取試驗證實RGD的4個N端插入突變以及3個C端插入突變會降低與integrin aIIbb3的結合能力. 但是2個緊接在RGD後面的插入突變[GST-RHO(51+A)以及GST-RHO(52+A)]以及位於Cys57之前的插入突變[GST-RHO(56+A)]則幾乎完全喪失與aIIbb3結合的能力. 此外, 血小板凝集抑制試驗以及血小板吸附試驗的結果都與攫取試驗的結果一致. 但是Pro53的取代突變種只會喪失與母系CHO細胞的吸附能力, 對於表現avb3或是aIIbb3的CHO細胞吸附則是稍微減少. 為了進一步評估rhodostomin分子骨架的功能, 本實驗比較rhodostomin[GST-RHO(RGD)], 人類MDC-15之disintegrin-like區域(GST-D15)以及rhodostomin-D15嵌合體對於integrin aIIbb3的結合能力. GST攫取試驗發現GST-D15對於integrin aIIbb3的結合能力遠低於GST-RHO(RGD), 而與rhodostomin-D15的嵌合體相當. 但是rhodostomin-D15嵌合體可以讓表現aIIbb3的CHO細胞吸附而GST-D15不行. 此外, 帶有Pro取代突變的GST-D15突變種(GST-D15/CP, GST-D15/MP)也可以讓表現aIIbb3的CHO細胞吸附. 總結上述結果, 本實驗提出堅強的證據證實: (1) rhodostomin真正參與integrin結合作用的區域是Arg49-Gly50-Asp51-Met52-Pro53. 除了RGD motif之外, 位於RGD的C端第2個氨基酸Pro53對於integrin的結合也是非常重要; (2) 本實驗第一次比較蛇毒disintegrin與人類MDC在integrin交互作用上的活性差異. 人類MDC之disintegrin-like區域(GST-D15)的integrin結合能力遠低於蛇毒disintegrin[GST-RHO(RGD)], 而其原因為RGD motif的鄰近序列不同所導致; (3) rhodostomin的分子骨架非常適合與fibrinogen的receptor, 即integrin avb3和aIIbb3結合. 同時, 利用人類MDC-15之RGD及其鄰近序列共9個氨基酸所製造之rhodostomin-D15嵌合體, 表現出近似GST-D15之integrin結合活性. 因此, 本實驗證實rhodostomin的分子骨架可以呈現來自其它disintegrin相關分子的9個氨基酸片段.

Abstract
Cell adhesion plays critical roles in directing the embryonic development, cell migration, proliferation, differentiation, as well as activation of immunocytes, aggregation of platelet, and cancer metastasis. Integrins are the major receptors mediated cell-cell and cell-extracellular matrix interactions, the bi-directional signal transducing activity of integrin integrates the extracellular signals and intracellular events. Disintegrins represent a protein family of low molecular weight, cysteine-rich and RGD- or KGD-containing peptides isolated from snake venom. They have been proved to inhibit platelet aggregation by blocking the binding between fibrinogen and platelet integrins. Rhodostomin(also called kistrin, RHO in abbreviation, a member of disintegrin) is derived from snake venom of Agkistrodon rhodostoma(also called Calloselasma rhodostoma, Malayan pit viper), which contains 68 amino acids, including 12 cysteine residues, 6 disulfide bonds, and an RGD motif at positions of 49-51. Intense studies have been done to define the biological functions of rhodostomin. Specifically, the rhodostomin-LDV chimeras which contain LDV motif in the natural RGD site had been proved to exhibit potency and specificity similar to the LDV-containing fragment of fibronectin. However, the role of presenting scaffold and the influence of relative distance between flanking residues and RGD motif of rhodostomin are unclear. In this study, by utilizing insertion mutagenesis and substitution mutagenesis, the importance of distance between RGD and flanking residues and the specific function of presenting scaffold of rhodostomin were characterized. GST pull-down assays indicated the 4 alanine insertions at N-terminal flanking region and 3 alanine insertions at C-terminal flanking region of RGD motif were able to reduce integrin aIIbb3 binding activity to a limited extent. In contrast, 2 insertions[GST-RHO(51+A) and GST-RHO(52+A)] immediately next to RGD and 1 insertion[GST-RHO(56+A)] in front of the Cys57 caused almost complete loss of binding activity to aIIbb3. The results of platelet aggregation assay and platelet adhesion assay for the insertion mutants were consistent with results of pull-down assay. However, Pro53-substituted mutants abolished parental CHO cells adhesion but mild reduced avb3- or aIIbb3-expressing CHO cells adhesion. To further evaluate the presenting function of rhodostomin scaffold, the aIIbb3 binding activity of rhodostomin[GST-RHO(RGD)], disintegrin-like domain of MDC-15(GST-D15) and fragment-graft rhodostomin-D15 chimeras were compared. The results of pull-down assay showed the binding activity of GST-D15 is much weaker than GST-RHO(RGD) and similar to rhodostomin-D15 chimeras. However, rhodostomin-D15 chimeras can mediate aIIbb3-expressing CHO cells adhesion but GST-D15 cannot. Besides, Pro-substituted D15 mutants(GST-D15/CP, GST-D15/MP) also mediate the adhesion of aIIbb3-expressing CHO cells. In summary, this study provides strong evidences that (i) the exact region of rhodostomin involved in integrin-binding interaction is Arg49-Gly50-Asp51-Met52-Pro53. In addition to the RGD motif, the Pro53 at position next 2 to RGD is also important in integrin binding; (ii) the low aIIbb3 binding activity of the disintegrin-like domain of MDC-15 is due to the influence of the flanking sequence of RGD motif; and (iii) the rhodostomin scaffold is optimized for interacting with fibrinogen receptors, integrin aIIbb3 and avb3. In addition, the integrin-binding activity of rhodostomin-D15 chimera is similar to GST-D15. This result indicated the scaffold of rhodostomin is suitable for presentation of large fragment as 9 amino acids from disintegrin-related molecules.

