Chromosomal localization of the murine RFC-1 gene encoding a folate transporter and its amplification in an antifolate resistant variant overproducing the transporter Journal Article
| Authors: | Roy, K.; Chiao, J. H.; Spengler, B. A.; Tolner, B.; Yang, C. H.; Biedler, J. L.; Sirotnak, F. M. |
| Article Title: | Chromosomal localization of the murine RFC-1 gene encoding a folate transporter and its amplification in an antifolate resistant variant overproducing the transporter |
| Abstract: | A variant of the L1210 cell (L1210/R83) selected in the presence of the lipophilic anti-folate, metoprine, and a concentration of the natural diastereoisomer of 5-formyltetrahydrofolate, lL5CHO-folateH4, suboptimum for growth exhibited a 35-fold increase compared to parental L1210 cells in one- carbon, reduced folate transport. This was evidenced by the increase in V(max) for [3H]MTX (methotrexate) influx and a commensurate increase in the amount of the 46 kilodalton (kDa) transport protein and reduced folate carrier (RFC-1) mRNA. The variant is resistant to lipophilic antifolates, but shows collateral sensitivity to classical folate analogues. Karyotype analysis of L1210/R83 cells revealed the presence of several new chromosome abnormalities. One of these was a large, submetacentric marker chromosome comprising a normal 10 and a longer, abnormally banded arm of uncertain origin which exhibited an interstitial, palely staining, HSR-like segment. The results of Southern and Northern blotting showed that the RFC-1 gene copy number and RNA transcript level were markedly increased (30-35 fold) in L1210/R83 cells. Fluorescence in situ hybridization (FISH) analysis revealed that the HSR-like segment in these cells was the site of amplified RFC-1 genes. Independent revertant subclones, obtained following growth in the absence of selection pressure, showed four- to 12-fold decreases in [3H]MTX influx V(max) and in amount of NHS [N-hydroxysuccinimide)-[3H]MTX affinity labeled one-carbon, reduced folate transporter compared to L1210/R83 cells. RFC-1 gene copy number also decreased, and the mean length of the HSR in these revertants declined 1.6- to 5-fold. Based upon genomic nucleotide sequencing, the RFC-1 gene in the normal mouse genome was localized to chromosome 10 in close association with the alpha 1 (Col18a1) collagen gene at 10B3(locus 41cM). The close association of these genes was confirmed by other data showing that the alpha 1 collagen gene was co-amplified in L1210/R83 cells. These results document the amplification at the site era putative HSR in an L1210 cell variant of the RFC-1 gene regulating expression of the one-carbon, reduced folate transporter. |
| Keywords: | controlled study; carrier protein; unclassified drug; nonhuman; methotrexate; animal cell; mouse; animals; mice; gene amplification; membrane proteins; drug resistance, neoplasm; tumor cells, cultured; chromosome aberration; fluorescence in situ hybridization; messenger rna; rna, messenger; folinic acid; lymphoma cell; carrier proteins; collagen; folic acid; chromosome aberrations; gene dosage; folic acid antagonist; folic acid antagonists; tetrahydrofolate dehydrogenase; karyotype; karyotyping; chromosomal localization; chromosome mapping; tritium; chromosome 10; membrane transport proteins; succinimide derivative; leukemia l1210; chromosome disorders; chromosomes, human, pair 10; humans; priority journal; article; metodiclorofen; n hydroxysuccinimide |
| Journal Title: | Cancer Genetics and Cytogenetics |
| Volume: | 105 |
| Issue: | 1 |
| ISSN: | 0165-4608 |
| Publisher: | Elsevier Science, Inc. |
| Date Published: | 1998-08-01 |
| Start Page: | 29 |
| End Page: | 38 |
| Language: | English |
| DOI: | 10.1016/s0165-4608(98)00005-3 |
| PUBMED: | 9689927 |
| PROVIDER: | scopus |
| DOI/URL: | |
| Notes: | Article -- Export Date: 12 December 2016 -- Source: Scopus |
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