The beta-globin promoter is important for recruitment of erythroid Krüppel-like factor to the locus control region in erythroid cells.
Erythroid Krüppel-like factor (EKLF), which binds to the CACCC box in the beta-globin promoter, is required for the expression of the beta-globin gene in adult erythroid cells. It was recently demonstrated that EKLF is also required for the activity of the beta-globin locus control region (LCR) 5'HS3. Some evidence suggests that the LCR and the beta-globin promoter interact in adult erythroid cells, and the network of protein-protein interactions that exists between these two elements may regulate how EKLF is recruited to the LCR. In this report, we use the PIN*POINT assay to study the role of the promoter on the recruitment of EKLF to 5'HS2 and 5'HS3 of the LCR. We find that recruitment of EKLF to 5'HS2 requires the TATA box, but recruitment to 5'HS3 depends on the CACCC and TATA boxes of the beta-globin promoter. Furthermore, recruitment of EKLF to 5'HS3 only occurred in beta-globin-expressing murine erythroid leukemia cells, whereas recruitment of EKLF to 5'HS2 occurred in both gamma-globin-expressing K562 cells and murine erythroid leukemia cells. Unlike EKLF, Sp1, which also binds to CACCC boxes, is not recruited to 5'HS3. We have also examined how one 5'HS affects the recruitment of EKLF to another 5'HS. We have found that the recruitment of EKLF to 5'HS3 depends on the presence of 5'HS2 in cis, but the recruitment to 5'HS2 does not depend on 5'HS3. Based on these results, we present a model that illustrates how EKLF may be recruited to the beta-globin locus.
Lee
J S
JS
Molecular Hematology Branch, National Heart Lung and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA.
Lee
C H
CH
Chung
J H
JH
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D003062:Codon; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004912:Erythrocytes / Q000378:metabolism*; D005914:Globins / Q000096:biosynthesis / Q000235:genetics*; D006801:Humans; D007438:Introns; D020014:K562 Cells; D051741:Kruppel-Like Transcription Factors; D007942:Leukemia, Experimental; D019901:Locus Control Region; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010750:Phosphoproteins / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D016385:TATA Box; D014157:Transcription Factors / Q000378:metabolism*; D014162:Transfection; D014407:Tumor Cells, Cultured; D016335:Zinc Fingers
D003062:Codon; D004268:DNA-Binding Proteins; D051741:Kruppel-Like Transcription Factors; D010750:Phosphoproteins; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; C080774:erythroid Kruppel-like factor; D005914:Globins