Constrasting effect of oncogene expression on two carrier-mediated systems internalizing folate compounds in fisher rat 3T3 cells Journal Article
| Authors: | Kühnel, J. M.; Chiao, J. H.; Sirotnak, F. M. |
| Article Title: | Constrasting effect of oncogene expression on two carrier-mediated systems internalizing folate compounds in fisher rat 3T3 cells |
| Abstract: | Folate compound transport into Fisher rat 3T3 (FR3T3) cells at physiological pH occurs predominantly by an acid pH-dependent, mobile carrier system. However, influx of [3H]MTX by this system is 3-4-fold higher at pH 6 than at pH 7.5, the optimum for RFC-1-mediated folate compound transport. This acid pH dependency reflects an alteration of influx V(max) rather than of influx K(m) in these cells at different pH. Acid pH-dependent folate compound transport interacts effectively with MTX, 5lLCHO-folateH4, 5lLCH3-folateH4 and folic acid as permeants (influx Ki = 2.7-5.3 μM). The relative inhibition of influx of [3H]MTX by the organic anions, probenecid, and PO4 was different than for RFC-1 mediated influx. The folate requirements for growth in culture of FR3T3 cells and cytotoxicity of MTX compared to L1210 cells reflects the interactions of these folate compounds with acid pH-dependent folate transport. 5lLCHO-folateH4 and PO4 act as exchange anions for this system but their transpositioning has variable effects on transport. 5lLCHO-folateH4 inhibits influx (decelerative equilibrium exchange) but stimulates efftux of [3H]MTX (accelerative equilibrium exchange) while PO4 inhibits efflux. In FR3T3 cells transfected with cmyc and Hras, influx V(max) for [3H]MTX is downregulated 4-fold and 9-fold, respectively. At the same time, RFC-1 expression, which is detectable in FR3T3 cells at the level of its mRNA and RFC-1 mediated folate compound transport, is increased 3-5-fold in these transfectants. The increase in RFC-1 expression in FR3T3Hras cells appears to result from a higher rate of transcription of the gene in these cells as determined by a luciferase reporter gene assay of RFC-1 promoter activity. This downregulation of the acid pH dependent system and concomitant upregulation of the RFC-1 mediated system markedly altered pH dependency for influx of [3H]MTX in these transfectants compared to that seen in untransfected cells. We conclude that the major route for internalization at a physiological pH of folate compounds in FR3T3 cells is by an acid pH-dependent carrier-mediated system independent of RFC-1 expression and is downregulated by oncogene expression. (C) 2000 Wiley-Liss, Inc. |
| Keywords: | controlled study; carrier protein; genetics; nonhuman; comparative study; methotrexate; animal cell; animal; metabolism; animals; gene expression; cell line; luciferase; membrane proteins; ph; genetic transcription; transfection; oncogene; genetic transfection; kinetics; messenger rna; nucleotide sequence; genes, myc; carrier proteins; membrane protein; rat; base sequence; rats; folic acid; dna primers; primer dna; genes, ras; phosphate; drug transport; oncogene ras; oncogene myc; internalization; hydrogen-ion concentration; probenecid; membrane transport proteins; reduced folate carrier; anion; biological transport, active; active transport; leukemia l1210; folic acid derivative; priority journal; article; leukemia l 1210 |
| Journal Title: | Journal of Cellular Physiology |
| Volume: | 184 |
| Issue: | 3 |
| ISSN: | 0021-9541 |
| Publisher: | John Wiley & Sons, Inc. |
| Date Published: | 2000-09-01 |
| Start Page: | 364 |
| End Page: | 372 |
| Language: | English |
| DOI: | 10.1002/1097-4652(200009)184:3<364::aid-jcp11>3.0.co;2-n |
| PUBMED: | 10911368 |
| PROVIDER: | scopus |
| DOI/URL: | |
| Notes: | Export Date: 18 November 2015 -- Source: Scopus |
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