Inhibition of β protein 1 expression enhances β-globin promoter activity and β-globin mRNA levels in the human erythroleukemia (K562) cell line

Abstract

Expression of globin genes within the β-globin cluster is regulated to produce different hemoglobin β chains during the embryonic (ε), fetal (γ), and adult (β) stages of erythroid cell development. Each distinct hemoglobin tetramer exhibits a different affinity for oxygen, resulting in optimal O 2 / CO 2 exchange between the developing fetus and mother. Thus, developmentally appropriate transcriptional activation of each different β chain is essential. This is achieved by the binding of transcriptional activator proteins to the successive β-chain promoters; the locus control region (LCR), consisting of five DNase I-hypersensitive sites (HS1-5) located from 8 to 22 kbp 5′ to the ε-globin gene, also Offprint requests to:

BP1 motif in the human β-globin promoter affects β-globin expression during embryonic/fetal erythropoiesis in transgenic mice bearing the human β-globin gene

“…EGFP-reporter constructs were prepared as described previously [9]. Briefly, a 690-bp DNA fragment of the adult human β-globin regulatory region, from −640 to +50 relative to the cap site (+1), was amplified by the polymerase chain reaction (PCR) from a 35.4-kb cosmid containing the β-globin gene locus.…”

“…Current hypotheses about the possible mechanisms for negative regulation include competition between genes for interaction with the locus control region (LCR) [1] and autonomous gene silencing involving sequences located in the proximal and distal promoters [2–4]. BP1 (β protein 1) plays an important role in β-globin gene regulation, because binding of BP1 to its site on the promoter of the adult β-globin gene has a silencing effect on β-globin transcription in vitro [5–9]. The adult β-globin gene includes two upstream silencer regions (silencer I and II), and BP1 binds to both of these regions [5,8].…”

“…In our earlier studies of BP1 function in K562 cells, an erythroleukemia cell line with an embryonic-fetal phenotype, we have shown that introduction of multiple mutations in two BP1 binding sites enhances β-globin promoter activity [6]. Inhibition of BP1 expression also has been demonstrated to increase levels of β-globin mRNA in K562 cells [9]. To better understand effects of the negative regulation of β-globin expression at the level of the whole organism, we have developed a transgenic mouse expressing the human β-globin gene mutated at the BP1 binding site.…”

A Homeobox Gene Related to Drosophila Distal-Less Promotes Ovarian Tumorigenicity by Inducing Expression of Vascular Endothelial Growth Factor and Fibroblast Growth Factor-2

“…Sequences of DLX4 and negative control siRNA oligomers (synthesized by Qiagen-Xeragon, Germantown, MD) were the same as used by other investigators 22 and were as follows: DLX4 siRNA: 5Ј-GCUCCUGAAGCA-GAAUUCUdTdT-3Ј; negative control siRNA: 5Ј-CAGUUAGAGGAGAAAAAGGdTdT-3Ј. One hundred thousand cells were seeded per well in six-well plates and allowed to adhere overnight.…”

Recombinant erythroid Kruppel-like factor fused to GATA1 up-regulates delta- and gamma-globin expression in erythroid cells

“…10,23 The ␦-globin reporter construct contains a 680-bp fragment of the human ␦-globin promoter (from Ϫ612 to ϩ68 relative to the cap site). The ␤-globin reporter construct contains a 690-bp fragment of the human ␤-globin promoter (from Ϫ640 to ϩ50).…”