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The Journal of Membrane Biology

, Volume 75, Issue 1, pp 11–20 | Cite as

Relationships between carrier-mediated transport of folate compounds by L1210 leukemia cells: Evidence for multiplicity of entry routes with different kinetic properties expressed in plasma membrane vesicles

  • C. -H. Yang
  • M. Dembo
  • F. M. Sirotnak
Articles

Summary

Various independent kinetic criteria for indicating multiplicity of carrier-mediated entry of folate compounds into L1210 cell plasma membrane vesicles are studied. We find a marked inconsistency between values for influxKm andKix in reciprocal experiments measuring competition between various folate compounds as well as inconsistent effects of transloading shown for 5-formyltetrahydrofolate influx, but not folic acid influx. These results argue strongly against a one-carrier model for transport of folate compounds. The most straightforward interpretation of our data is that two distinct transport systems mediate entry of folate compounds in L1210 plasma membrane vesicles. If a two-carrier model is correct, then our data indicate that one of the carriers has low capacity and high affinity for folate coenzymes and methotrexate. This system is apparently negligible as a transport route for folic acid. Transtimulation of initial influx by substrates of the low capacity system is obtained following transloading with coenzymes but not by transloading with folic acid. Our data indicate that the second folate transport system postulated by the two-carrier model has a low affinity for all the folate compounds studied. Nevertheless, the putative second system is significant, especially for folic acid transport, because it has a much higher capacity than the first transport system. In contrast to the first system, transloading with any of the folate compounds studied had no effect on initial influx mediated by the second folate transport system. The two systems are also differentially inhibited bypCMBS, DIDS and SITS and the influxVmax for the high-affinity/low-capacity system was altered in a vesicle preparation derived from a methotrexate resistant L1210 cell line.

