A base substitution in the promoter associated with the human haptoglobin 2-1 modified phenotype decreases transcriptional activity and responsiveness to interleukin-6 in human hepatoma cells.
An A-to-C base substitution at nucleotide position -61 in the promoter region of the human haptoglobin gene (Hp) has been shown to be strongly associated with the haptoglobin 2-1 modified (Hp2-1mod) phenotype. In order to investigate whether this base substitution is the cause of reduced expression of the Hp2 allele relative to the Hp1 allele in individuals with the Hp2-1mod phenotype, we used the chloramphenicol acetyl transferase (CAT) expression system to evaluate promoter function. In HepG2 cells, which normally express their endogenous haptoglobin genes, CAT plasmid constructs with the -61C base change in the promoter had about 10-fold-lower transcriptional activity after transfection than did the Hp control construct. The -61C substitution also rendered the construct unresponsive to treatment by interleukin-6 after transfection into Hep3B2 cells, which normally do not express haptoglobin but do so in response to stimulation by acute-phase reactants. In addition, two base substitutions, T to A and A to G, at positions -104 and -55G, respectively, in the promoter region of the Hp1 allele, are also associated with the Hp2-1mod phenotype. CAT constructs with both substitutions (-104A-55G) and with one substitution (-55G) showed activity similar to that in the Hp control when transfected into both HepG2 and Hep3B2 cells, although interleukin-6 induction was less than with the Hp control construct. These results further support the hypothesis that the Hp2-1mod phenotype results, in part, from the -61C mutation in the promoter region of the Hp2 gene.
Grant
D J
DJ
Department of Pathology, University of North Carolina, Chapel Hill 27599.
Maeda
N
N
D000483:Alleles; D006528:Carcinoma, Hepatocellular / Q000235:genetics; D015500:Chloramphenicol O-Acetyltransferase / Q000378:metabolism; D006242:Haptoglobins / Q000187:drug effects / Q000235:genetics* / Q000502:physiology; D006801:Humans; D015850:Interleukin-6 / Q000494:pharmacology*; D008113:Liver Neoplasms / Q000235:genetics; D010641:Phenotype; D010957:Plasmids; D017354:Point Mutation; D011401:Promoter Regions, Genetic / Q000187:drug effects; D011994:Recombinant Proteins / Q000494:pharmacology; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured
D006242:Haptoglobins; D015850:Interleukin-6; D011994:Recombinant Proteins; D015500:Chloramphenicol O-Acetyltransferase