封面
中文摘要
英文摘要
名詞縮寫
1.　緒論
1.1　引言
1.2　細胞吸附作用的生理功能與參與的分子
1.3　Integrin的結構與生理功能
1.4　血小板之凝血機轉
1.5　蛇毒蛋白和disintegrin
1.6　MDC的分子結構及可能的生理功能
1.7　近代針對細胞吸附作用的藥物發展
1.8　本實驗之研究動機與重要性
1.9　本實驗之主要目標
1.10　本實驗之研究策略
2.　材料
2.1　藥品
2.2　實驗套組
2.3　抗體
2.4　酵素
2.5　大腸桿菌株
2.6　細胞株
2.7　質體
2.8　引子(Primers)
3.　方法
3.1　Rhodostomin片段之序列相似性比對
3.2　質體之構築
3.3　大腸桿菌之轉形(Transformation)
3.4　GST融合蛋白之純化
3.5　人類血小板之製備
3.6　SDS－聚丙烯醯電泳(SDS-PAGE)
3.7　西方點墨法(Western blot)
3.8　GST攫取試驗
3.9　血小板凝集試驗
3.10　細胞吸附試驗
4.　結果
4.1　Rhodostomin片段之序列相似性比對
4.2　以插入突變鑑定RGD序列與鄰近氨基酸相對位置之重要性
4.3　以取代突變進一步探討Pro53對於rhodostomin與integrin結合之影響
4.4　人類MDC-15之disintegrin相似區域與integrin結合之能力明顯低於rhodostomin
4.5　以區域置換突變來探討rhodostomin分子骨架之重要性
4.6　利用D15之取代突變種來採討Pro之功能
4.7　總結
5.　討論
5.1　序列相似性比對之分析結果
5.2　GST融合蛋白之生物活性
5.3　Ala插入突變對於rhodostomin生物活性的影響
5.4　Pro53對於rhodostomin生物活性的重要性
5.5　Rhodostomin分子骨架的功能
5.6　MDC-15之disintegrin-like區域可能的生物功能
5.7　Integrin與ligand交互作用的可能機轉
5.8　蛇毒研究之未來展望及臨床應用
6.　參考文獻
7.　圖表
8.　附錄
8.1　發表論文
8.2　CD-ROM光碟權利宣告及使用方式

1.楊玉齡, 及 羅時成. (1996) 台灣蛇毒傳奇. 天下文化出版股份有限公司, 台北, 台灣.
2.Adams, J.C. (1997) Cell adhesion-spreading frontiers, intricate insights. Trends Cell Biol. 7, 107-110.
3.Adler, M., Lazarus, R.A., Dennis, M.S., Wagner, G. (1991) Solution structure of kistrin, a potent platelet aggregation inhibitor and GP IIb-IIIa antagonist. Science 253, 445-448.
4.Alberts, B., Bray, D., Lewis, J., Raff, M., Roberts, K., and Watson J.D. (1994a) The evolution of the cell. in “Molecular biology of the cell”, 3rd ed., Garland Publishing, Inc., New York. Chapter 1, 3-40.
5.Alberts, B., Bray, D., Lewis, J., Raff, M., Roberts, K., and Watson J.D. (1994b) Cell junctions, cell adhesion, and the extracellular matrix. in “Molecular biology of the cell”, 3rd ed., Garland Publishing, Inc., New York. Chapter 19, 949-1009.
6.Aplin, A.E., Howe, A., Alahari, S.K., and Juliano, R.L. (1998) Signal transduction and signal modulation by cell adhesion receptors: The role of integrin, cadherins, immunoglobulin-cell adhesion molecules, and selectins. Pharmacol. Rev. 50, 197-263.
7.Arcangelis, A.D., and Georges-Labouesse, E. (2000) Integrin and ECM functions. Roles in vertebrate development. Trends Genet. 16, 389-395.
8.Au, L.-C., Huang, Y.-B., Huang, T.-F., Teh, G.-W., Lin, H.-H., and Choo, K.-B. (1991) A common precursor for a putative hemorrhagic protein and rhodostomin, a platelet aggregation inhibitor of the Calloselasma rhodostoma: Molecular cloning and sequence analysis. Biochem. Biophys. Res. Commun. 181, 585-593.
9.Au, L.-C., Chou, J.-S., Chang, K.-J., Teh, G.-W., and Lin, S.-B. (1993) Nucleotide sequence of a full-length cDNA encoding a common precursor of platelet aggregation inhibitor and hemorrhagic protein from Calloselasma rhodostoma venom. Biochim. Biophys. Acta. 1173, 243-245.
10.Bigler, D., Chen, M., Water, S., and White, J.M. (1997) A model for sperm-egg binding and fusion based on ADAMs and integrins. Trends Cell Biol. 7, 220-225.
11.Black, R.A., and White, J.M. (1998) ADAMs: Focus on the protease domain. Curr. Opin. Cell Biol. 10, 654-659.
12.Blobel, C.P., Myles, D.G., Primakoff, P., and White, J.M. (1990) Proteolytic processing of a protein involved in sperm-egg fusion correlates with acquisition of fertilization competence. J. Cell Biol. 111, 69-78.
13.Blobel, C.P., and White, J.M. (1992) Structure, function and evolutionary relationship of proteins containing a disintegrin domain. Curr. Opin. Cell Biol. 4, 760-765.
14.Blobel, C.P., Wolfsberg, T.G., Turck, C.W., Myles, D.G., Primakoff, P., and White, J.M. (1992) A potential fusion peptide and an integrin ligand domain in a protein active in sperm-egg fusion. Nature 356, 248-252.
15.Bornstein, P., Armstrong, L.C., Hankenson, K.D., Kyriakides, T.R., and Yang, Z. (2000) Thrombospondin 2, a matricellular protein with diverse functions. Matrix Biol. 19, 557-568.
