Skip to main content
Elsevier - PMC COVID-19 Collection logoLink to Elsevier - PMC COVID-19 Collection
. 2004 Apr 28;36(4):847–856. doi: 10.1016/0092-8674(84)90034-5

Microinjection of mRNA coding for an anti-golgi antibody inhibits intracellular transport of a viral membrane protein

Brian Burke 1,, Graham Warren 1
PMCID: PMC7133283  PMID: 6323023

Abstract

Messenger RNA was prepared from a hybridoma cell line secreting a monoclonal antibody (53FC3) directed against a luminal epitope of a Golgi membrane protein (Mr = 135 kd) found in rodent cells. When this mRNA was microinjected into the cytoplasm of BHK cells, mouse IgG was seen to accumulate in the Golgi complex after 5–6 hr of incubation. No accumulation was seen in 3T3 cells which lack the epitope recognized by 53FC3. When microinjected BHK cells were infected with vesicular stomatitis virus, surface expression of the viral G protein was considerably reduced when compared with neighboring noninjected cells.

References

  1. Ansorge W. Improved system for capillary microinjection into living cells. Exp. Cell Res. 1982;140:31–37. doi: 10.1016/0014-4827(82)90152-5. [DOI] [PubMed] [Google Scholar]
  2. Antman K.H., Livingston D.M. Intracellular neutralization of SV40 tumor antigens following microinjection of specific antibody. Cell. 1980;19:627–635. doi: 10.1016/s0092-8674(80)80039-0. [DOI] [PubMed] [Google Scholar]
  3. Ash J.F., Louvard D., Singer S.J. Vol. 74. 1977. Antibody induced linkages of plasma membrane proteins to intracellular actomycin-containing filaments in cultured fibroblasts; pp. 5584–5588. (Proc. Nat. Acad. Sci. USA). [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bergeron J.J.M. Golgi fractions from livers of control and ethanol intoxicated rats. Biochim. Biophys. Acta. 1979;555:493–503. doi: 10.1016/0005-2736(79)90402-4. [DOI] [PubMed] [Google Scholar]
  5. Bergman J.E., Tokuyasu K.T., Singer S.J. Vol. 78. 1981. Passage of an integral membrane protein, the vesicular stomatitis virus glycoprotein, through the Golgi apparatus en route to the plasma membrane; pp. 1746–1750. (Proc. Nat. Acad. Sci. USA). [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Blobel G., Dobberstein B. Transfer of proteins across membranes. I. Presence of proteolytically processed and unprocessed nascent immunoglobulin light chains on membrane bound ribosomes of murine myeloma. J. Cell Biol. 1977;67:835–851. doi: 10.1083/jcb.67.3.835. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Brandtzaeg P. Conjugates of immunoglobulin G with different fluorochromes. I. Characterization of anionic exchange chromatography. Scand. J. Immunol. 1973;2:273–290. doi: 10.1111/j.1365-3083.1973.tb02037.x. [DOI] [PubMed] [Google Scholar]
  8. Bretz R., Stäubli W. Detergent influence on rat liver galactosyl transferase activities towards different acceptors. Eur. J. Biochem. 1977;77:181–192. doi: 10.1111/j.1432-1033.1977.tb11656.x. [DOI] [PubMed] [Google Scholar]
  9. Bretz R., Bretz H., Palade G. Distribution of terminal glycosyl transferases in hepatic Golgi fractions. J. Cell Biol. 1980;84:87–101. doi: 10.1083/jcb.84.1.87. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Burke B., Griffiths G., Reggio H., Louvard D., Warren G. A monoclonal antibody against a 135K Golgi membrane protein. EMBO J. 1982;1:1621–1628. doi: 10.1002/j.1460-2075.1982.tb01364.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Cleveland D.W., Pittinger M.F., Feramisco J.R. Elevation of tubulin levels by microinjection suppresses new tubulin synthesis. Nature. 1983;305:738–740. doi: 10.1038/305738a0. [DOI] [PubMed] [Google Scholar]
