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. 1990 Aug 1;111(2):721–729. doi: 10.1083/jcb.111.2.721

An interaction between alpha-actinin and the beta 1 integrin subunit in vitro

PMCID: PMC2116186  PMID: 2116421

Abstract

A number of cytoskeletal-associated proteins that are concentrated in focal contacts, namely alpha-actinin, vinculin, talin, and integrin, have been shown to interact in vitro such that they suggest a potential link between actin filaments and the membrane. Because some of these interactions are of low affinity, we suspect the additional linkages also exist. Therefore, we have used a synthetic peptide corresponding to the cytoplasmic domain of beta 1 integrin and affinity chromatography to identify additional integrin-binding proteins. Here we report our finding of an interaction between the cytoplasmic domain of beta 1 integrin and the actin-binding protein alpha-actinin. Beta 1- integrin cytoplasmic domain peptide columns bound several proteins from Triton extracts of chicken embryo fibroblasts. One protein at approximately 100 kD was identified by immunoblot analysis as alpha- actinin. Solid phase binding assays indicated that alpha-actinin bound specifically and directly to the beta 1 peptide with relatively high affinity. Using purified heterodimeric chicken smooth muscle integrin (a beta 1 integrin) or the platelet integrin glycoprotein IIb/IIIa complex (a beta 3 integrin), binding of alpha-actinin was also observed in similar solid phase assays, albeit with a lower affinity than was seen using the beta 1 peptide. alpha-Actinin also bound specifically to phospholipid vesicles into which glycoprotein IIb/IIIa had been incorporated. These results lead us to suggest that this integrin-alpha- actinin linkage may contribute to the attachment of actin filaments to the membrane in certain locations.