16.Boudreau, N.J., and Jones, P.L. (1999) Extracellular matrix and integrin signaling: The shape of things to come. Biochem. J. 339, 481-488.
17.Calvete, J.J., Schäfer, W., Soszka, T., Lu, W., Cook, J.J., Jameson, B.A., and Niewiarowski, S. (1991) Identification of the disulfide bond pattern in albolabrin an RGD-containing peptide from the venom of Trimeresurus albolabris: Significance for the expression of platelet aggregation inhibitory activity. Biochemistry-US 30, 5225-5229.
18.Calvete, J.J., Jurgens, M., Marcinkiewicz, C., Romero, A., Schrader, M., and Niewiarowski, S. (2000) Disulphide-bond pattern and molecular modeling of the dimeric disintegrin EMF-10, a potent and selective integrin a5b1 antagonist from Eristocophis macmahoni venom. Biochem. J. 345, 573-581.
19.Camper, L., Hellman, U., and Lundgren-Åkerlund, E. (1998) Isolation, cloning, and sequence analysis of the integrin subunit a10, a b1-associated collagen binding integrin expressed on chondrocytes. J. Biol. Chem. 273, 20383-20389.
20.Chang, H.-H., Hu, S.-T., Huang, T.-F., Chen, S.-H., Lee, Y.-H. W., and Lo., S.J. (1993) Rhodostomin, an RGD-containing peptide expressed from a synthetic gene in Escherichia coli, facilitates the attachment of human hepatoma cells. Biochem. Biophys. Res. Commun. 190, 242-249.
21.Chang, H.-H., Tsai, W.-J., and Lo, S.J. (1997) Glutathione S-transferase- rhodostomin fusion protein inhibits platelet aggregation and induces platelet shape change. Toxicon 35, 195-204.
22.Chang, H.-H., and Lo, S.J. (1998) Full-spreading platelets induced by the recombinant rhodostomin are via binding to integrins and correlated with FAK phosphorylation. Toxicon 36, 1087-1099.
23.Chang, H.-H., Lin, C.-H., and Lo, S.J. (1999) Recombinant rhodostomin substrates induce human platelet full transformation and active calcium oscillation. Exp. Cell Res. 250, 387-400.
24.Chang, H.-H., Chang, C.-P., Chang, J.-C., Dung, S.-Z., and Lo, S.J. (1997) Application of recombinant rhodostomin in studying cell adhesion. J. Biomed. Sci. 4, 235-243.
25.Chang, J.-C. (1997) Characterization of recombinant rhodostomin-binding proteins on the membrane of mouse melanoma cells and human platelets. Master thesis. Institute of Microbiology and Immunology, National Yang-Ming University.
26.Chen, M.S., Almeida, E.A.C., Huovila, A.-P.J., Takahashi, Y., Shaw, L.M., Mercurio, A.M., and White, J.M. (1999) Evidence that distinct states of the integrin a6b1 interact with laminin and an ADAM. J. Cell Biol. 144, 549-561.
27.Chiang, H.-S., Swaim, M.W., and Huang, T.-F. (1994) Characterization of platelet aggregation induced by human colon adenocarcinoma cells and its inhibition by snake venom peptides, trigramin and rhodostomin. Brit. J. Haematol. 87, 325-331.
28.Chung, J., Gao, A.-G., and Frazier, W.A. (1997) Thrombospondin acts via integrin-associated protein to activate the platelet integrin aIIbb3. J. Biol. Chem. 272, 14740-14746.
29.Clark, E.A., and Brugge, J.S. (1995) Integrins and signal transduction pathways: The road taken. Science 268, 233-239.
30.Clark, E.A., King, W.G., Brugge, J.S., Symons, M., and Hynes, R.O. (1998) Integrin-mediated signals regulated by members of the Rho family of GTPase. J. Cell Biol. 142, 573-586.
31.Davis, B.D., Dulbecco, R., Eisen, H.N., and Ginsberg, H.S. (1990a) Nutrition; energy, membrane transport; chemotaxis. in “Microbiology”, 4th ed., Harper and Row, Inc., New York. Chapter 4, 65-90.
32.Davis, B.D., Dulbecco, R., Eisen, H.N., and Ginsberg, H.S. (1990b) Host-parasite relations in bacterial infections. in “Microbiology”, 4th ed., Harper and Row, Inc., New York. Chapter 21, 485-505.
33.Dedhar, S., and Hannigan, G.E. (1996) Integrin cytoplasmic interactions and bi-directional transmembrane signaling. Curr. Opin. Cell Biol. 8, 657-669.
34.Dennis, M.S., Henzel, W.J., Pitti, R.M., Lipari, M.T., Napier, M.A., Deisher, T.A., Bunting, S., and Lazarus, R.A. (1989) Platelet glycoprotein IIb-IIIa protein antagonists from snake venoms: Evidence for a family of platelet-aggregation inhibitors. Proc. Natl. Acad. Sci. U.S.A. 87, 2471-2475.
35.Dennis, M.S., Carter, P., and Lazarus, R.A. (1993) Binding interactions of kistrin with platelet glycoprotein IIb-IIIa: Analysis by site-directed mutagenesis. Proteins 15, 312-321.
36.Dickeson, S.K., and Santoro, S.A. (1998) Ligand recognition by the I domain-containing integrins. Cell. Mol. Life Sci. 54, 556-566.
37.D’Souza, S.E., Haas, T.A., Piotrowicz, R.S., Byers-Ward, V., McGrath, D.E., Soule, H.R., Cierniewski, C., Plow, E.F., and Smith, J.W. (1994) Ligands and cation binding are dual functions of a discrete segment of the integrin b3 subunit: Cation displacement is involved in ligand binding. Cell 79, 659-667.
38.Evans, J.P., Schultz, R.M., and Kopf, G.S. (1998) Role of the dinintegrin domains of mouse fertilins a and b in fertilization. Biol. Reprod. 59, 145-152.
