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. 1984 Dec 1;99(6):2254–2259. doi: 10.1083/jcb.99.6.2254

Segregation of the polypeptide translocation apparatus to regions of the endoplasmic reticulum containing ribophorins and ribosomes. II. Rat liver microsomal subfractions contain equimolar amounts of ribophorins and ribosomes

PMCID: PMC2113550  PMID: 6501424

Abstract

Ribophorins I and II, two transmembrane glycoproteins characteristic of the rough endoplasmic reticulum (ER) are thought to be part of the translocation apparatus for proteins made on membrane bound polysomes. To study the stoichiometry between ribophorins and membrane-bound ribosomes we have determined the RNA and ribophorin content in rat liver microsomes or in microsomal subfractions of different density (i.e., ribosome content). The specificity of antibodies against the ribophorins was demonstrated by Western blot analysis of rat liver rough microsomes separated by 2-dimensional gel electrophoresis. The ribophorin content of microsomal subfractions was determined by indirect immunoprecipitation and for ribophorin I by a radioimmune assay. In the latter assay a molar ratio of ribophorin I/ribosomes approaching one was calculated for total microsomes as well as in the gradient subfractions. We therefore suggest that ribophorins mediate the binding of ribosomes to endoplasmic reticulum membranes or play a role in co-translational process which depend on this binding, such as the insertion of nascent polypeptides into the membrane or their transfer into the cisternal lumen.

