Skip to main content
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1976 May;73(5):1631–1635. doi: 10.1073/pnas.73.5.1631

Sodium-stimulated amino acid uptake into isolated membrane vesicles from Balb/c 3T3 cells transformed by simian virus 40.

D C Quinlan, J R Parnes, R Shalom, T Q Garvey 3rd, K J Isselbacher, J Hochstadt
PMCID: PMC430353  PMID: 179092

Abstract

Mediated uptake of amino acids by membrane vesicles isolated from Balb/c 3T3 cells transformed by simian virus 40 has been demonstrated. Initial rates of transport of radioactively labeled L-leucine and alpha-aminoisobutyric acid were enhanced by the addition of NaCl (100 mM) to the reaction mixture at the start of the uptake process. This enhancement included a prominent "overshoot" during initial uptake. Slight stimulation of alpha-aminoisobutyric acid uptake was seen with K+, but none with Li+. The mediated nature of the uptake event for L-leucine was shown by saturation kinetics and by inhibition with L-valine. The transport assay measured predominantly intravesicular amino acid uptake rather than binding, as shown by the variation of uptake in response to changes in extravesicular osmolarity. Electron microscopy confirmed the presence of closed vesicles. Thus, amino acid transport has been characterized in an in vitro membrane vesicles system which should prove useful for studies of growth control.