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Selected References

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  1. Arahata K., Ishiura S., Ishiguro T., Tsukahara T., Suhara Y., Eguchi C., Ishihara T., Nonaka I., Ozawa E., Sugita H. Immunostaining of skeletal and cardiac muscle surface membrane with antibody against Duchenne muscular dystrophy peptide. Nature. 1988 Jun 30;333(6176):861–863. doi: 10.1038/333861a0. [DOI] [PubMed] [Google Scholar]
  2. Argraves W. S., Dickerson K., Burgess W. H., Ruoslahti E. Fibulin, a novel protein that interacts with the fibronectin receptor beta subunit cytoplasmic domain. Cell. 1989 Aug 25;58(4):623–629. doi: 10.1016/0092-8674(89)90097-4. [DOI] [PubMed] [Google Scholar]
  3. Argraves W. S., Suzuki S., Arai H., Thompson K., Pierschbacher M. D., Ruoslahti E. Amino acid sequence of the human fibronectin receptor. J Cell Biol. 1987 Sep;105(3):1183–1190. doi: 10.1083/jcb.105.3.1183. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Beckerle M. C. Identification of a new protein localized at sites of cell-substrate adhesion. J Cell Biol. 1986 Nov;103(5):1679–1687. doi: 10.1083/jcb.103.5.1679. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Buck C. A., Horwitz A. F. Cell surface receptors for extracellular matrix molecules. Annu Rev Cell Biol. 1987;3:179–205. doi: 10.1146/annurev.cb.03.110187.001143. [DOI] [PubMed] [Google Scholar]
  6. Burridge K., Fath K., Kelly T., Nuckolls G., Turner C. Focal adhesions: transmembrane junctions between the extracellular matrix and the cytoskeleton. Annu Rev Cell Biol. 1988;4:487–525. doi: 10.1146/annurev.cb.04.110188.002415. [DOI] [PubMed] [Google Scholar]
  7. Burridge K., Mangeat P. An interaction between vinculin and talin. Nature. 1984 Apr 19;308(5961):744–746. doi: 10.1038/308744a0. [DOI] [PubMed] [Google Scholar]
  8. Burridge K., McCullough L. The association of alpha-actinin with the plasma membrane. J Supramol Struct. 1980;13(1):53–65. doi: 10.1002/jss.400130106. [DOI] [PubMed] [Google Scholar]
  9. COHEN C., HOLMES K. C. X-ray diffraction evidence for alpha-helical coiled-coils in native muscle. J Mol Biol. 1963 May;6:423–432. doi: 10.1016/s0022-2836(63)80053-4. [DOI] [PubMed] [Google Scholar]
  10. Campbell K. P., Kahl S. D. Association of dystrophin and an integral membrane glycoprotein. Nature. 1989 Mar 16;338(6212):259–262. doi: 10.1038/338259a0. [DOI] [PubMed] [Google Scholar]
  11. Chen W. T., Singer S. J. Immunoelectron microscopic studies of the sites of cell-substratum and cell-cell contacts in cultured fibroblasts. J Cell Biol. 1982 Oct;95(1):205–222. doi: 10.1083/jcb.95.1.205. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Davison M. D., Baron M. D., Critchley D. R., Wootton J. C. Structural analysis of homologous repeated domains in alpha-actinin and spectrin. Int J Biol Macromol. 1989 Apr;11(2):81–90. doi: 10.1016/0141-8130(89)90047-0. [DOI] [PubMed] [Google Scholar]
  13. Feramisco J. R., Burridge K. A rapid purification of alpha-actinin, filamin, and a 130,000-dalton protein from smooth muscle. J Biol Chem. 1980 Feb 10;255(3):1194–1199. [PubMed] [Google Scholar]
  14. Fyrberg E., Kelly M., Ball E., Fyrberg C., Reedy M. C. Molecular genetics of Drosophila alpha-actinin: mutant alleles disrupt Z disc integrity and muscle insertions. J Cell Biol. 1990 Jun;110(6):1999–2011. doi: 10.1083/jcb.110.6.1999. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Gehlsen K. R., Dillner L., Engvall E., Ruoslahti E. The human laminin receptor is a member of the integrin family of cell adhesion receptors. Science. 1988 Sep 2;241(4870):1228–1229. doi: 10.1126/science.2970671. [DOI] [PubMed] [Google Scholar]
  16. Glenney J. R., Jr, Zokas L. Novel tyrosine kinase substrates from Rous sarcoma virus-transformed cells are present in the membrane skeleton. J Cell Biol. 1989 Jun;108(6):2401–2408. doi: 10.1083/jcb.108.6.2401. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hammonds R. G., Jr Protein sequence of DMD gene is related to actin-binding domain of alpha-actinin. Cell. 1987 Oct 9;51(1):1–1. doi: 10.1016/0092-8674(87)90002-x. [DOI] [PubMed] [Google Scholar]
  18. Horwitz A., Duggan K., Buck C., Beckerle M. C., Burridge K. Interaction of plasma membrane fibronectin receptor with talin--a transmembrane linkage. Nature. 1986 Apr 10;320(6062):531–533. doi: 10.1038/320531a0. [DOI] [PubMed] [Google Scholar]
  19. Hynes R. O. Integrins: a family of cell surface receptors. Cell. 1987 Feb 27;48(4):549–554. doi: 10.1016/0092-8674(87)90233-9. [DOI] [PubMed] [Google Scholar]
  20. Ignatius M. J., Reichardt L. F. Identification of a neuronal laminin receptor: an Mr 200K/120K integrin heterodimer that binds laminin in a divalent cation-dependent manner. Neuron. 1988 Oct;1(8):713–725. doi: 10.1016/0896-6273(88)90170-5. [DOI] [PubMed] [Google Scholar]
  21. Kelly T., Molony L., Burridge K. Purification of two smooth muscle glycoproteins related to integrin. Distribution in cultured chicken embryo fibroblasts. J Biol Chem. 1987 Dec 15;262(35):17189–17199. [PubMed] [Google Scholar]
  22. Koenig M., Monaco A. P., Kunkel L. M. The complete sequence of dystrophin predicts a rod-shaped cytoskeletal protein. Cell. 1988 Apr 22;53(2):219–228. doi: 10.1016/0092-8674(88)90383-2. [DOI] [PubMed] [Google Scholar]
