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. 1990 Oct 1;271(1):179–183. doi: 10.1042/bj2710179

Distribution of polypeptides binding guanosine 5'-[gamma-[35S]thio]triphosphate and anti-(ras protein) antibodies in liver subcellular fractions. Evidence for endosome-specific components.

N Ali 1, W H Evans 1
PMCID: PMC1149530  PMID: 2121130

Abstract

The subcellular distribution in rat liver of polypeptides binding guanosine 5'-[gamma-[35S]thio]triphosphate [( 35S]GTP[S]) and seven antibodies against ras oncoproteins was evaluated. Multiple low-Mr (21,000-28,000) GTP-binding proteins were detected, but their relative distribution among the membrane fractions varied. A more specific compartmentation of polypeptides which bind antibodies generated against ras proteins was evident, with an Mr-28,000 polypeptide and a probable Mr-56,000 dimer, identified by six of the antibodies tested, being confined mainly to endosomes. An Mr-23,000 polypeptide was detected by some of the antibodies in all of the membrane fractions, but especially in the plasma membranes.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Ali N., Milligan G., Evans W. H. Distribution of G-proteins in rat liver plasma-membrane domains and endocytic pathways. Biochem J. 1989 Aug 1;261(3):905–912. doi: 10.1042/bj2610905. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Barbacid M. ras genes. Annu Rev Biochem. 1987;56:779–827. doi: 10.1146/annurev.bi.56.070187.004023. [DOI] [PubMed] [Google Scholar]
  3. Beckers C. J., Balch W. E. Calcium and GTP: essential components in vesicular trafficking between the endoplasmic reticulum and Golgi apparatus. J Cell Biol. 1989 Apr;108(4):1245–1256. doi: 10.1083/jcb.108.4.1245. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bergeron J. J. Golgi fractions from livers of control and ethanol-intoxicated rats. Enzymic and morphologic properties following rapid isolation. Biochim Biophys Acta. 1979 Aug 23;555(3):493–503. doi: 10.1016/0005-2736(79)90402-4. [DOI] [PubMed] [Google Scholar]
  5. Bhullar R. P., Haslam R. J. Detection of 23-27 kDa GTP-binding proteins in platelets and other cells. Biochem J. 1987 Jul 15;245(2):617–620. doi: 10.1042/bj2450617. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Bourne H. R. Do GTPases direct membrane traffic in secretion? Cell. 1988 Jun 3;53(5):669–671. doi: 10.1016/0092-8674(88)90081-5. [DOI] [PubMed] [Google Scholar]
  7. Burgoyne R. D., Morgan A. Low molecular mass GTP-binding proteins of adrenal chromaffin cells are present on the secretory granule. FEBS Lett. 1989 Mar 13;245(1-2):122–126. doi: 10.1016/0014-5793(89)80204-2. [DOI] [PubMed] [Google Scholar]
  8. 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]
  9. Buss J. E., Solski P. A., Schaeffer J. P., MacDonald M. J., Der C. J. Activation of the cellular proto-oncogene product p21Ras by addition of a myristylation signal. Science. 1989 Mar 24;243(4898):1600–1603. doi: 10.1126/science.2648572. [DOI] [PubMed] [Google Scholar]
  10. Comerford J. G., Dawson A. P. The effect of limited proteolysis on GTP-dependent Ca2+ efflux and GTP-dependent fusion in rat liver microsomal vesicles. Biochem J. 1989 Mar 15;258(3):823–829. doi: 10.1042/bj2580823. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Comerford J. G., Gibson J. R., Dawson A. P., Gibson I. ras p21 and other Gn proteins are detected in mammalian cell lines by [gamma-35S]GTP gamma S binding. Biochem Biophys Res Commun. 1989 Mar 31;159(3):1269–1274. doi: 10.1016/0006-291x(89)92247-x. [DOI] [PubMed] [Google Scholar]
  12. Cross S. L. More functions yet: no ras for the weary. The Role of Small GTP-Binding Proteins in Cellular Function, sponsored by the Oncogenes and Mitogens Program Project, University of Virginia Cancer Center, Charlottesville, VA, USA, November 2-3, 1989. New Biol. 1990 Feb;2(2):136–138. [PubMed] [Google Scholar]
  13. Damonte G., Sdraffa A., Zocchi E., Guida L., Polvani C., Tonetti M., Benatti U., Boquet P., De Flora A. Multiple small molecular weight guanine nucleotide-binding proteins in human erythrocyte membranes. Biochem Biophys Res Commun. 1990 Feb 14;166(3):1398–1405. doi: 10.1016/0006-291x(90)91022-k. [DOI] [PubMed] [Google Scholar]
  14. Evans W. H., Flint N. Subfractionation of hepatic endosomes in Nycodenz gradients and by free-flow electrophoresis. Separation of ligand-transporting and receptor-enriched membranes. Biochem J. 1985 Nov 15;232(1):25–32. doi: 10.1042/bj2320025. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Evans W. H. Preparation of low-density "endosome" and "endosome"-depleted Golgi fractions from rat liver. Methods Enzymol. 1985;109:246–257. doi: 10.1016/0076-6879(85)09090-5. [DOI] [PubMed] [Google Scholar]
  16. Gilman A. G. G proteins: transducers of receptor-generated signals. Annu Rev Biochem. 1987;56:615–649. doi: 10.1146/annurev.bi.56.070187.003151. [DOI] [PubMed] [Google Scholar]
  17. Goud B., Salminen A., Walworth N. C., Novick P. J. A GTP-binding protein required for secretion rapidly associates with secretory vesicles and the plasma membrane in yeast. Cell. 1988 Jun 3;53(5):753–768. doi: 10.1016/0092-8674(88)90093-1. [DOI] [PubMed] [Google Scholar]
