Interaction of epsilon-globin cis-acting control elements with erythroid-specific regulatory macromolecules.
We have used a competition assay to investigate the influence of erythroid-specific cellular factors on transcription from the human epsilon-globin major cap site promoter and the minor promoter located 200 base pairs (bp) upstream from the epsilon-globin cap site. In the human erythroid cell line K562, competition of the epsilon-globin major cap site promoter linked to the chloramphenicol acetyltransferase (CAT) gene (epsilon P-CAT) with the same promoter fragment linked to a neomycin resistance gene (epsilon P-NEO) leads to a reduction in CAT activity. This indicates the specific presence of K562 cells of factor(s) which interact with the 200-bp promoter fragment (isolated from the gene body or flanking sequences) to activate transcription from the epsilon-globin major cap site. Competition of the epsilon-globin major promoter (as epsilon P-CAT) with the upstream minor epsilon-globin promoter (as epsilon P2-NEO) also leads to a reduction in CAT activity, indicating that both promoters share erythroid-specific trans-acting factors. The reverse competition (epsilon P2-CAT with epsilon P-NEO) leads to an increase in CAT activity, suggesting that the existence of erythroid-specific factor(s) which repress transcription from the 200-bp-upstream epsilon-globin promoter.
Wu
J
J
Department of Genetics, College of Physicians and Surgeons, Columbia University, New York, NY 10032.
Grindlay
G J
GJ
Johnson
C
C
Allan
M
M
D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation; D005809:Genes, Regulator; D005914:Globins / Q000235:genetics*; D006801:Humans; D007700:Kinetics; D010957:Plasmids; D011401:Promoter Regions, Genetic; D012315:RNA Caps; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015183:Restriction Mapping; D014158:Transcription, Genetic; D014162:Transfection
D012315:RNA Caps; D011993:Recombinant Fusion Proteins; D005914:Globins; D015500:Chloramphenicol O-Acetyltransferase