Abstract
A point mutation in a highly conserved region of the beta 1 subunit, Asp130 to Ala (D130A) substitution, abrogates the Arg-Gly-Asp (RGD)- dependent binding of alpha 5 beta 1 to fibronectin (FN) without disrupting gross structure or heterodimer assembly. The D130A mutation also interferes with binding to invasin, a ligand that lacks RGD sequence. In spite of the lack of detectable FN binding by alpha 5 beta 1(D130A), it was recruited to adhesion plaques formed on FN by endogenous hamster receptors. Thus, intact ligand binding function is not required for recruitment of alpha 5 beta 1 to adhesion plaques. Overexpression of beta 1(D130A) partially interfered with endogenous alpha 5 beta 1 function, thus defining a dominant negative beta 1 integrin mutation.
Full Text
The Full Text of this article is available as a PDF (2.7 MB).
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Akiyama S. K., Yamada S. S., Chen W. T., Yamada K. M. Analysis of fibronectin receptor function with monoclonal antibodies: roles in cell adhesion, migration, matrix assembly, and cytoskeletal organization. J Cell Biol. 1989 Aug;109(2):863–875. doi: 10.1083/jcb.109.2.863. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Amaya E., Musci T. J., Kirschner M. W. Expression of a dominant negative mutant of the FGF receptor disrupts mesoderm formation in Xenopus embryos. Cell. 1991 Jul 26;66(2):257–270. doi: 10.1016/0092-8674(91)90616-7. [DOI] [PubMed] [Google Scholar]
- 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]
- Aruffo A., Seed B. Molecular cloning of a CD28 cDNA by a high-efficiency COS cell expression system. Proc Natl Acad Sci U S A. 1987 Dec;84(23):8573–8577. doi: 10.1073/pnas.84.23.8573. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Braciale T. J., Henkel T. J., Lukacher A., Braciale V. L. Fine specificity and antigen receptor expression among influenza virus-specific cytolytic T lymphocyte clones. J Immunol. 1986 Aug 1;137(3):995–1002. [PubMed] [Google Scholar]
- Brown P. J., Juliano R. L. Monoclonal antibodies to distinctive epitopes on the alpha and beta subunits of the fibronectin receptor. Exp Cell Res. 1988 Aug;177(2):303–318. doi: 10.1016/0014-4827(88)90464-8. [DOI] [PubMed] [Google Scholar]
- Brown P. J., Juliano R. L. Selective inhibition of fibronectin-mediated cell adhesion by monoclonal antibodies to a cell-surface glycoprotein. Science. 1985 Jun 21;228(4706):1448–1451. doi: 10.1126/science.4012302. [DOI] [PubMed] [Google Scholar]
- Carter W. G., Ryan M. C., Gahr P. J. Epiligrin, a new cell adhesion ligand for integrin alpha 3 beta 1 in epithelial basement membranes. Cell. 1991 May 17;65(4):599–610. doi: 10.1016/0092-8674(91)90092-d. [DOI] [PubMed] [Google Scholar]
- Cheresh D. A., Smith J. W., Cooper H. M., Quaranta V. A novel vitronectin receptor integrin (alpha v beta x) is responsible for distinct adhesive properties of carcinoma cells. Cell. 1989 Apr 7;57(1):59–69. doi: 10.1016/0092-8674(89)90172-4. [DOI] [PubMed] [Google Scholar]
- D'Souza S. E., Ginsberg M. H., Burke T. A., Lam S. C., Plow E. F. Localization of an Arg-Gly-Asp recognition site within an integrin adhesion receptor. Science. 1988 Oct 7;242(4875):91–93. doi: 10.1126/science.3262922. [DOI] [PubMed] [Google Scholar]
- Dejana E., Colella S., Conforti G., Abbadini M., Gaboli M., Marchisio P. C. Fibronectin and vitronectin regulate the organization of their respective Arg-Gly-Asp adhesion receptors in cultured human endothelial cells. J Cell Biol. 1988 Sep;107(3):1215–1223. doi: 10.1083/jcb.107.3.1215. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Elices M. J., Osborn L., Takada Y., Crouse C., Luhowskyj S., Hemler M. E., Lobb R. R. VCAM-1 on activated endothelium interacts with the leukocyte integrin VLA-4 at a site distinct from the VLA-4/fibronectin binding site. Cell. 1990 Feb 23;60(4):577–584. doi: 10.1016/0092-8674(90)90661-w. [DOI] [PubMed] [Google Scholar]