39.Frenette, P.S., and Wagner, D.D. (1996) Molecular medicine. Adhesion molecules-part I. N. Engl. J. Med. 334, 1526-1529.
40.Frisch, S.M., and Ruoslahti, E. (1997) Integrins and anoikis. Curr. Opin. Cell Biol. 9, 701-706.
41.Frojmovic, M.M. (1998) Platelet aggregation in flow: Differential roles for adhesive receptors and ligands. Am. Heart J. 135, S119-S131.
42.Gao, A.-G., Lindberg, F.P., Dimitry, J.M., Brown, E.J., and Frazier, W.A. (1996) Thrombospondin modulates avb3 function through integrin-associated protein. J. Cell Biol. 135, 533-544.
43.George, J.N., Caen, J.P., and Nurden, A.T. (1990) Glanzmann’s thrombasthenia: The spectrum of clinical disease. Blood 75, 1383-1395.
44.Giancotti, F.G., and Ruoslahti, E. (1999) Integrin signaling. Science 285, 1028-1032.
45.Green, L.J., Mould, A.P., and Humphries, M.J. (1998) The integrin b subunit. Int. J. Biochem. Cell Biol. 30, 179-184.
46.Gould, R.J., Polokoff, M.A., Friedman, P.A., Huang, T.-F., Holt, J.C., Cook, J.J., and Niewiarowski, S. (1990) Dinintegrin: A family of integrin inhibitory ptoteins from viper venoms. Proc. Soc. Exp. Biol. Med. 195, 168-171.
47.Guex, N., Diemand, A., and Peitsch, M.C. (1999) Protein modeling for all. Trends Biochem. Sci. 24, 364-367.
48.Guex, N., and Peitsch, M.C. (1997) SWISS-MODEL and the Swiss-PdbViewer: An environment for comparative protein modeling. Electrophoresis 18, 2714-2723.
49.Guilherme, A., Torres, K., and Czech, M.P. (1998) Cross-talk between insulin receptor and integrin a5b1 signaling pathway. J. Biol. Chem. 273, 22899-22903.
50.Gumbiner, B.M. (1996) Cell adhesion: The molecular basis of tissue architecture and morphogenesis. Cell 84, 345-357.
51.Guo, R.-T., Chou, L.-J., Chen, Y.-C., Chen, C.-Y., Pari, K., Jen, C.J., Lo, S.J., Huang, S.-L., Lee, C.-Y., Chang, T.-W., and Chaung, W.-J. (2001) Expression in Pichia pastoris and characterization by circular dichroism and NMR of rhodostomin. Proteins 43, 499-508.
52.Haas, T.A., and Plow, E.F. (1994) Integrin-ligand interactions: A year in review. Curr. Opin. Cell Biol. 6, 656-662.
53.Hautanen, A., Gailit, J., Mann, D.M., and Ruoslahti, E. (1989) Effects on modifications of the RGD sequence and its context on recognition by the fibronectin receptor. J. Biol. Chem. 264, 1437-1442..
54.Hemler, M.E. (1998) Integrin associated proteins. Curr. Opin. Cell Biol. 10, 578-585.
55.Herren, B., Raines, E.W., and Ross, R. (1997) Expression of a disintegrin-like protein in cultured human vascular cells and in vivo. FASEB J. 11, 173-180.
56.Horwitz, A.F., and Hunter, T. (1996) Cell adhesion: Integrating circuitry. Trends Cell Biol. 6, 460-461.
57.Howe, A., Aplin, A.E., Alahari, S.K., and Juliano, R.L. (1998) Integrin signaling and cell growth control. Curr. Opin. Cell Biol. 10, 220-231.
58.Huang, T.-F., Wu, Y.-J., and Ouyang, C. (1987a) Characterization of a potent platelet aggregation inhibiyor from Agkistrodon rhodostoma snake venom. Biochim. Biophy. Acta. 925, 248-257.
59.Huang, T.-F., Holt, J.C., Lukasiewicz, H., and Niewiarowski, S. (1987b) Trigramin: A low molecular weight peptide inhibiting fibrinogen interaction with platelet receptors expressed on glycoprotein IIb-IIIa complex. J. Biol. Chem. 262, 16157-16163.
60.Huang, T.-F., Holt, J.C., Kirby, E.P., and Niewiarowski, S. (1989) Trigramin: Promary structure and its inhibition of von Willebrand factor binding to glycoprotein IIb/IIIa complex on human platelets. Biochemistry-US 28, 661-666.
61.Huang, T.-F. (1998) What have snakes taught us about integrins? Cell. Mol. Life Sci. 54, 527-540.
62.Huang, T.-F., and Niewiarowski, S. (1994) Disintegrins: The naturally -occurring antagonists of platelet fibrinogen receptor. J. Toxicol. 13, 253-273.
63.Humphries, M.J. (1996) Integrin activation: The link between ligand binding and signal transduction. Curr. Opin. Cell Biol. 8, 632-640.
64.Humphries, M.J. (1996) Integrin cell adhesion receptors and the concept of agonism. Trends Pharmacol. Sci. 21, 29-32.
65.Humphries, M.J., and Newham, P. (1998) The structure of cell-adhesion molecules. Trends Cell Biol. 8, 78-83.
66.Huovila, A.-P. J., Almeida, E.A., and White, J.M. (1996) ADAMs and cell fusion. Curr. Opin. Cell Biol. 8, 692-699.
67.Hynes, R.O. (1987) Integrins: A family of cell surface receptors. Cell 48, 549-554.
68.Hynes, R.O. (1992) Integrin: Versatility, modulation, and signaling in cell adhesion. Cell 69, 11-25.
69.Hynes, R.O. (1999) Cell adhesion: Old and new questions. Trends Cell Biol. 9, M33-M37.
70.Hynes, R.O., and Zhao, Q. (2000) The evolution of cell adhesion. J. Cell Biol. 150, F89-F95.
