Regulation of human apolipoprotein A-I gene expression by equine estrogens

Xia Zhang,Jei-Jun Jiao,Bhagu R Bhavnani,Shui-Pang Tam

doi:10.1016/s0022-2275(20)31505-4

Abstract

Estrogen replacement therapies, such as conjugated equine estrogen (CEE, Premarin®), reduce the risk of coronary heart disease among postmenopausal women. In the present study, a HepG2 stable cell line (HepG2/S) that harbors a luciferase reporter gene cassette with the human apolipoprotein A-I (apoA-I) promoter region was used to examine the activity of CEE components in modulating human apoA-I promoter activity. A number of estrogens modulated apoA-I promoter activity, with equilenin (Eqn) being the most potent. Eqn produced a 3-fold increase in apoA-I promoter activity and a similar increase in apoA-I mRNA without affecting its degradation rate. Nuclear runoff assays indicated that the transcription rate of the apoA-I gene was increased 2.5-fold in Eqn-treated cells. When HepG2/S cells were exposed to Eqn, apoA-I protein secretion increased by 80%, whereas the level of secreted apoA-II remained unchanged. Transient transfection studies with human apoA-I promoter constructs derived from pGL3-luciferase reporter plasmid were used to identify the cis-acting element involved in Eqn-mediated induction. The results demonstrated that the apoA-I electrophile/antioxidant response element (EpRE/ARE) might be responsible for the increase in apoA-I promoter activity by Eqn. Cotransfection experiments using estrogen receptor (ERα and/or ERβ) expression vectors have indicated that neither receptor can potentiate the Eqn-mediated induction of apoA-I promoter activity. In addition, mobility shift analysis using antibody against either ERα or ERβ cannot detect the presence of these receptors in the DNA-protein complex. The data indicate that Eqn can induce the promoter activity of the human apoA-I gene, leading to an increase in apoA-I mRNA levels and apoA-I protein secretion through an ER-independent pathway involving apoA-I EpRE/ARE.—Zhang, X., J-J. Jiao, B. R. Bhavnani, and S-P. Tam. Regulation of human apolipoprotein A-I gene expression by equine estrogens.

Highlights

Estrogen replacement therapies, such as conjugated equine estrogen (CEE, Premarin®), reduce the risk of coronary heart disease among postmenopausal women
We examined the effects of various equine estrogens on apolipoprotein A-I (apoA-I) promoter activity, using a stable hepatoma cell line G2 (HepG2) cell line (HepG2/S) that permanently harbors a gene cassette containing 256 bp of the 5Ј-flanking region of a human apoA-I gene fused to the luciferase gene
Using functional DNA analysis of the human apoA-I gene promoter, the individual role of each estrogenic component of CEE that may play in the regulation of apoA-I gene expression was investigated

Summary

Introduction

Estrogen replacement therapies, such as conjugated equine estrogen (CEE, Premarin®), reduce the risk of coronary heart disease among postmenopausal women. A HepG2 stable cell line (HepG2/S) that harbors a luciferase reporter gene cassette with the human apolipoprotein A-I (apoA-I) promoter region was used to examine the activity of CEE components in modulating human apoA-I promoter activity. Nuclear runoff assays indicated that the transcription rate of the apoA-I gene was increased 2.5-fold in Eqn-treated cells. The results demonstrated that the apoA-I electrophile/antioxidant response element (EpRE/ARE) might be responsible for the increase in apoA-I promoter activity by Eqn Cotransfection experiments using estrogen receptor (ER␣ and/or ER␤) expression vectors have indicated that neither receptor can potentiate the Eqnmediated induction of apoA-I promoter activity. The data indicate that Eqn can induce the promoter activity of the human apoA-I gene, leading to an increase in apoA-I mRNA levels and apoA-I protein secretion through an ER-independent pathway involving apoA-I EpRE/ARE.—Zhang, X., J-J.

Methods

Results

Conclusion