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. 1985 Oct 1;101(4):1323–1331. doi: 10.1083/jcb.101.4.1323

beta-Internexin, a ubiquitous intermediate filament-associated protein

PMCID: PMC2113939  PMID: 3900089

Abstract

In this article we show a Triton-insoluble, intermediate filament- associated protein of approximately 70 kD to be expressed ubiquitously in diverse mammalian cell types. This protein, assigned the name beta- internexin, exhibits extreme homology in each of the various cell lines as demonstrated by identical limited peptide maps, similar mobilities on two-dimensional gels, and detection in Triton-soluble and -insoluble extracts. beta-Internexin also shares some degree of homology with alpha-internexin, an intermediate filament-associated protein isolated and purified from rat spinal cord, which accounts for the immunologic cross-reactivity displayed by these polypeptides. Light microscopic immunolocalization of beta-internexin with a monoclonal antibody (mAb- IN30) reveals it to be closely associated with the vimentin network in fibroblasts. The antigen is also observed to collapse with the vimentin reticulum during the formation of a juxtanuclear cap induced by colchicine treatment. Ultrastructural localization, using colloidal gold, substantiates the affinity of beta-internexin for cytoplasmic filaments and, in addition, demonstrates its apparent exclusion from the intranuclear filament network. We examine also the resemblance of beta-internexin to a microtubule-associated polypeptide and the constitutively synthesized mammalian heat shock protein (HSP 68/70).