71.Jia, L.-G., Wang, X.-M., Shannon, J.D., Bjarnason, J.B., and Fox, J.W. (1997) Function of disintegrin-like/cysteine-rich domains of atrolysin A. J. Biol. Chem. 272, 13094-13102.
72.Jiang, S.-T., Chiang, H.-C., Cheng, M.-H., Yang, T.-P., Chuang, W.-J., and Tang, M.-J. (1999) Role of fibronectin deposition in cystogenesis of Madin-Darby canine kidney cells. Kidney Int. 56, 92-103.
73.Jiang, S.-T., Chuang, W.-J., and Tang, M.-J. (2000) Role of fibronectin deposition in branching morphogenesis of Madin-Darby canine kidney cells. Kidney Int. 57, 1860-1867.
74.Kamata, T., Irie, A., Tokuhira, M., and Takada, Y. (1996) Critical residues of integrin aIIb subunit for binding of aIIbb3(glycoprotein IIb-IIIa) to fibrinogen and ligand-mimetic antibodies(PAC-1, OP-G2, and LJ-CP3) J. Biol. Chem. 271, 18610-18615.
75.Kini, R.M., and Evans, H.J. (1992) Structural domains in venom proteins: Evidence that metallopoteinases and nonenzymatic platelet aggregation inhibitors(disintegrins) from venoms are derived by proteolysis from a common precursor. Toxicon 30, 265-293.
76.Kosfeld, M.D., and Frazier, W.A. (1992) Identification of active peptide sequences in the carboxyl-terminal cell binding domain of human thrombospondin-1. J. Biol. Chem. 267, 16230-16236.
77.Krätzschmar J., Lum, L., and Blobel, C.P. (1996) Metargidin, a membrane-anchored metalloprotease-disintegrin protein with an RGD integrin binding sequence. J. Biol. Chem. 271, 4593-4596.
78.Kuhara, T., Kagami, S., and Kuroda, Y. (1997) Expression of b1-integrins on activated mesangial cells in human glomerulonephritis. J. Am. Soc. Nephrol. 8, 1679-1687.
79.Kumar, C.C. (1998) Signaling by integrin receptors. Oncogene 17, 1365-1373.
80.Laemmli, U.K. (1970) Cleavage of structural protein during the assembly of the head of bacteriophage T4. Nature 227, 680-685.
81.Lawler, J. (2000) The functions of throbospondin-1 and —2. Curr. Opin. Cell Biol. 12, 634-640.
82.Lee, C.-Y. (1972) Chemistry and pharmacology of polypeptide toxins in snake venoms. Annu. Rev. Pharmacol. 12, 265-286.
83.Lee, C.-Y., Ouyang, C., and Chang, C.-C. (1973) Collected paper on snake venom. Pharmacological Institute, National Taiwan University.
84.Lee, L.-W., Peng, H.-C., Ko, W.-C., Hung, W.-C., Su, C.-H., Lin, C.-H., Huang, T.-F., Yen, M.-H., and Sheu, J.-R. (1999) Triflavin potentiates the antiplatelet activity of platelet activating factor receptor antagonist on activated neurophil-induced platelet aggregation. Eur. J. Pharmacol. 294, 231-238.
85.Leung, L.L.K. (1984) Role of thrombospondin in platelet aggregation. J. Clin. Invest. 74, 1764-1772.
86.Liu, C.-Z., Wang, Y.-W., Shen, M.-C., and Huang, T.-F. (1994) Analysis of human platelet glycoprotein IIb-IIIa by fluorescein isothiocyanate- conjugated disintegrins with flow cytometry. Thromb. Haemost. 72, 919-925.
87.Lodish, H., Berk, A., Zipursky, S.L., Matsudaira, P., Baltimore, D., and Darnell, J.E. (1999a) Integrating cells into tissues. in “Molecular cell biology”, 4th ed., W. H. Freeman and Company, New York. Chapter 22, 968-1002.
88.Lodish, H., Berk, A., Zipursky, S.L., Matsudaira, P., Baltimore, D., and Darnell, J.E. (1999b) Protein structure and function. in “Molecular cell biology”, 4th ed., W. H. Freeman and Company, New York. Chapter 3, 968-1002.
89.Lombardi, P., Pelagalli, A., Avallone, L., d’Angelo, D., Belisario, M.A., d’Angelo, A., and Staiano, N. (1999) Species-dependent specificity of platelet aggregation inhibitors from snake venom. J. Comp. Path. 121, 185-190.
90.Longhurst C.M., and Jennings, L.K. (1998) Integrin-mediated signal transduction. Cell. Mol. Life Sci. 54, 514-526.
91.Lu, X., Deadman, J.J., Williams, J.A., Kakkar, V.V., and Rahman, S. (1993) Synthetic RGD peptides derived from the adhesive domains of snake-venom proteins: Evaluation as inhibitors of platelet aggregation. Biochem. J. 296, 21-24.
92.Lu, X., Rahman, S., Kakkar, V.V., and Authi, K.S. (1996) Substitutions of proline 42 to alanine and methionine 46 to asparagines around the RGD Domain of the neurotoxin dendroaspin alter its preferential antagonism to that resembling the disintegrin elegabtin. J. Biol. Chem. 271, 289-294.
93.Lu, X., Sun, Y., Shang, D., Wattam, B., Egglezou, S., Hughes, T., Hyde, E., Scully, M., and Kakkar, V. (2001) Evaluation of the role of proline residues flanking the RGD motif of dendroaspin, an inhibitor of platelet aggregation and cell adhesion. Biochem. J. 355, 633-638.
94.Marcinkiewicz, C., Calvete, J.J., Marcinkiewicz, M.M., Raida, M., Vijay-Kumar, S., Huang, Z., Lobb, R.R., and Niewiarowski, S. (1999a) EC3, a novel heterodimeric disintegrin from Echis carinatus venom, inhibits a4 and a5 integrins in an RGD-independent manner. J. Biol. Chem. 274, 12468-12473.
