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Expression of Endogenous and Transfected Apolipoprotein II and Vitellogenin II Genes in an Estrogen Responsive Chicken Liver Cell Line

Department of Pharmacological Sciences, Health Sciences Center, State University of New York at Stony Brook Stony Brook, New York 11794
Department of Molecular Biology Graduate Program, Health Sciences Center, State University of New York at Stony Brook Stony Brook, New York 11794
Institute for Cancer Research, Fox Chase Cancer Center Philadelphia, Pennsylvania 19111
Biochemistry Department, Dalhousie University Halifax, Nova Scotia, B3H 4H7, Canada

Address requests for reprints to: David L. Williams, Department of Pharmacological Sciences, Health Science Center, State University of New York at Stony Brook, Stony Brook, New York 11794.

Abstract

A recently described chicken liver cell line, LMH, was characterized to evaluate responsiveness to estrogen. Expression of the endogenous apolipoprotein (apo) II gene was induced by 17 β-estradiol when LMH cells were cultured with chicken serum. The response was low and yielded apoll mRNA at only 0.3% of the level seen in estrogenized rooster liver. Higher levels of apoll mRNA were achieved when LMH cells were transiently transfected with an expression plasmid for estrogen receptor. A transfected apoll gene was strongly expressed only when cotransfected with receptor. Expression of the endogenous vitellogenin (VTG) II gene was not detected. However, when cotransfected with a receptor expression plasmid, VTG II reporter plasmids were expressed in LMH cells in response to 17 β- estradiol. These results suggest that estrogen responsiveness of LMH cells is limited by the availability of functional receptor. Low levels of estrogen receptor mRNA were detected in LMH cells, and receptor binding sites and mRNA were greatly increased following transient transfection with a receptor expression plasmid. Using this transient transfection protocol, several VTG II reporter plasmids were compared in LMH cells and chick embryo fibroblasts. A plasmid containing VTG II estrogen response elements linked to a heterologous promoter was regulated by estrogen in both cell types. In contrast, reporter plasmids containing the VTG II promoter were regulated by estrogen in LMH cells but were not expressed at all in chick embryo fibro-blasts. These results suggest that regulation of the VTG II gene involves cell type-specific elements in addition to estrogen response elements. LMH cells will provide a homologous cell culture model for the analysis of liver-specific and estrogen-responsive transcriptional elements in avian genes.

FOOTNOTES

This work was supported by NIH Grant DK-18171 (to D.L.W.), NIH Grant CA-06927, and Public Health Service Grant 35535 (to J.B.E.B.), Medical Research Council of Canada Grant MT4880 (to C.B.L.), and NIH predoctoral award (National Research Service Award) GM-08065 (to R.B.).

Received for publication September 8, 1989. Revision received November 16, 1989. Accepted for publication November 28, 1989.

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