Full text

PDF
1631

Images in this article

Selected References

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

  1. CHRISTENSEN H. N., RIGGS T. R., RAY N. E. Concentrative uptake of amino acids by erythrocytes in vitro. J Biol Chem. 1952 Jan;194(1):41–51. [PubMed] [Google Scholar]
  2. Christensen H. N., De Cespedes C., Handlogten M. E., Ronquist G. Modified transport substrates as probes for intramembrane gradients. Ann N Y Acad Sci. 1974 Feb 18;227:355–379. doi: 10.1111/j.1749-6632.1974.tb14400.x. [DOI] [PubMed] [Google Scholar]
  3. Colby C., Romano A. H. Phosphorylation but not transport of sugars is enhanced in virus-transformed mouse 3T3 cells. J Cell Physiol. 1975 Feb;85(1):15–23. doi: 10.1002/jcp.1040850103. [DOI] [PubMed] [Google Scholar]
  4. Colombini M., Johnstone R. M. Na+-gradient-stimulated AIB transport in membrane vesicles from Ehrlich ascites cells. J Membr Biol. 1974;18(3-4):315–334. doi: 10.1007/BF01870120. [DOI] [PubMed] [Google Scholar]
  5. Cunningham D. D., Pardee A. B. Transport changes rapidly initiated by serum addition to "contact inhibited" 3T3 cells. Proc Natl Acad Sci U S A. 1969 Nov;64(3):1049–1056. doi: 10.1073/pnas.64.3.1049. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Foster D. O., Pardee A. B. Transport of amino acids by confluent and nonconfluent 3T3 and polyoma virus-transformed 3T3 cells growing on glass cover slips. J Biol Chem. 1969 May 25;244(10):2675–2681. [PubMed] [Google Scholar]
  7. Harris E. D., Jr, DiBona D. R., Krane S. M. Collagenases in human synovial fluid. J Clin Invest. 1969 Nov;48(11):2104–2113. doi: 10.1172/JCI106177. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Hassell J. A., Colby C., Romano A. H. The effect of serum on the transport and phosphorylation of 2-deoxyglucose by untransformed and transformed mouse 3T3 cells. J Cell Physiol. 1975 Aug;86(1):37–45. doi: 10.1002/jcp.1040860106. [DOI] [PubMed] [Google Scholar]
  9. Hatanaka M., Augl C., Gilden R. V. Evidence for a functional change in the plasma membrane of murine sarcoma virus-infected mouse embryo cells. Transport and transport-associated phosphorylation of 14C-2-deoxy-D-glucose. J Biol Chem. 1970 Feb 25;245(4):714–717. [PubMed] [Google Scholar]
  10. Hochstadt J. The role of the membrane in the utilization of nucleic acid precursors. CRC Crit Rev Biochem. 1974 Mar;2(2):259–310. doi: 10.3109/10409237409105449. [DOI] [PubMed] [Google Scholar]
  11. Holley R. W. A unifying hypothesis concerning the nature of malignant growth. Proc Natl Acad Sci U S A. 1972 Oct;69(10):2840–2841. doi: 10.1073/pnas.69.10.2840. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Isselbacher K. J. Increased uptake of amino acids and 2-deoxy-D-glucose by virus-transformed cells in culture. Proc Natl Acad Sci U S A. 1972 Mar;69(3):585–589. doi: 10.1073/pnas.69.3.585. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Kaback H. R. Transport studies in bacterial membrane vesicles. Science. 1974 Dec 6;186(4167):882–892. doi: 10.1126/science.186.4167.882. [DOI] [PubMed] [Google Scholar]
  14. 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]
  15. Li C. C., Hochstadt J. Transport mechanisms in isolated plasma membranes. Nucleoside processing by membrane vesicles from mouse fibroblast cells grown in defined medium. J Biol Chem. 1976 Feb 25;251(4):1175–1180. [PubMed] [Google Scholar]
  16. Martin G. S., Venuta S., Weber M., Rubin H. Temperature-dependent alterations in sugar transport in cells infected by a temperature-sensitive mutant of Rous sarcoma virus. Proc Natl Acad Sci U S A. 1971 Nov;68(11):2739–2741. doi: 10.1073/pnas.68.11.2739. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. OXENDER D. L., CHRISTENSEN H. N. DISTINCT MEDIATING SYSTEMS FOR THE TRANSPORT OF NEUTRAL AMINO ACIDS BY THE EHRLICH CELL. J Biol Chem. 1963 Nov;238:3686–3699. [PubMed] [Google Scholar]
  18. Otsuka H., Moskowitz M. Difference in transport of leucine in attached and suspended 3T3 cells. J Cell Physiol. 1975 Jun;85(3):665–673. doi: 10.1002/jcp.1040850319. [DOI] [PubMed] [Google Scholar]
  19. PARDEE A. B. CELL DIVISION AND A HYPOTHESIS OF CANCER. Natl Cancer Inst Monogr. 1964 May;14:7–20. [PubMed] [Google Scholar]
  20. Quinlan D. C., Hochstadt J. An altered rate of uridine transport in membrane vesicles isolated from growing and quiescent mouse 3T3 fibroblast cells. Proc Natl Acad Sci U S A. 1974 Dec;71(12):5000–5003. doi: 10.1073/pnas.71.12.5000. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Quinlan D. C., Hochstadt J. Group translocation of the ribose moiety of inosine by vesicles of plasma membrane from T(3 cells transformed by Simian virus 40. J Biol Chem. 1976 Jan 25;251(2):344–354. [PubMed] [Google Scholar]
  22. RIGGS T. R., WALKER L. M., CHRISTENSEN H. N. Potassium migration and amino acid transport. J Biol Chem. 1958 Dec;233(6):1479–1484. [PubMed] [Google Scholar]
  23. Shohat B., Gitter S., Lavie D. Effect of withaferin A on Ehrlich ascites tumor cells--cytological observations. Int J Cancer. 1970 Mar 15;5(2):244–252. doi: 10.1002/ijc.2910050212. [DOI] [PubMed] [Google Scholar]
  24. Sigrist-Nelson K., Murer H., Hopfer U. Active alanine transport in isolated brush border membranes. J Biol Chem. 1975 Jul 25;250(14):5674–5680. [PubMed] [Google Scholar]
  25. Simoni R. D., Postma P. W. The energetics of bacterial active transport. Annu Rev Biochem. 1975;44:523–554. doi: 10.1146/annurev.bi.44.070175.002515. [DOI] [PubMed] [Google Scholar]
  26. Todaro G. J., Lazar G. K., Green H. The initiation of cell division in a contact-inhibited mammalian cell line. J Cell Physiol. 1965 Dec;66(3):325–333. doi: 10.1002/jcp.1030660310. [DOI] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES