TY - JOUR
T1 - A single in vitro point mutation in the first non-translated exon silences transcription of the human apolipoprotein B gene in HepG2 cells
AU - Chuang, Samuel S.
AU - Das, Hriday K.
N1 - Funding Information:
This research was supported by grant HL49481 from the National Institutes of Health.
PY - 1999/1/4
Y1 - 1999/1/4
N2 - Hepatic cell-specific expression of the human apolipoprotein B (apoB) gene is controlled by at least four cis-acting elements located within the - 128 to +122 promoter region (S.S. Chuang, H.K. Das, Identification of trans- acting factors that interact with cis-acting elements present in the first nontranslated exon of the human apolipoprotein B gene, Biochem. Biophys. Res. Commun. 220 (1996) 553-562). Two cis-acting positive elements (-104 to -85; - 84 to -60) are located upstream from the start of transcription. A negative element (+20 to +40) and a strong positive element (+43 to +53) are located in the first non-translated exon of the human apolipoprotein B gene. Trans- acting factors BRF-2, BRF-1, BRF-3, and BRF-4 interact with the above four cis-acting elements respectively. In this study, we examine the roles of the upstream positive elements -104 to -85 and -84 to -60 in modulating transcriptional regulation of the apoB gene by downstream elements +20 to +40 and +43 to +53. Using in vitro mutagenesis and transient transfection experiments in HepG2 cells, the cis-acting element -84 to -60 has been found to be absolutely necessary for the function of the upstream element -104 to - 85 and downstream elements +20 to +40 and +43 to +53. In vitro mutagenesis of the downstream positive element +43 to +53 and transfection of the mutant promoter constructs in HepG2 cells reveal that nucleotide G at position +51 is essential for the strong positive activity of the element +43 to +53. A single substitution point mutation of nucleotide G to either A or T at position +51 reduces apolipoprotein B gene transcription substantially in HepG2 cells. These results suggest that a single substitution mutation in vivo, of nucleotide G to either A or T at position +51 in the downstream positive promoter element +43 to +53 may potentially cause hypobetalipoproteinemia, a heterozygous form of an autosomal-dominant disorder.
AB - Hepatic cell-specific expression of the human apolipoprotein B (apoB) gene is controlled by at least four cis-acting elements located within the - 128 to +122 promoter region (S.S. Chuang, H.K. Das, Identification of trans- acting factors that interact with cis-acting elements present in the first nontranslated exon of the human apolipoprotein B gene, Biochem. Biophys. Res. Commun. 220 (1996) 553-562). Two cis-acting positive elements (-104 to -85; - 84 to -60) are located upstream from the start of transcription. A negative element (+20 to +40) and a strong positive element (+43 to +53) are located in the first non-translated exon of the human apolipoprotein B gene. Trans- acting factors BRF-2, BRF-1, BRF-3, and BRF-4 interact with the above four cis-acting elements respectively. In this study, we examine the roles of the upstream positive elements -104 to -85 and -84 to -60 in modulating transcriptional regulation of the apoB gene by downstream elements +20 to +40 and +43 to +53. Using in vitro mutagenesis and transient transfection experiments in HepG2 cells, the cis-acting element -84 to -60 has been found to be absolutely necessary for the function of the upstream element -104 to - 85 and downstream elements +20 to +40 and +43 to +53. In vitro mutagenesis of the downstream positive element +43 to +53 and transfection of the mutant promoter constructs in HepG2 cells reveal that nucleotide G at position +51 is essential for the strong positive activity of the element +43 to +53. A single substitution point mutation of nucleotide G to either A or T at position +51 reduces apolipoprotein B gene transcription substantially in HepG2 cells. These results suggest that a single substitution mutation in vivo, of nucleotide G to either A or T at position +51 in the downstream positive promoter element +43 to +53 may potentially cause hypobetalipoproteinemia, a heterozygous form of an autosomal-dominant disorder.
KW - Apolipoprotein B gene
KW - Downstream
KW - Promoter activity
KW - Transcription factor
UR - http://www.scopus.com/inward/record.url?scp=0033521705&partnerID=8YFLogxK
U2 - 10.1016/S0005-2760(98)00117-9
DO - 10.1016/S0005-2760(98)00117-9
M3 - Article
C2 - 9989290
AN - SCOPUS:0033521705
SN - 1388-1981
VL - 1436
SP - 600
EP - 605
JO - Biochimica et Biophysica Acta - Molecular and Cell Biology of Lipids
JF - Biochimica et Biophysica Acta - Molecular and Cell Biology of Lipids
IS - 3
ER -