  23. Kunicki T. J., Nugent D. J., Staats S. J., Orchekowski R. P., Wayner E. A., Carter W. G. The human fibroblast class II extracellular matrix receptor mediates platelet adhesion to collagen and is identical to the platelet glycoprotein Ia-IIa complex. J Biol Chem. 1988 Apr 5;263(10):4516–4519. [PubMed] [Google Scholar]
  24. LOWEY S., COHEN C. Studies on the structure of myosin. J Mol Biol. 1962 Apr;4:293–308. doi: 10.1016/s0022-2836(62)80007-2. [DOI] [PubMed] [Google Scholar]
  25. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  26. Lazarides E., Burridge K. Alpha-actinin: immunofluorescent localization of a muscle structural protein in nonmuscle cells. Cell. 1975 Nov;6(3):289–298. doi: 10.1016/0092-8674(75)90180-4. [DOI] [PubMed] [Google Scholar]
  27. Marcantonio E. E., Hynes R. O. Antibodies to the conserved cytoplasmic domain of the integrin beta 1 subunit react with proteins in vertebrates, invertebrates, and fungi. J Cell Biol. 1988 May;106(5):1765–1772. doi: 10.1083/jcb.106.5.1765. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Mimura N., Asano A. Further characterization of a conserved actin-binding 27-kDa fragment of actinogelin and alpha-actinins and mapping of their binding sites on the actin molecule by chemical cross-linking. J Biol Chem. 1987 Apr 5;262(10):4717–4723. [PubMed] [Google Scholar]
  29. Otto J. J. Detection of vinculin-binding proteins with an 125I-vinculin gel overlay technique. J Cell Biol. 1983 Oct;97(4):1283–1287. doi: 10.1083/jcb.97.4.1283. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Parise L. V., Phillips D. R. Fibronectin-binding properties of the purified platelet glycoprotein IIb-IIIa complex. J Biol Chem. 1986 Oct 25;261(30):14011–14017. [PubMed] [Google Scholar]
  31. Parise L. V., Phillips D. R. Reconstitution of the purified platelet fibrinogen receptor. Fibrinogen binding properties of the glycoprotein IIb-IIIa complex. J Biol Chem. 1985 Sep 5;260(19):10698–10707. [PubMed] [Google Scholar]
  32. Ruoslahti E., Pierschbacher M. D. New perspectives in cell adhesion: RGD and integrins. Science. 1987 Oct 23;238(4826):491–497. doi: 10.1126/science.2821619. [DOI] [PubMed] [Google Scholar]
  33. Sonnenberg A., Modderman P. W., Hogervorst F. Laminin receptor on platelets is the integrin VLA-6. Nature. 1988 Dec 1;336(6198):487–489. doi: 10.1038/336487a0. [DOI] [PubMed] [Google Scholar]
  34. Suzuki A., Goll D. E., Singh I., Allen R. E., Robson R. M., Stromer M. H. Some properties of purified skeletal muscle alpha-actinin. J Biol Chem. 1976 Nov 10;251(21):6860–6870. [PubMed] [Google Scholar]
  35. Tamkun J. W., DeSimone D. W., Fonda D., Patel R. S., Buck C., Horwitz A. F., Hynes R. O. Structure of integrin, a glycoprotein involved in the transmembrane linkage between fibronectin and actin. Cell. 1986 Jul 18;46(2):271–282. doi: 10.1016/0092-8674(86)90744-0. [DOI] [PubMed] [Google Scholar]
  36. Tidball J. G. Alpha-actinin is absent from the terminal segments of myofibrils and from subsarcolemmal densities in frog skeletal muscle. Exp Cell Res. 1987 Jun;170(2):469–482. doi: 10.1016/0014-4827(87)90321-1. [DOI] [PubMed] [Google Scholar]
  37. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Wachsstock D. H., Wilkins J. A., Lin S. Specific interaction of vinculin with alpha-actinin. Biochem Biophys Res Commun. 1987 Jul 31;146(2):554–560. doi: 10.1016/0006-291x(87)90564-x. [DOI] [PubMed] [Google Scholar]
  39. Watkins S. C., Hoffman E. P., Slayter H. S., Kunkel L. M. Immunoelectron microscopic localization of dystrophin in myofibres. Nature. 1988 Jun 30;333(6176):863–866. doi: 10.1038/333863a0. [DOI] [PubMed] [Google Scholar]
  40. Wayner E. A., Carter W. G., Piotrowicz R. S., Kunicki T. J. The function of multiple extracellular matrix receptors in mediating cell adhesion to extracellular matrix: preparation of monoclonal antibodies to the fibronectin receptor that specifically inhibit cell adhesion to fibronectin and react with platelet glycoproteins Ic-IIa. J Cell Biol. 1988 Nov;107(5):1881–1891. doi: 10.1083/jcb.107.5.1881. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Wilkins J. A., Chen K. Y., Lin S. Detection of high molecular weight vinculin binding proteins in muscle and nonmuscle tissues with an electroblot-overlay technique. Biochem Biophys Res Commun. 1983 Nov 15;116(3):1026–1032. doi: 10.1016/s0006-291x(83)80245-9. [DOI] [PubMed] [Google Scholar]
  42. Wilkins J. A., Risinger M. A., Lin S. Studies on proteins that co-purify with smooth muscle vinculin: identification of immunologically related species in focal adhesions of nonmuscle and Z-lines of muscle cells. J Cell Biol. 1986 Oct;103(4):1483–1494. doi: 10.1083/jcb.103.4.1483. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Zubrzycka-Gaarn E. E., Bulman D. E., Karpati G., Burghes A. H., Belfall B., Klamut H. J., Talbot J., Hodges R. S., Ray P. N., Worton R. G. The Duchenne muscular dystrophy gene product is localized in sarcolemma of human skeletal muscle. Nature. 1988 Jun 2;333(6172):466–469. doi: 10.1038/333466a0. [DOI] [PubMed] [Google Scholar]

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