95.Marcinkiewicz, C., Calvete, J.J., Vijay-Kumar, S., Marcinkiewicz, M.M., Raida, M., Schick, P., Lobb, R.R., and Niewiarowski, S. (1999b) Structural and functional characterization of EMF10, a heterodimeric disintegrin from Eristocophis macmahoni venom that selectively inhibits a5b1 integrin. Biochemistry-US 38, 13302-13309.
96.McLane, M.A., Kowalska, M.A., Silver, L., Shattil, S.J., and Niewiarowski, S. (1994) Interaction of disintegrins with the aIIbb3 receptor on resting and activated human platelets. Biochem. J. 301, 429-436.
97.Meredith, Jr. J.E., and Schwartz, M.A. (1997) Integrins, adhesion and apoptosis. Trends Cell Biol. 7, 146-150.
98.Mondal, K., Lofquist, A.K., Watson, J.M., Morris, J.S., Price, L.K., and Haskill, J.S. (1995) Adhesion and direct integrin engagement differentially regulate gene transcription, transcript stabilization and translation. Biochem. Soc. T. 23, 460-464.
99.Mould, A.P., Akiyama, S.K., and Humphries, M.J. (1996) The inhibitory anti-b1 integrin monoclonal antibody 13 recognizes an epitope that is attenuated by ligand occupancy. J. Biol. Chem. 271, 20365-20374.
100.Mould, A.P., Askari, J.A., Aota S.-I., Yamada, K.M., Irie, A., Takada, Y., Mardon, H.J., and Humphries, M.J. (1997) Defining the topology of integrin a5b1-fibronectin interactions using inhibitory anti-a5 and anti-b1 monoclonal antibodies. J. Biol. Chem. 271, 20365-20374.
101.Mould, A.P., Garratt, A.N., Puzon-McLaughlin, W., Takada, Y., and Humphries, M.J. (1998) Regulation of integrin function: Evidance that bivalent-cation-induced conformational changes lead to the unmasking of ligand-binding sites within integrin a5b1. Biochem. J. 331, 821-828.
102.Mousa, S.A., and Cheresh, D.A. (1997) Recent advances in cell adhesion molecules and extracellular matrix proteins: Potential clinical implications. Drug Discov. Today 2, 187-199.
103.Murase, S., and Schuman, E.M. (1999) The role of cell adhesion molecules in synaptic plasticity and memory. Curr. Opin. Cell Biol. 11, 549-553.
104.Mustard, J.F., Kinlough-Rathbone, R.L., and Packham, M.A. (1989) Isolation of human platelet from plasma by centrifugation and washing. Method Enzymol. 169, 3-11.
105.Naz, R.K., Phillips, T.M., and Rosenblum, B.B. (1986) Characterization of the fertilization antigen 1 for the development of a contraceptive vaccine. Proc. Natl. Acad. Sci. U.S.A. 83, 5713-5717.
106.Newham, P., Craig, S.E., Clark, K., Mould, A.P., and Humphries, M.J. (1998) Analysis of ligand-induced and ligand-attenuated epitopes on the leukocyte integrin a4b1: VCAM-1, mucosal addressin cell adhesion molecule-1, and fibronectin induce distinct conformational change. J. Immunol. 160, 4508-4517.
107.Nobes, C.D., and Hall, A. (1999) Rho GTPase control polarity, protrusion, and adhesion during cell movement. J. Cell Biol. 144, 1235-1244.
108.Nurden, P., Heilmann, E., Paponneau, A., and Nurden, A. (1994) Two-way trafficking of membrane glycoproteins on thrombin-activated human platelets. Semin. Hematol. 31, 240-250.
109.Ouyang, C., Teng, C.-M., and Huang, T.-F. (1992) Characterization of snake venom components acting on blood coagulation and platelet function. Toxicon 30, 945-966.
110.Pabla, R., Weyrich, A.S., Dixon, D.A., Bray, P.F., McIntyre, T.M., Prescott, S.M., and Zimmerman, G.A. (1999) Integrin-dependent control of translation: Engagement of integrin aIIbb3 regulates synthesis of proteins in activated human platelets. J. Cell Biol. 144, 175-184.
111.Parise, L.V. (1999) Integrin aIIbb3 signalling in platelet adhesion and aggregation. Curr. Opin. Cell Biol. 11, 597-601.
112.Peitsch, M.C. (1995) Protein modeling by E-mail. Bio/Technology 13, 658-660.
113.Peyman, A., Wehner, V., Knolle, J., Stilz, H.U., Breipohl, G., Scheunemann, K.-H., Carniato, D., Ruxer, J.-M. Gourvest, J.-F., Gadek, T.R., and Bodary, S. (2000) RGD mimetics containing a central hydantoin scaffold: avb3 vs aIIbb3 selectivity requirements. Bioorg. Med. Chem. Lett. 10, 179-182.
114.Pfaff, M., Tangemann, K., Müller, B., Gurrath, M., Müller, G., Kessler, H., Timpl, R., and Engel, J. (1994) Selective recognition of cyclic RGD peptides of NMR defined conformation by aIIbb3, avb3, and a5b1 integrins. J. Biol. Chem. 269, 20233-20238.
115.Pierschbacher, M.D., and Ruoslahti, E. (1984) Cell attachment activity of fibronectin can be duplicated by small synthetic fragments of the molecule. Nature 309, 30-33.
116.Pierschbacher, M.D., and Ruoslahti, E. (1987) Influence of stereochemistry of the sequence Arg-Gly-Asp-Xaa on binding specificity in cell adhesion. J. Biol. Chem. 262, 17294-17298.
117.Plow, E.F., Haas, T.A., Zhang, L., Loftus, J., and Smith, J.W. (2000) Ligand binding to integrins. J. Biol. Chem. 275, 21785-21788.