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Selected References

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  1. Adelman M. R., Sabatini D. D., Blobel G. Ribosome-membrane interaction. Nondestructive disassembly of rat liver rough microsomes into ribosomal and membranous components. J Cell Biol. 1973 Jan;56(1):206–229. doi: 10.1083/jcb.56.1.206. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Amar-Costesec A., Todd J. A., Kreibich G. Segregation of the polypeptide translocation apparatus to regions of the endoplasmic reticulum containing ribophorins and ribosomes. I. Functional tests on rat liver microsomal subfractions. J Cell Biol. 1984 Dec;99(6):2247–2253. doi: 10.1083/jcb.99.6.2247. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bolton A. E., Hunter W. M. The labelling of proteins to high specific radioactivities by conjugation to a 125I-containing acylating agent. Biochem J. 1973 Jul;133(3):529–539. doi: 10.1042/bj1330529. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Borgese N., Mok W., Kreibich G., Sabatini D. D. Ribosomal-membrane interaction: in vitro binding of ribosomes to microsomal membranes. J Mol Biol. 1974 Sep 25;88(3):559–580. doi: 10.1016/0022-2836(74)90408-2. [DOI] [PubMed] [Google Scholar]
  5. Burnette W. N. "Western blotting": electrophoretic transfer of proteins from sodium dodecyl sulfate--polyacrylamide gels to unmodified nitrocellulose and radiographic detection with antibody and radioiodinated protein A. Anal Biochem. 1981 Apr;112(2):195–203. doi: 10.1016/0003-2697(81)90281-5. [DOI] [PubMed] [Google Scholar]
  6. Fisher P. A., Berrios M., Blobel G. Isolation and characterization of a proteinaceous subnuclear fraction composed of nuclear matrix, peripheral lamina, and nuclear pore complexes from embryos of Drosophila melanogaster. J Cell Biol. 1982 Mar;92(3):674–686. doi: 10.1083/jcb.92.3.674. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Gilmore R., Walter P., Blobel G. Protein translocation across the endoplasmic reticulum. II. Isolation and characterization of the signal recognition particle receptor. J Cell Biol. 1982 Nov;95(2 Pt 1):470–477. doi: 10.1083/jcb.95.2.470. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Kreibich G., Czakó-Graham M., Grebenau R., Mok W., Rodriguez-Boulan E., Sabatini D. D. Characterization of the ribosomal binding site in rat liver rough microsomes: ribophorins I and II, two integral membrane proteins related to ribosome binding. J Supramol Struct. 1978;8(3):279–302. doi: 10.1002/jss.400080307. [DOI] [PubMed] [Google Scholar]
  9. Kreibich G., Freienstein C. M., Pereyra B. N., Ulrich B. L., Sabatini D. D. Proteins of rough microsomal membranes related to ribosome binding. II. Cross-linking of bound ribosomes to specific membrane proteins exposed at the binding sites. J Cell Biol. 1978 May;77(2):488–506. doi: 10.1083/jcb.77.2.488. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Kreibich G., Marcantonio E. E., Sabatini D. D. Ribophorins I and II: membrane proteins characteristic of the rough endoplasmic reticulum. Methods Enzymol. 1983;96:520–530. doi: 10.1016/s0076-6879(83)96045-7. [DOI] [PubMed] [Google Scholar]
  11. Kreibich G., Sabatini D. D. Selective release of content from microsomal vesicles without membrane disassembly. II. Electrophoretic and immunological characterization of microsomal subfractions. J Cell Biol. 1974 Jun;61(3):789–807. doi: 10.1083/jcb.61.3.789. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Kreibich G., Ulrich B. L., Sabatini D. D. Proteins of rough microsomal membranes related to ribosome binding. I. Identification of ribophorins I and II, membrane proteins characteristics of rough microsomes. J Cell Biol. 1978 May;77(2):464–487. doi: 10.1083/jcb.77.2.464. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  14. Lazarides E. Two general classes of cytoplasmic actin filaments in tissue culture cells: the role of tropomyosin. J Supramol Struct. 1976;5(4):531(383)–563(415). doi: 10.1002/jss.400050410. [DOI] [PubMed] [Google Scholar]
  15. Marcantonio E. E., Grebenau R. C., Sabatini D. D., Kreibich G. Identification of ribophorins in rough microsomal membranes from different organs of several species. Eur J Biochem. 1982 May;124(1):217–222. doi: 10.1111/j.1432-1033.1982.tb05928.x. [DOI] [PubMed] [Google Scholar]
  16. Meyer D. I., Dobberstein B. A membrane component essential for vectorial translocation of nascent proteins across the endoplasmic reticulum: requirements for its extraction and reassociation with the membrane. J Cell Biol. 1980 Nov;87(2 Pt 1):498–502. doi: 10.1083/jcb.87.2.498. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Meyer D. I., Dobberstein B. Identification and characterization of a membrane component essential for the translocation of nascent proteins across the membrane of the endoplasmic reticulum. J Cell Biol. 1980 Nov;87(2 Pt 1):503–508. doi: 10.1083/jcb.87.2.503. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Meyer D. I., Krause E., Dobberstein B. Secretory protein translocation across membranes-the role of the "docking protein'. Nature. 1982 Jun 24;297(5868):647–650. doi: 10.1038/297647a0. [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. Rodriguez Boulan E., Sabatini D. D., Pereyra B. N., Kreibich G. Spatial orientation of glycoproteins in membranes of rat liver rough microsomes. II. Transmembrane disposition and characterization of glycoproteins. J Cell Biol. 1978 Sep;78(3):894–909. doi: 10.1083/jcb.78.3.894. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Sharma R. N., Behar-Bennelier M., Rolleston F. S., Murray R. K. Electrophoretic studies on liver endoplasmic reticulum membrane polypeptides and on their phosphorylation in vivo and in vitro. J Biol Chem. 1978 Mar 25;253(6):2033–2043. [PubMed] [Google Scholar]
  22. Vlasuk G. P., Walz F. G., Jr Liver endoplasmic reticulum polypeptides resolved by two-dimensional gel electrophoresis. Anal Biochem. 1980 Jun;105(1):112–120. doi: 10.1016/0003-2697(80)90431-5. [DOI] [PubMed] [Google Scholar]
  23. Walter P., Blobel G. Purification of a membrane-associated protein complex required for protein translocation across the endoplasmic reticulum. Proc Natl Acad Sci U S A. 1980 Dec;77(12):7112–7116. doi: 10.1073/pnas.77.12.7112. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Walter P., Blobel G. Translocation of proteins across the endoplasmic reticulum III. Signal recognition protein (SRP) causes signal sequence-dependent and site-specific arrest of chain elongation that is released by microsomal membranes. J Cell Biol. 1981 Nov;91(2 Pt 1):557–561. doi: 10.1083/jcb.91.2.557. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Walter P., Blobel G. Translocation of proteins across the endoplasmic reticulum. II. Signal recognition protein (SRP) mediates the selective binding to microsomal membranes of in-vitro-assembled polysomes synthesizing secretory protein. J Cell Biol. 1981 Nov;91(2 Pt 1):551–556. doi: 10.1083/jcb.91.2.551. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Walter P., Ibrahimi I., Blobel G. Translocation of proteins across the endoplasmic reticulum. I. Signal recognition protein (SRP) binds to in-vitro-assembled polysomes synthesizing secretory protein. J Cell Biol. 1981 Nov;91(2 Pt 1):545–550. doi: 10.1083/jcb.91.2.545. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Warren G., Dobberstein B. Protein transfer across microsomal membranes reassembled from separated membrane components. Nature. 1978 Jun 15;273(5663):569–571. doi: 10.1038/273569a0. [DOI] [PubMed] [Google Scholar]
  28. Wray W., Boulikas T., Wray V. P., Hancock R. Silver staining of proteins in polyacrylamide gels. Anal Biochem. 1981 Nov 15;118(1):197–203. doi: 10.1016/0003-2697(81)90179-2. [DOI] [PubMed] [Google Scholar]

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