- Engvall E., Ruoslahti E. Binding of soluble form of fibroblast surface protein, fibronectin, to collagen. Int J Cancer. 1977 Jul 15;20(1):1–5. doi: 10.1002/ijc.2910200102. [DOI] [PubMed] [Google Scholar]
- Giancotti F. G., Ruoslahti E. Elevated levels of the alpha 5 beta 1 fibronectin receptor suppress the transformed phenotype of Chinese hamster ovary cells. Cell. 1990 Mar 9;60(5):849–859. doi: 10.1016/0092-8674(90)90098-y. [DOI] [PubMed] [Google Scholar]
- Hemler M. E., Huang C., Takada Y., Schwarz L., Strominger J. L., Clabby M. L. Characterization of the cell surface heterodimer VLA-4 and related peptides. J Biol Chem. 1987 Aug 25;262(24):11478–11485. [PubMed] [Google Scholar]
- Holzmann B., McIntyre B. W., Weissman I. L. Identification of a murine Peyer's patch--specific lymphocyte homing receptor as an integrin molecule with an alpha chain homologous to human VLA-4 alpha. Cell. 1989 Jan 13;56(1):37–46. doi: 10.1016/0092-8674(89)90981-1. [DOI] [PubMed] [Google Scholar]
- Isberg R. R. Discrimination between intracellular uptake and surface adhesion of bacterial pathogens. Science. 1991 May 17;252(5008):934–938. doi: 10.1126/science.1674624. [DOI] [PubMed] [Google Scholar]
- Isberg R. R., Leong J. M. Multiple beta 1 chain integrins are receptors for invasin, a protein that promotes bacterial penetration into mammalian cells. Cell. 1990 Mar 9;60(5):861–871. doi: 10.1016/0092-8674(90)90099-z. [DOI] [PubMed] [Google Scholar]
- Kaufman R. J., Sharp P. A. Amplification and expression of sequences cotransfected with a modular dihydrofolate reductase complementary dna gene. J Mol Biol. 1982 Aug 25;159(4):601–621. doi: 10.1016/0022-2836(82)90103-6. [DOI] [PubMed] [Google Scholar]
- Kunkel T. A., Roberts J. D., Zakour R. A. Rapid and efficient site-specific mutagenesis without phenotypic selection. Methods Enzymol. 1987;154:367–382. doi: 10.1016/0076-6879(87)54085-x. [DOI] [PubMed] [Google Scholar]
- LaFlamme S. E., Akiyama S. K., Yamada K. M. Regulation of fibronectin receptor distribution. J Cell Biol. 1992 Apr;117(2):437–447. doi: 10.1083/jcb.117.2.437. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 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]
- Leong J. M., Fournier R. S., Isberg R. R. Identification of the integrin binding domain of the Yersinia pseudotuberculosis invasin protein. EMBO J. 1990 Jun;9(6):1979–1989. doi: 10.1002/j.1460-2075.1990.tb08326.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Loftus J. C., O'Toole T. E., Plow E. F., Glass A., Frelinger A. L., 3rd, Ginsberg M. H. A beta 3 integrin mutation abolishes ligand binding and alters divalent cation-dependent conformation. Science. 1990 Aug 24;249(4971):915–918. doi: 10.1126/science.2392682. [DOI] [PubMed] [Google Scholar]
- MacKrell A. J., Blumberg B., Haynes S. R., Fessler J. H. The lethal myospheroid gene of Drosophila encodes a membrane protein homologous to vertebrate integrin beta subunits. Proc Natl Acad Sci U S A. 1988 Apr;85(8):2633–2637. doi: 10.1073/pnas.85.8.2633. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Obara M., Kang M. S., Yamada K. M. Site-directed mutagenesis of the cell-binding domain of human fibronectin: separable, synergistic sites mediate adhesive function. Cell. 1988 May 20;53(4):649–657. doi: 10.1016/0092-8674(88)90580-6. [DOI] [PubMed] [Google Scholar]
- Pierschbacher M. D., Ruoslahti E., Sundelin J., Lind P., Peterson P. A. The cell attachment domain of fibronectin. Determination of the primary structure. J Biol Chem. 1982 Aug 25;257(16):9593–9597. [PubMed] [Google Scholar]
- Plow E. F., Birdwell C., Ginsberg M. H. Identification and quantitation of platelet-associated fibronectin antigen. J Clin Invest. 1979 Mar;63(3):540–543. doi: 10.1172/JCI109334. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Pytela R., Pierschbacher M. D., Ruoslahti E. Identification and isolation of a 140 kd cell surface glycoprotein with properties expected of a fibronectin receptor. Cell. 1985 Jan;40(1):191–198. doi: 10.1016/0092-8674(85)90322-8. [DOI] [PubMed] [Google Scholar]