118.Plow, E.F., Pierschbacher, M.D., Ruoslahti, E., Marguerie, G., and Ginsberg, M.H. (1987) Arginyl-Glycyl-Aspartic acid sequences and fibrinogen binding to platelets. Blood 70, 110-115.
119.Porter, J.C., and Hogg, N. (1998) Integrins take partners: Cross-talk between integrins and other membrane receptors. Trends Cell Biol. 8, 390-396.
120.Prater, C.A., Plotkin, J., Jaye, D., and Frazier, W.A. (1991) The properdin-like type I repeats of human thrombospondin contain a cell attachment site. J. Cell Biol. 112, 1031-1040.
121.Primakoff, P., Hyatt, H., and Tredick-Kline, J. (1987) Identification and purification of a sperm surface protein with a potential role in sperm-egg membrane fusion. J. Cell Biol. 104, 141-149.
122.Quinn, M., and Fitzgerald, D.J. (1998) Long-term administration of glycoprotein IIb/IIIa antagonists. Am. Heart J. 135, S113-S118.
123.Rahman, S., Lu, X., Kakkar, V.V., and Authi, K.S. (1995) The integrin aIIbb3 contains distinct and interacting binding sites for snake-venom RGD(Arg-Gly-Asp) proteins. Biochem. J. 335, 247-257.
124.Rahman, S., Aitken, A., Flynn, G., Formstone, C., and Savidge, G.F. (1998) Modulation of RGD sequence motifs regulates disintegrin recognition of aIIbb3 and a5b1 integrin complexes. Biochem. J. 335, 247-257.
125.Re, F., Zanetti, A., Sironi, M., Polentarutti, N., Lanfrancone, L., Dejana, E., and Colotta, F. (1994) Inhibition of anchorage-dependent cell spreading triggers apoptosis in culture human endothelial cells. J. Cell Biol. 127, 537-546.
126.Retta, S.F., Balzac, F., Ferraris, P., Belkin, A.M., Fässler, R., Humphries M.J., Leo, G.D., Silengo, L., and Tarone, G. (1998) b1-integrin cytoplasmic subdomains involved in dominant negative function. Mol. Biol. Cell 9, 715-731.
127.Rooke, J., Pan, D., Xu, T., and Rubin, G.M. (1996) KUZ, a conserved metalloprotease-disintegrin protein with two roles in Drosophila neurogenesis. Science 273, 1227-1231.
128.Ruoslahti, E., and Pierschbacher, M.D. (1986) Arg-Gly-Asp: A versatile cell recognition signal. Cell 44, 517-518.
129.Ruoslahti, E., and Pierschbacher, M.D. (1987) New perspectives in cell adhesion: RGD and integrins. Science 238, 491-497.
130.Ruoslahti, E., and Reed, J.C. (1994) Anchorage dependence, integrins, and apoptosis. Cell 77, 477-478.
131.Sarkar, S., Rooney, M.M. and Lord, S.T. (1999) Activation of integrin-b3-associated syk in platelets. Biochem. J. 338, 677-680.
132.Savage, B., Almus-Jacobs, F., and Ruggeri, Z.M. (1998) Specific synergy of multiple substrate-receptor interactions in platelet thrombus fomation under flow. Cell 94, 657-666.
133.Scaloni, A., Martino, E.D., Miraglia, N., Pelagalli, A., Morte, R.D., Staiano, N., and Pucci, P. (1996) Amino acid sequence and molecular modelling of glycoprotein IIb-IIIa and fibronectin receptor iso-antagonists from Trimeresurus elegans venom. Biochem. J. 319, 775-782.
134.Scarborough, R.M., Rose, J.W., Naughton, M.A., Phillips, D.R., Nannizzi, L., Arfsten, A., Campbell, A.M., and Charo, I.F. (1993a) Characterization of the integrin specificities of disintegrins isolated from American pit viper venom. J. Biol. Chem. 268, 1058-1065.
135.Scarborough, Naughton, M.A., Teng, W., Rose, J.W., Phillips, D.R., Nannizzi, L., Arfsten, A., Campbell, A.M., and Charo, I.F. (1993b) Design of potent and specific integrin antagonists. J. Biol. Chem. 268, 1066-1073.
136.Schmitz, G., Rothe, G., Ruf, A., Barlage, S., Tschöpe, D., Clemetson, K.J., Goodall, A.H., Michelson, A.D., Nurden, A.T., and Shankey, T.V. European working group on clinical cell analysis: Consensus protocol for the flow cytometric characterization of platelet function. Thromb. Haemost. 79, 885-896.
137.Schoenwaelder, S.M., and Burridge, K. (1999) Bidirectional signaling between the cytokeleton and integrins. Curr. Opin. Cell Biol. 11, 274-286.
138.Schwartz, M.A., and Ingber, D.E. (1994) Integrating with integrins. Mol. Biol. Cell 5, 389-393.
139.Schwartz, M.A., Schaller, M.D., and Ginsberg, M.H. (1995) Integrin: Emerging paradigms of signal transduction. Annu. Rev. Cell Dev. Biol. 11, 549-599.
140.Sechler, J.L., Corbett, S.A., and Schwarzbauer, J.E. (1997) Modulatory roles for integrin activation and the synergy site of fibronectin during matrix assembly. Mol. Biol. Cell 8, 2563-2573.
141.Shattil, S.J., Ginsberg, M.H., and Brugge, J.S. (1994) Adhesive signaling in platelets. Curr. Opin. Cell Biol. 6, 695-704.
142.Shattil, S.J., Gao, J., and Kashiwagi, H. (1997) Not just another pretty face: Regulation of platelet function at the cytoplasmic face of integrin aIIbb3. Thromb. Haemost. 78, 220-225.
143.Shattil, S.J., Kashiwagi, H., and Pampori, N. (1998) Integrin signaling: The platelet paradigm. Blood 91, 2645-2657.