  18. Gutierrez L., Magee A. I., Marshall C. J., Hancock J. F. Post-translational processing of p21ras is two-step and involves carboxyl-methylation and carboxy-terminal proteolysis. EMBO J. 1989 Apr;8(4):1093–1098. doi: 10.1002/j.1460-2075.1989.tb03478.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Haldar S., Beatty C., Tsujimoto Y., Croce C. M. The bcl-2 gene encodes a novel G protein. Nature. 1989 Nov 9;342(6246):195–198. doi: 10.1038/342195a0. [DOI] [PubMed] [Google Scholar]
  20. Kawata M., Kawahara Y., Araki S., Sunako M., Tsuda T., Fukuzaki H., Mizoguchi A., Takai Y. Identification of a major GTP-binding protein in bovine aortic smooth muscle membranes as smg p21, a GTP-binding protein having the same effector domain as ras p21s. Biochem Biophys Res Commun. 1989 Sep 29;163(3):1418–1427. doi: 10.1016/0006-291x(89)91137-6. [DOI] [PubMed] [Google Scholar]
  21. Kikuchi A., Sasaki T., Araki S., Hata Y., Takai Y. Purification and characterization from bovine brain cytosol of two GTPase-activating proteins specific for smg p21, a GTP-binding protein having the same effector domain as c-ras p21s. J Biol Chem. 1989 Jun 5;264(16):9133–9136. [PubMed] [Google Scholar]
  22. Klausner R. D. Sorting and traffic in the central vacuolar system. Cell. 1989 Jun 2;57(5):703–706. doi: 10.1016/0092-8674(89)90783-6. [DOI] [PubMed] [Google Scholar]
  23. 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]
  24. Lanoix J., Roy L., Paiement J. Detection of GTP-binding proteins in purified derivatives of rough endoplasmic reticulum. Biochem J. 1989 Sep 1;262(2):497–503. doi: 10.1042/bj2620497. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Lapetina E. G., Lacal J. C., Reep B. R., Molina y Vedia L. A ras-related protein is phosphorylated and translocated by agonists that increase cAMP levels in human platelets. Proc Natl Acad Sci U S A. 1989 May;86(9):3131–3134. doi: 10.1073/pnas.86.9.3131. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Mayorga L. S., Diaz R., Colombo M. I., Stahl P. D. GTP gamma S stimulation of endosome fusion suggests a role for a GTP-binding protein in the priming of vesicles before fusion. Cell Regul. 1989 Nov;1(1):113–124. doi: 10.1091/mbc.1.1.113. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Mayorga L. S., Diaz R., Stahl P. D. Regulatory role for GTP-binding proteins in endocytosis. Science. 1989 Jun 23;244(4911):1475–1477. doi: 10.1126/science.2499930. [DOI] [PubMed] [Google Scholar]
  28. Mizoguchi A., Kim S., Ueda T., Takai Y. Tissue distribution of smg p25A, a ras p21-like GTP-binding protein, studied by use of a specific monoclonal antibody. Biochem Biophys Res Commun. 1989 Aug 15;162(3):1438–1445. doi: 10.1016/0006-291x(89)90835-8. [DOI] [PubMed] [Google Scholar]
  29. Nigam S. K. Subcellular distribution of small GTP binding proteins in pancreas: identification of small GTP binding proteins in the rough endoplasmic reticulum. Proc Natl Acad Sci U S A. 1990 Feb;87(4):1296–1299. doi: 10.1073/pnas.87.4.1296. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Santos E., Nebreda A. R. Structural and functional properties of ras proteins. FASEB J. 1989 Aug;3(10):2151–2163. doi: 10.1096/fasebj.3.10.2666231. [DOI] [PubMed] [Google Scholar]
  31. Segev N., Mulholland J., Botstein D. The yeast GTP-binding YPT1 protein and a mammalian counterpart are associated with the secretion machinery. Cell. 1988 Mar 25;52(6):915–924. doi: 10.1016/0092-8674(88)90433-3. [DOI] [PubMed] [Google Scholar]
  32. Shears S. B., Evans W. H., Kirk C. J., Michell R. H. Preferential localization of rat liver D-myo-inositol 1,4,5-trisphosphate/1,3,4,5-tetrakisphosphate 5-phosphatase in bile-canalicular plasma membrane and 'late' endosomal vesicles. Biochem J. 1988 Dec 1;256(2):363–369. doi: 10.1042/bj2560363. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Sigal I. S., Gibbs J. B., D'Alonzo J. S., Scolnick E. M. Identification of effector residues and a neutralizing epitope of Ha-ras-encoded p21. Proc Natl Acad Sci U S A. 1986 Jul;83(13):4725–4729. doi: 10.1073/pnas.83.13.4725. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Toki C., Oda K., Ikehara Y. Demonstration of GTP-binding proteins and ADP-ribosylated proteins in rat liver Golgi fraction. Biochem Biophys Res Commun. 1989 Oct 16;164(1):333–338. doi: 10.1016/0006-291x(89)91722-1. [DOI] [PubMed] [Google Scholar]
  35. Urumow T., Wieland O. H. A small G-protein involved in phosphatidylinositol-4-phosphate kinase activation. FEBS Lett. 1990 Apr 9;263(1):15–17. doi: 10.1016/0014-5793(90)80694-e. [DOI] [PubMed] [Google Scholar]
  36. Wisher M. H., Evans W. H. Functional polarity of the rat hepatocyte surface membrane. Isolation and characterization of plasma-membrane subfractions from the blood-sinusoidal, bile-Canalicular and contiguous surfaces of the hepatocyte. Biochem J. 1975 Feb;146(2):375–388. doi: 10.1042/bj1460375. [DOI] [PMC free article] [PubMed] [Google Scholar]

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