  12. Colman A., Besley J., Valle G. Interactions of mouse immunoglobulin chains within Xenopus oocytes. J. Mol. Biol. 1982;160:459–474. doi: 10.1016/0022-2836(82)90308-4. [DOI] [PubMed] [Google Scholar]
  13. Fleischer B. Orientation of glycoprotein galactosyltransferase and sialyltransferase enzymes in vesicles derived from rat liver Golgi apparatus. J. Cell Biol. 1981;89:246–255. doi: 10.1083/jcb.89.2.246. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Glenney J.R., Glenney P., Weber K. Vol. 79. 1982. Erythroid spectrin, brain fodrin, and intestinal brush border proteins (TW-260/240) are related molecules containing a common calmodulin-binding subunit bound to a variant cell type-specific subunit; pp. 4002–4005. (Proc. Nat. Acad. Sci.). [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Griffiths G., Quinn P., Warren G. Dissection of the Golgi complex. I. Monensin inhibits the transport of viral membrane proteins from medial to trans Golgi cisternae in baby hamster kidney cells infected with Semliki Forest virus. J. Cell Biol. 1983;96:835–850. doi: 10.1083/jcb.96.3.835. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hasilik A., Neufeld E. Biosynthesis of lysosomal enzymes in fibroblasts: phosphorylation of mannose residues. J. Biol. Chem. 1980;255:4946–4950. [PubMed] [Google Scholar]
  17. Hiller G., Weber K. Golgi detection in mitotic and interphase cells by antibodies to secreted galactosyl transferase. Exp. Cell Res. 1982;142:85–94. doi: 10.1016/0014-4827(82)90412-8. [DOI] [PubMed] [Google Scholar]
  18. Howe J.G., Hershey W.B. A sensitive immunoblotting method for measuring protein synthesis initiation factor levels in lysate of E. coli. J. Biol. Chem. 1981;256:12836–12839. [PubMed] [Google Scholar]
  19. Hubbard S.C., Ivatt R.J. Synthesis and processing of asparagine-linked oligosaccharides. Ann. Rev. Biochem. 1981;50:55–83. doi: 10.1146/annurev.bi.50.070181.003011. [DOI] [PubMed] [Google Scholar]
  20. Jamieson Z.D., Palade G. Intracellular transport of secretory proteins in the pancreatic exocrine cell. III. Dissociation of intracellular transport from protein synthesis. J. Cell Biol. 1968;39:580–588. doi: 10.1083/jcb.39.3.580. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Lin J.J.-C., Feramisco J.R. Disruption of the in vivo distribution of the intermediate filaments in living fibroblasts through the microinjection of a specific monoclonal antibody. Cell. 1981;24:185–193. doi: 10.1016/0092-8674(81)90514-6. [DOI] [PubMed] [Google Scholar]
  22. Louvard D., Reggio H., Warren G. Antibodies to the Golgi complex and the rough endoplasmic reticulum. J. Cell Biol. 1982;92:92–107. doi: 10.1083/jcb.92.1.92. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Mabuchi I., Okuno M. The effects of myosin antibody on the division of starfish blastomeres. J. Cell Biol. 1977;74:251–264. doi: 10.1083/jcb.74.1.251. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. 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;87:498–502. doi: 10.1083/jcb.87.2.498. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Novikoff P.M., Tulsiani D.R.P., Touster O., Yam A., Novikoff A.B. Vol. 80. 1983. Immunocytochemical localisation of c<-D-Mannosidase II in the Golgi apparatus of rat liver; pp. 4364–4368. (Proc. Nat. Acad. Sci. USA). [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Palade G. Intracellular aspects of the process of protein synthesis. Science. 1975;189:347–352. doi: 10.1126/science.1096303. [DOI] [PubMed] [Google Scholar]