Key Words

Folate compounds transport multiplicity L1210 cells 

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References

  1. 1.
    Blakely, R.L. 1969. The Biochemistry of Folic Acid and Related Pteridines. pp. 139–181. John Wiley and Sons, N.Y.Google Scholar
  2. 2.
    Coward, J.K., Chello, P.L., Cashmore, A.R., Parameswaren, K.H., DeAngeles, L.M., Bertino, J.R. 1975. 5-Methyl-5,6,7,8-tetrahydropteroyl oligo-γ-l-glutamates: synthesis and kinetic studies with methionine synthetase from bovine brain.Biochemistry 14:1548–1552PubMedGoogle Scholar
  3. 3.
    Eilam, Y., Stein, W.D. 1974. Kinetic studies of transport across red cell membranes.Methods Membr. Biol. 2:283–354Google Scholar
  4. 4.
    Goldman, I.D. 1971. The characteristics of membrane transport of amethopterin and the natural occurring folates.Ann. N.Y. Acad. Sci. 186:400–422PubMedGoogle Scholar
  5. 5.
    Goldman, I.D. 1971. A model system for the study of hetero-exchange diffusion: Methotrexate-folate interactions in L1210 leukemia and Ehrlich ascites tumor cells.Biochim Biophys. Acta 233:624–634PubMedGoogle Scholar
  6. 6.
    Hissen, P.J., Hilf, R. 1978. Characteristics of proline transport into R3230 AC mammary tumor cells.Biochim. Biophys Acta 408:401–412Google Scholar
  7. 7.
    Hochstadt, J., Quinlan, D.C., Rader, R., Li, C.C. 1975. Use of isolated membrane vesicles in transport studies.Methods Membrane Biol. 5:117–126Google Scholar
  8. 8.
    Horne, D.W., Briggs, W.T., Wagner, C. 1976. A functional active transport system for methotrexate in freshly isolated hepatocytes.Biochem. Biophys. Res. Commun. 68:70–76PubMedGoogle Scholar
  9. 9.
    Horne, D.W., Briggs, W.T., Wagner, C. 1978. Transport of 5-methyltetrahydrofolate acid and folic acid in freshly isolated hepatocytes.J. Biol. Chem. 253:3529–3536PubMedGoogle Scholar
  10. 10.
    Huennekens, F.M., Vitols, K.S., Suresh, M.R., Henderson, G.R. 1981. Transport of folate compounds in L1210 cells: Components and mechanisms.In: Molecular Actions and Targets for Cancer Chemotherapeutic Agents. A.C. Sartorelli, J.S. Lazo and J.R. Bertino, editors. pp. 333–346. Academic Press, New York.Google Scholar
  11. 11.
    Hutchison, D.J., Robinson, D.J., Martin, D., Ittensohn, D.L., Dillenberg, J. 1963. Effects of selected cancer chemotherapeutic drugs on the survival time of mice with L1210 leukemia: Relative responses of antimetabolite-resistant strains.Cancer Res. 22:57–72Google Scholar
  12. 12.
    Jackson, R.C., Niethammer, D., Huennekens, F.M. 1975. Enzyme and transport mechanisms of amethopterin resistance in L1210 mouse leukemia cells.Cancer Biochem. Biophys. 1:151–155Google Scholar
  13. 13.
    Lichtenstein, L.S., Oliverio, V.T., Goldman, I.D. 1969. Characteristics of folic acid transport in the L1210 leukemia cell.Biochim. Biophys. Acta 193:456–467PubMedGoogle Scholar
  14. 14.
    Lowry, O.H., Rosebrough, N.J., Farr, A.L., Randall, R.J. 1951. Protein measurement with the Folin phenol reagent.J. Biol. Chem. 193:265–275PubMedGoogle Scholar
  15. 15.
    Nahas, A., Nixon, P.F., Bertino, J.R. 1972. Uptake and metabolism of N5-formyltetrahydrofolate by L1210 leukemia cells.Cancer Res. 32:1416–1421PubMedGoogle Scholar
  16. 16.
    Niethammer, D., Jackson, R.C. 1975. Changes of molecular properties associated with the development of resistance against methotrexate in human lymphoblastoid cells.Eur. J. Cancer 11:845–854PubMedGoogle Scholar
  17. 17.
    Rader, J.R., Niethammer, D., Huennekens, F.M. 1974. Effects of sulfhydryl inhibitors upon transport of folate compounds into L1210 cells.Biochem. Pharmacol. 23:2057–2059PubMedGoogle Scholar
  18. 18.
    Shortman, K., Williams, N., Adams, P. 1972. The separation of different cell class from lymphoid organs. V. Simple procedures for removal of cell debris, damaged cells and erythroid cells from lymphoid cell suspensions.J. Immunol. Methods 1:273–287PubMedGoogle Scholar
  19. 19.
    Silber, R., Huennekens, F.M., Gabrio, B.W. 1963. Studies on the interaction of tritium-labelled aminopterin with dihydrofolate reductase.Arch. Biochem. Biophys. 100:525–530PubMedGoogle Scholar
  20. 20.
    Sirotnak, F.M. 1980. Correlates of folate analog transport, pharmacokinetics and selective antitumor action.Pharmacol. Ther. 8:71–103Google Scholar
  21. 21.
    Sirotnak, F.M., Chello, P.L., Moccio, D.M., Kisliuk, R.L., Combepine, R.L., Gaumont, H., Montgomery, J.A. 1979. Stereospecificity at carbon 6 of formyltetrahydrofolate as a competitive inhibitor of transport and cytotoxicity of methotrexatein vitro.Biochem. Pharmacol. 28:2993–2997PubMedGoogle Scholar
  22. 22.
    Sirotnak, F.M., Donati, G.J., Hutchison, D.J. 1963. Folic acid derivatives synthesized during growth ofDiplococcus pneumoniae.J. Bacteriol. 85:658–665PubMedGoogle Scholar
  23. 23.
    Sirotnak, F.M., Donsbach, R.C. 1976. Kinetic correlates of methotrexate transport and therapeutic responsiveness in murine tumors.Cancer Res. 36:1151–1158PubMedGoogle Scholar
  24. 24.
    Sirotnak, F.M., Moccio, D.M., Goutas, L.J., Kelleher, L.E., Montgomery, J.A. 1982. Biochemical correlates of responsiveness and collateral sensitivity of some methotrexate-resistant murine tumors to the lipophilic antifolate, metoprine.Cancer Res. 42:924–928PubMedGoogle Scholar
  25. 25.
    Sirotnak, F.M., Moccio, D.M., Kelleher, L.E., Goutas, L.J. 1981. Relative frequency and kinetic properties of transport-defective phenotypes among methotrexate-resistant L1210 clonal cell lines derivedin vitro.Cancer Res. 41:4447–4452Google Scholar
  26. 26.
    Suresh, M.R., Henderson, G.B., Huennekens, F.M. 1979. Folate uptake in L1210 cells: Medication by an adenine transport system.Biochem. Biophys. Res. Commun. 87:135–139PubMedGoogle Scholar
  27. 27.
    Yang, C.-H., Dembo, M., Sirotnak, F.M. 1982. Two compartment behavior during transport of folate compounds in L1210 cell plasma membrane vesicles.J. Membrane Biol. 68:19–28Google Scholar
  28. 28.
    Yang, C.-H., Peterson, R.H.F., Sirotnak, F.M., Chello, P.L. 1979. Folate analog transport by plasma membrane vesicles isolated from L1210 leukemia cells.J. Biol. Chem. 254:1402–1407PubMedGoogle Scholar

Copyright information

© Springer-Verlag New York Inc. 1983

Authors and Affiliations

  • C. -H. Yang
    • 1
    • 2
  • M. Dembo
    • 1
    • 2
  • F. M. Sirotnak
    • 1
    • 2
  1. 1.Laboratory for Molecular TherapeuticsMemorial Sloan-Kettering Cancer CenterNew York
  2. 2.Theoretical DivisionLos Alamos National LaboratoryLos Alamos

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