Retinoic acid-resistant HL-60R cells harbor a point mutation in the retinoic acid receptor ligand-binding domain that confers dominant negative activity.
Retinoic acid (RA) induces granulocytic differentiation of acute promyelocytic leukemia (APL) cells and is a useful therapeutic agent for patients with this disease. In the HL-60 promyelocytic leukemia cell line, this RA-induced granulocytic differentiation appears to be directly mediated through the RA receptor (RAR-alpha). We have previously identified a mutant subclone of HL-60 (designated HL-60R) that exhibits relative resistance to RA and that harbors RA receptors with markedly reduced affinity for RA. In the present study, we have now identified the genetic basis for this aberrant RA receptor activity. DNA sequencing of polymerase chain reaction-amplified cDNA products corresponding to the RAR-alpha ligand-binding domain shows a point mutation in RAR-alpha codon 411 in this mutant HL-60R subclone. This specific C-->T mutation generates a termination codon resulting in the truncation of 52 amino acids at the COOH terminal end of RAR-alpha. In cotransfection studies, expression vectors harboring this mutated RAR-alpha exhibit dominant negative activity with respect to the trans-activating function of the normal RAR-alpha. Although our observations are limited to HL-60 cells, similar RA receptor mutations might play an important role in the acquisition of RA resistance in RA-treated APL patients.
Robertson
K A
KA
Molecular Medicine Program, Fred Hutchinson Cancer Center, Seattle, WA 98104.
Emami
B
B
Collins
S J
SJ
D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D002352:Carrier Proteins / Q000235:genetics*; D002454:Cell Differentiation / Q000187:drug effects; D004247:DNA / Q000737:chemistry; D004351:Drug Resistance / Q000235:genetics; D006801:Humans; D015473:Leukemia, Promyelocytic, Acute / Q000235:genetics* / Q000473:pathology; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D018168:Receptors, Retinoic Acid; D015533:Transcriptional Activation; D014162:Transfection; D014212:Tretinoin / Q000494:pharmacology*; D014407:Tumor Cells, Cultured
D002352:Carrier Proteins; D018168:Receptors, Retinoic Acid; D014212:Tretinoin; D004247:DNA