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Selected References

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  1. Aamodt E. J., Williams R. C., Jr Microtubule-associated proteins connect microtubules and neurofilaments in vitro. Biochemistry. 1984 Dec 4;23(25):6023–6031. doi: 10.1021/bi00320a019. [DOI] [PubMed] [Google Scholar]
  2. Ben-Ze'ev A. Inhibition of vimentin synthesis and disruption of intermediate filaments in simian virus 40-infected monkey kidney cells. Mol Cell Biol. 1984 Sep;4(9):1880–1889. doi: 10.1128/mcb.4.9.1880. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bonner W. M., Laskey R. A. A film detection method for tritium-labelled proteins and nucleic acids in polyacrylamide gels. Eur J Biochem. 1974 Jul 1;46(1):83–88. doi: 10.1111/j.1432-1033.1974.tb03599.x. [DOI] [PubMed] [Google Scholar]
  4. Borisy G. G., Olmsted J. B., Marcum J. M., Allen C. Microtubule assembly in vitro. Fed Proc. 1974 Feb;33(2):167–174. [PubMed] [Google Scholar]
  5. Cleveland D. W., Fischer S. G., Kirschner M. W., Laemmli U. K. Peptide mapping by limited proteolysis in sodium dodecyl sulfate and analysis by gel electrophoresis. J Biol Chem. 1977 Feb 10;252(3):1102–1106. [PubMed] [Google Scholar]
  6. Dippold W. G., Lloyd K. O., Li L. T., Ikeda H., Oettgen H. F., Old L. J. Cell surface antigens of human malignant melanoma: definition of six antigenic systems with mouse monoclonal antibodies. Proc Natl Acad Sci U S A. 1980 Oct;77(10):6114–6118. doi: 10.1073/pnas.77.10.6114. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Elder J. H., Pickett R. A., 2nd, Hampton J., Lerner R. A. Radioiodination of proteins in single polyacrylamide gel slices. Tryptic peptide analysis of all the major members of complex multicomponent systems using microgram quantities of total protein. J Biol Chem. 1977 Sep 25;252(18):6510–6515. [PubMed] [Google Scholar]
  8. Faulk W. P., Taylor G. M. An immunocolloid method for the electron microscope. Immunochemistry. 1971 Nov;8(11):1081–1083. doi: 10.1016/0019-2791(71)90496-4. [DOI] [PubMed] [Google Scholar]
  9. GREENWOOD F. C., HUNTER W. M., GLOVER J. S. THE PREPARATION OF I-131-LABELLED HUMAN GROWTH HORMONE OF HIGH SPECIFIC RADIOACTIVITY. Biochem J. 1963 Oct;89:114–123. doi: 10.1042/bj0890114. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Geuze H. J., Slot J. W., van der Ley P. A., Scheffer R. C. Use of colloidal gold particles in double-labeling immunoelectron microscopy of ultrathin frozen tissue sections. J Cell Biol. 1981 Jun;89(3):653–665. doi: 10.1083/jcb.89.3.653. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Hirokawa N., Glicksman M. A., Willard M. B. Organization of mammalian neurofilament polypeptides within the neuronal cytoskeleton. J Cell Biol. 1984 Apr;98(4):1523–1536. doi: 10.1083/jcb.98.4.1523. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Hsu S. M., Raine L., Fanger H. Use of avidin-biotin-peroxidase complex (ABC) in immunoperoxidase techniques: a comparison between ABC and unlabeled antibody (PAP) procedures. J Histochem Cytochem. 1981 Apr;29(4):577–580. doi: 10.1177/29.4.6166661. [DOI] [PubMed] [Google Scholar]
  13. Hughes E. N., August J. T. Coprecipitation of heat shock proteins with a cell surface glycoprotein. Proc Natl Acad Sci U S A. 1982 Apr;79(7):2305–2309. doi: 10.1073/pnas.79.7.2305. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Köhler G., Milstein C. Derivation of specific antibody-producing tissue culture and tumor lines by cell fusion. Eur J Immunol. 1976 Jul;6(7):511–519. doi: 10.1002/eji.1830060713. [DOI] [PubMed] [Google Scholar]
  15. 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]
  16. Laskey R. A., Mills A. D. Quantitative film detection of 3H and 14C in polyacrylamide gels by fluorography. Eur J Biochem. 1975 Aug 15;56(2):335–341. doi: 10.1111/j.1432-1033.1975.tb02238.x. [DOI] [PubMed] [Google Scholar]
  17. Leterrier J. F., Liem R. K., Shelanski M. L. Interactions between neurofilaments and microtubule-associated proteins: a possible mechanism for intraorganellar bridging. J Cell Biol. 1982 Dec;95(3):982–986. doi: 10.1083/jcb.95.3.982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Liem R. K., Chin S. S., Moraru E., Wang E. Monoclonal antibodies to epitopes on different regions of the 200 000 dalton neurofilament protein. Probes for the geometry of the filament. Exp Cell Res. 1985 Feb;156(2):419–428. doi: 10.1016/0014-4827(85)90548-8. [DOI] [PubMed] [Google Scholar]
  19. O'Farrell P. H. High resolution two-dimensional electrophoresis of proteins. J Biol Chem. 1975 May 25;250(10):4007–4021. [PMC free article] [PubMed] [Google Scholar]
  20. Pachter J. S., Liem R. K. alpha-Internexin, a 66-kD intermediate filament-binding protein from mammalian central nervous tissues. J Cell Biol. 1985 Oct;101(4):1316–1322. doi: 10.1083/jcb.101.4.1316. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Shelanski M. L., Gaskin F., Cantor C. R. Microtubule assembly in the absence of added nucleotides. Proc Natl Acad Sci U S A. 1973 Mar;70(3):765–768. doi: 10.1073/pnas.70.3.765. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Sloboda R. D., Dentler W. L., Rosenbaum J. L. Microtubule-associated proteins and the stimulation of tubulin assembly in vitro. Biochemistry. 1976 Oct 5;15(20):4497–4505. doi: 10.1021/bi00665a026. [DOI] [PubMed] [Google Scholar]
  23. 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]
  24. Wang C., Gomer R. H., Lazarides E. Heat shock proteins are methylated in avian and mammalian cells. Proc Natl Acad Sci U S A. 1981 Jun;78(6):3531–3535. doi: 10.1073/pnas.78.6.3531. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Wang E., Cairncross J. G., Yung W. K., Garber E. A., Liem R. K. An intermediate filament-associated protein, p50, recognized by monoclonal antibodies. J Cell Biol. 1983 Nov;97(5 Pt 1):1507–1514. doi: 10.1083/jcb.97.5.1507. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Weatherbee J. A., Luftig R. B., Weihing R. R. Purification and reconstitution of HeLa cell microtubules. Biochemistry. 1980 Aug 19;19(17):4116–4123. doi: 10.1021/bi00558a033. [DOI] [PubMed] [Google Scholar]
  27. Weingarten M. D., Lockwood A. H., Hwo S. Y., Kirschner M. W. A protein factor essential for microtubule assembly. Proc Natl Acad Sci U S A. 1975 May;72(5):1858–1862. doi: 10.1073/pnas.72.5.1858. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Welch W. J., Feramisco J. R. Nuclear and nucleolar localization of the 72,000-dalton heat shock protein in heat-shocked mammalian cells. J Biol Chem. 1984 Apr 10;259(7):4501–4513. [PubMed] [Google Scholar]
  29. Welch W. J., Feramisco J. R. Purification of the major mammalian heat shock proteins. J Biol Chem. 1982 Dec 25;257(24):14949–14959. [PubMed] [Google Scholar]

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