144.Sims, P.J., Ginsberg, M.H., Plow, E.F., and Shattil, S.J. (1991) Effect of platelet activation on the conformation of the plasma membrane glycoprotein IIb-IIIa complex. J. Biol. Chem. 266, 7345-7352.
145.Sixma, J.J., Zanten, G.H.V., Huizinga, E.G., Plas, R.M.V.D., Verkley, M., Wu, Y.-P., Gros, P., and Groot, P.G.D. (1997) Platelet adhesion to collagen: An update. Thromb. Haemost. 78, 434-438.
146.Smith, D.B., and Johnson, K.S. (1988) Single-step purification of polypeptides expressed in Escherichia coli as fusions with glutathione S-transferase. Gene 67, 31-40.
147.Smith, J.W., Piotrowicz, R.S., and Mathis, D. (1994a) A mechanism for divalent cation regulation of b3-integrins. J. Biol. Chem. 269, 960-967.
148.Smith, J.W., Hu, D., Satterthwait, A., Pinz-Sweeney, S., and Barbas, C.F. (1994b) Building synthetic antibodies as adhesive ligands for integrins. J. Biol. Chem. 269, 32788-32795.
149.Stryer, L. (1988a) Prelude. in “Biochemistry”, 3rd ed., W. H. Freeman and Company, New York. Chapter 1, 3-14.
150.Stryer, L. (1988b) Biosynthesis of amino acids and heme. in “Biochemistry”, 3rd ed., W. H. Freeman and Company, New York. Chapter 24, 592-593.
151.Stryer, L. (1988c) Protein structure and function. in “Biochemistry”, 3rd ed., W. H. Freeman and Company, New York. Chapter 2, 32-35.
152.Takagi, J., Kamata, T., Meredith, J., Puzon-McLaughlin, W., and Takada, Y. (1997) Changing ligand specificities of avb1 and avb3 integrins by swapping a short diverse sequence of the b subunit. J. Biol. Chem. 272, 19794-19800.
153.Tamku, J.W., DeSimone, D.W., Fonda, D., Patel, R.S., Buck, C., Horwitz, A.F., and Hynes, R.O. (1986) Structure of integrin, a glycoprotein involved in the transmembrane linkage between fibronectin and actin. Cell 46, 271-282.
154.Teng, C.-M., and Huang, T.-F. (1991) Snake venom constituents that affect platelet function. Platelets 2, 77-87.
155.Tlsty, T.D. (1998) Cell-adhesion-dependent influences on genomic instability and carcinogenesis. Curr. Opin. Cell Biol. 10, 647-653.
156.Towbin, H., Staehelin, T., and Gordon, J. (1979) Electrophoretic transfr of proteins from polyacrylamide gels to nitrocellulose sheets: Procedure and applications. Proc. Natl. Acad. Sci. U.S.A. 76,4350-4354.
157.Tsai, J.-W. (2000) Using optical tweezers in biological force measurements. Master thesis. Institute of Microbiology and Immunology, National Yang-Ming University.
158.Tselepis, V.H., Green, L.J., and Humphries, M.J. (1997) An RGD to LDV motif conversion within the disintegrin kistrin generates an integrin antagonist that retains poteny but exhibits altered receptor specificity. J. Biol. Chem. 272, 21341-21348.
159.Velling, T., Kusche-Gullberg, M., Sejersen, T., and Gullberg, D. (1999) cDNA cloning and chromosomal localization of human a11 integrin. J. Biol. Chem. 274, 25735-25742.
160.Wattam, B., Shang, D., Rahman, S., Egglezou, S., Scully, M., Kakkar, V., and Lu, X. (2001) Arg-Tyr-Asp(RYD) and Arg-Cys-Asp(RCD) motifs in dendroaspin promote selective inhibition of b1 and b3 integrins. Biochem. J. 356, 11-17.
161.Wierzbicka-Patynowski, I., Niewiarowsk, S., Marcinkiewicz, C., Calvete, J.J., Marcinkiewicz, M.M., and McLane, M.A. (1999) Structural requirements of echistatin for the recognition of avb3 and a5b1 integrins. J. Biol. Chem. 274, 37809-37814.
162.Wolfsberg, T.G., Primakoff, P., Myles, D.G., and White, J.M. (1995) ADAM, a novel family of membrane proteins containing a disintegrin and metalloprotease domain: Multipotential functions in cell-cell and cell-matrix interaction. J. Cell Biol. 131, 275-278.
163.Woods, Jr. V.L., Wolff, L.E., and Keller, D.M. (1986) Resting platelets contain a substantial centrally located pool of glycoprotein IIb-IIIa complex which may be accessible to some but not other extracellular proteins. J. Biol. Chem. 261, 15242-15251.
164.Woods, A., and Couchman, J.R. (2000) Integrin modulation by lateral association. J. Biol. Chem. 275, 24233-24236.
165.Yagami-Hiromasa, T., Sato, T., Kurisaki, T., Kamijo, K., Nabeshima, Y.-I., and Fujisawa-Sehara, A. (1995) A metalloprotease-disintegrin participating in myoblast fusion. Nature 377, 652-656.
166.Yeh, C.-H., Peng, H.-C., and Huang, T.-F. (1999) Cytokines modulate integrin avb3-mediated human endothelial cell adhesion and calcium signaling. Exp. Cell Res. 251, 57-66.
167.Yeh, C.-H., Peng, H.-C., Yang, R.-S., and Huang, T.-F. (2001) Rhodostomin, a snake venom disintegrin, inhibits angiogenesis elicited by basic fibroblast growth factor and suppresses tumor growth by a selective avb3 blockade of endothelial cells. Mol. Pharmacol. 59, 1333-1342.
168.Zhang, X.-P., Kamata, T., Yokoyama, K., Puzon-McLaughlin, W., and Takada, Y. (1998) Specific interaction of the recombinant disintegrin-like domain of MDC-15(metargidin, ADAM-15) with integrin avb3. J. Biol. Chem. 273, 7345-7350.