- Schreiner C. L., Bauer J. S., Danilov Y. N., Hussein S., Sczekan M. M., Juliano R. L. Isolation and characterization of Chinese hamster ovary cell variants deficient in the expression of fibronectin receptor. J Cell Biol. 1989 Dec;109(6 Pt 1):3157–3167. doi: 10.1083/jcb.109.6.3157. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Smith J. W., Cheresh D. A. The Arg-Gly-Asp binding domain of the vitronectin receptor. Photoaffinity cross-linking implicates amino acid residues 61-203 of the beta subunit. J Biol Chem. 1988 Dec 15;263(35):18726–18731. [PubMed] [Google Scholar]
- Solowska J., Guan J. L., Marcantonio E. E., Trevithick J. E., Buck C. A., Hynes R. O. Expression of normal and mutant avian integrin subunits in rodent cells. J Cell Biol. 1989 Aug;109(2):853–861. doi: 10.1083/jcb.109.2.853. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Subramani S., Mulligan R., Berg P. Expression of the mouse dihydrofolate reductase complementary deoxyribonucleic acid in simian virus 40 vectors. Mol Cell Biol. 1981 Sep;1(9):854–864. doi: 10.1128/mcb.1.9.854. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Takada Y., Elices M. J., Crouse C., Hemler M. E. The primary structure of the alpha 4 subunit of VLA-4: homology to other integrins and a possible cell-cell adhesion function. EMBO J. 1989 May;8(5):1361–1368. doi: 10.1002/j.1460-2075.1989.tb03516.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Takada Y., Hemler M. E. The primary structure of the VLA-2/collagen receptor alpha 2 subunit (platelet GPIa): homology to other integrins and the presence of a possible collagen-binding domain. J Cell Biol. 1989 Jul;109(1):397–407. doi: 10.1083/jcb.109.1.397. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Takada Y., Murphy E., Pil P., Chen C., Ginsberg M. H., Hemler M. E. Molecular cloning and expression of the cDNA for alpha 3 subunit of human alpha 3 beta 1 (VLA-3), an integrin receptor for fibronectin, laminin, and collagen. J Cell Biol. 1991 Oct;115(1):257–266. doi: 10.1083/jcb.115.1.257. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Takada Y., Strominger J. L., Hemler M. E. The very late antigen family of heterodimers is part of a superfamily of molecules involved in adhesion and embryogenesis. Proc Natl Acad Sci U S A. 1987 May;84(10):3239–3243. doi: 10.1073/pnas.84.10.3239. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Ueno H., Colbert H., Escobedo J. A., Williams L. T. Inhibition of PDGF beta receptor signal transduction by coexpression of a truncated receptor. Science. 1991 May 10;252(5007):844–848. doi: 10.1126/science.1851331. [DOI] [PubMed] [Google Scholar]
- Urlaub G., Chasin L. A. Isolation of Chinese hamster cell mutants deficient in dihydrofolate reductase activity. Proc Natl Acad Sci U S A. 1980 Jul;77(7):4216–4220. doi: 10.1073/pnas.77.7.4216. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Van Nhieu G. T., Isberg R. R. The Yersinia pseudotuberculosis invasin protein and human fibronectin bind to mutually exclusive sites on the alpha 5 beta 1 integrin receptor. J Biol Chem. 1991 Dec 25;266(36):24367–24375. [PubMed] [Google Scholar]
- Ylänne J., Virtanen I. The Mr 140,000 fibronectin receptor complex in normal and virus-transformed human fibroblasts and in fibrosarcoma cells: identical localization and function. Int J Cancer. 1989 Jun 15;43(6):1126–1136. doi: 10.1002/ijc.2910430628. [DOI] [PubMed] [Google Scholar]
- Zoller M. J., Smith M. Oligonucleotide-directed mutagenesis using M13-derived vectors: an efficient and general procedure for the production of point mutations in any fragment of DNA. Nucleic Acids Res. 1982 Oct 25;10(20):6487–6500. doi: 10.1093/nar/10.20.6487. [DOI] [PMC free article] [PubMed] [Google Scholar]