| Abstract: |
The balanced t(15;17) rearrangement found in acute promyelocytic leukemia (APL) cells fuses PML on chromosome 15 to the retinoic acid receptor α (RARα) on chromosome 17. PML/RARα is expressed in APL cells with the non-rearranged alleles, PML and RARα. Clinical remissions induced by all-trans-retinoic acid (RA) treatment of APL patients are linked to expression of PML/RARα, a transcription factor with reported dominant negative functions. The roles of PML and RARα in the RA response of APL have not yet been fully explored. This study examines these roles by individually transfecting RARα and PML into NB4 APL cells. NB4 is the sole APL cell line containing the t(15;17). RA treatment represses NB4 cell growth and induces a myeloid phentoype. Full length cDNAs for RARα and PML were individually cloned into a CMV-driven expression vector containing the neomycin resistance gene. Surprisingly, none of the obtained stable transfectants expressed exogenous RARα or PML mRNAs even when reverse transcription polymerase chain reaction (RT-PCR) detection assays were used. All clones expressed the neomycin resistance gene and were similar to parental NB4 cells in their growth and differentiation properties. An explanation explored for this lack of gene expression was that increased levels of RARα or PML might suppress APL cell growth. To examine this possibility, transfection experiments were repeated using an episomal vector-based expression system containing an SV40 driven RARα or PML cDNA and the hygromycin B resistance gene. A new selection strategy augmented expression of the desired cDNAs. A control episomal vector lacked a cDNA insert. Following electroporation and selection, exogenous RARα expression was obtained. Compared to controls, the growth of these transfectants was markedly inhibited before and after RA-treatment and these cells more prominently induced myeloid maturation markers. In contrast, exogenous PML expression was transient since these transfectants did not appear to propagate in culture. These findings indicate: (1) a growth disadvantage for NB4 cells having increased expression of RARα or PML and (2) increased RARα expression augmented RA-mediated maturation of NB4 cells. This implicates a role for RARα or PML in regulating the growth or differentiation of APL cells. It is hypothesized this occurs through antagonism of PML/RARα actions in these leukemic cells. |
| Keywords: |
controlled study; human cell; polymerase chain reaction; allele; cell division; reverse transcription polymerase chain reaction; neoplasm proteins; alleles; transcription factor; cell differentiation; drug resistance; transcription factors; leukemia, promyelocytic, acute; antibiotic resistance; molecular sequence data; messenger rna; rna, messenger; acute myeloblastic leukemia; oncogene proteins, fusion; simian virus 40; base sequence; retinoic acid; complementary dna; growth inhibition; chromosomes, human, pair 17; acute promyelocytic leukemia; chromosomes, human, pair 15; retinoic acid receptor; neomycin; tretinoin; pml; receptors, retinoic acid; chromosome translocation 17; human; priority journal; article; hygromycin b; chromosome translocation 15; support, non-u.s. gov't; support, u.s. gov't, p.h.s.; translocation (genetics); rarα
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