Up-regulation of LRP16 mRNA by 17β-estradiol through activation of estrogen receptor α (ERα), but not ERβ, and promotion of human breast cancer MCF-7 cell proliferation: A preliminary report

Abstract

LRP16 is a novel gene cloned from lymphocytic cells, and its function is not known. The expression level of LRP16 mRNA was up-regulated by estrogen in breast cancer MCF-7 cells based on the computed aided serial analysis of gene expression (SAGE) analysis. In this study, we investigate the effect of 17β-estradiol (17β-E2) on the expression of LRP16 mRNA and the effects of overexpression of LRP16 on the proliferation of cultured MCF-7 cells and the possible mechanisms involved. The expression level of LRP16 mRNA induced by 17β-E2 was determined by Northern blot analysis. LRP16 promoter-controlled luciferase expression vector (pGL3-So) was co-transfected with various nuclear receptors, including estrogen receptor α and β (ERα and ERβ), glucocorticoid receptor α (GRα), androgen receptor (AR) and peroxisome-proliferator activated receptor γ and α (PPARγ and PPARα) into COS-7 cells, and the relative luciferase activity was measured using Dual-luciferase report assay systems. The effect of overexpression of LRP16 on MCF-7 proliferation was examined by the Trypan Blue exclusion method, and the cell cycle was analyzed by flow cytometry. The expression levels of cyclin E, p53 and p21WAF1/CIP1 proteins were determined by Western blot analysis. The results showed (1) 17β-E2 induced a five- to eightfold increase in LRP16 mRNA levels in MCF-7 cells; (2) the relative luciferase activities in the COS-7 cells co-transfected by pGL3-So and ERα or AR were 7.8-fold and 11-fold respectively of those in the control cells transfected by pGL3-So alone; (3) overexpression of LRP16 stimulated MCF-7 cell proliferation, and the numbers of cells in the S-phase of the cell cycle in cells transfected with LRP16 increased about 10% compared with the control cells; and (4) cyclin E levels were much higher in cells with overexpression of LRP16 than in the control cells, while the expression levels of p53 and p21WAF1/CIP1 were not different between the two groups of cells. From these results we concluded that estrogen up-regulates the expression level of LRP16 mRNA through activation of ERα and that overexpression of LRP16 promotes MCF-7 cell proliferation probably by increasing cyclin E.