Importance of low affinity Elf-1 sites in the regulation of lymphoid-specific inducible gene expression.
Elf-1 is an Ets family transcription factor that regulates a number of inducible lymphoid-specific genes, including those encoding interleukin 3 (IL-3), granulocyte/macrophage colony-stimulating factor (GM-CSF), and the IL-2 receptor (IL-2R) alpha chain. A minimal oligonucleotide spanning the IL-2R alpha Elf-1 site (-97/-84) bound Elf-1 poorly, but binding activity markedly increased when this oligonucleotide was multimerized or flanking sequences were added. This result is consistent with the requirement of accessory proteins for efficient Elf-1 binding, as has been demonstrated for the GM-CSF and IL-3 promoters. A binding site selection analysis revealed the optimal Elf-1 consensus motif to be A(A/t)(C/a)CCGGAAGT(A/S), which is similar to the consensus motif for the related Drosophila E74 protein. This minimal high affinity site could bind Elf-1 and functioned as a stronger transcription element than the -97/-84 IL-2R alpha oligonucleotide when cloned upstream of a heterologous promoter. In contrast, in the context of the IL-2R alpha promoter, conversion of the naturally occurring low affinity Elf-1 site to an optimal site decreased inducible activation of a reporter construct in Jurkat cells. This finding may be explained by the observation that another Ets family protein, ER GB/Fli-1, can efficiently bind only to the optimal site, and in this context, interferes with Elf-1 binding. Therefore, high affinity Elf-1 sites may lack sufficient binding specificity, whereas naturally occurring low affinity sites presumably favor the association of Elf-1 in the context of accessory proteins. These findings offer an explanation for the lack of optimal sites in any of the known Elf-1-regulated genes.
John
S
S
Laboratory of Molecular Immunology, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
Marais
R
R
Child
R
R
Light
Y
Y
Leonard
W J
WJ
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002460:Cell Line; D016384:Consensus Sequence; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000378:metabolism; D004330:Drosophila; D005786:Gene Expression Regulation; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000096:biosynthesis; D016325:HIV Long Terminal Repeat; D015497:HIV-1 / Q000235:genetics; D015498:HIV-2 / Q000235:genetics; D007313:Insecta; D007377:Interleukin-3 / Q000096:biosynthesis; D008214:Lymphocytes / Q000276:immunology / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009838:Oligodeoxyribonucleotides; D016133:Polymerase Chain Reaction; D015375:Receptors, Interleukin-2 / Q000096:biosynthesis*; D012689:Sequence Homology, Nucleic Acid; D014162:Transfection
D017931:DNA Primers; D004268:DNA-Binding Proteins; D007377:Interleukin-3; D009838:Oligodeoxyribonucleotides; D015375:Receptors, Interleukin-2; D016178:Granulocyte-Macrophage Colony-Stimulating Factor