  27. Quinn P., Griffiths G., Warren G. Dissection of the Golgi complex. II. Density separation of specific Golgi functions in virally infected cells treated with monensin. J. Cell Biol. 1983;96:851–856. doi: 10.1083/jcb.96.3.851. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Reitman M.L., Kornfeld S. Lysosomal enzyme targeting: Nacetylglucosaminylphosphotransferase selectively phosphorylates native lysosomal enzymes. J. Biol. Chem. 1981;256:11977–11980. [PubMed] [Google Scholar]
  29. Roth J., Berger E.G. Immunocytochemical localisation of galactosyl transferase in HeLa cells: codistribution with thiamine pyrophosphatase in trans Golgi cisternae. J. Cell Biol. 1982;92:223–229. doi: 10.1083/jcb.93.1.223. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Rothman J.E. The Golgi apparatus: two organelles in tandem. Science. 1981;213:1212–1219. doi: 10.1126/science.7268428. [DOI] [PubMed] [Google Scholar]
  31. Schmidt M.F.G., Bracha M., Schlessinger M.J. Vol. 76. 1979. Evidence for covalent attachment of fatty acids to Sindbis virus glycoproteins; pp. 1687–1691. (Proc. Nat. Acad. Sci. USA). [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Silverstein S.C., Steinman R.M., Cohn Z.A. Endocytosis. Ann. Rev. Biochem. 1977;46:669–722. doi: 10.1146/annurev.bi.46.070177.003321. [DOI] [PubMed] [Google Scholar]
  33. Tartakoff A., Vassalli P. Plasma cell immunoglobulin secretion: arrest is accompanied by alterations in the Golgi complex. J. Exp. Med. 1977;146:1332–1345. doi: 10.1084/jem.146.5.1332. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Tartakoff A., Vassalli P. Comparative study of intracellular transport of secretory proteins. J. Cell Biol. 1978;79:694–707. doi: 10.1083/jcb.79.3.694. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Temynck T., Avrameas S. Polymerisation and immobilization of proteins using ethylchloroformate and glutaraldehyde. Scand. J. Immunol. Suppl. 1976;3:29–35. [Google Scholar]
  36. Timm B., Kondor-Koch C., Lehrach H., Riedel H., Edstrom J.-E., Garoff H. Expression of viral membrane proteins from cloned cDNA by microinjection into eukaryotic cell nuclei. Meth. Enzymol. 1983;96:496–511. doi: 10.1016/s0076-6879(83)96043-3. [DOI] [PubMed] [Google Scholar]
  37. Valle G., Jones E.A., Colman A. Anti-ovalbumin monoclonal antibodies interact with their antigen in internal membranes of Xenopus oocytes. Nature. 1982;300:71–74. doi: 10.1038/300071a0. [DOI] [PubMed] [Google Scholar]
  38. von Figura K., Klein U. Isolation and characterisation of phosphorylated oligosaccharides from c<-N-acetylglucosaminidase that are recognised by cell-surface receptors. Eur. J. Biochem. 1979;94:347–354. doi: 10.1111/j.1432-1033.1979.tb12900.x. [DOI] [PubMed] [Google Scholar]
  39. Warren G., Dobberstein B. Protein transfer across microsomal membranes reassembled from separated membrane components. Nature. 1978;273:569–571. doi: 10.1038/273569a0. [DOI] [PubMed] [Google Scholar]
  40. Warren G., Featherstone C., Griffiths G., Burke B. Newly synthesized G protein of vesicular stomatitis virus is not transported to the cell surface during mitosis. J. Cell Biol. 1983;97:1623–1628. doi: 10.1083/jcb.97.5.1623. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Yamaizumi M., Uchida T., Okada Y., Furusawa M. Neutralization of diphtheria toxin in living cells by microinjection of antifragment A contained within resealed erythrocyte ghosts. Cell. 1978;13:227–232. doi: 10.1016/0092-8674(78)90191-5. [DOI] [PubMed] [Google Scholar]

Articles from Cell are provided here courtesy of Elsevier

RESOURCES