Transcriptional regulation of Fhl1 by estradiol in rat myoblastocytes

Fhl1 (Four and a Half LIM domain 1) regulates muscle growth and development. In addition, skeletal myoblast growth is significantly affected by gender differences, implicating estrogen in the regulation of muscle development. We sought to determine if estrogen influences Fhl1 gene expression levels in rat L6GNR4 myoblastocytes that express the estrogen receptor β (ERβ), while luciferase assay, electrophoretic mobility shift assay (EMSA), and chromatin immunoprecipitation (ChIP) assay were employed to confirm the interaction between ERβ and Fhl1. Treatment of L6GNR4 cells with physiological levels of 17β-estradiol (E2) results in markedly decreased endogenous Fhl1 expression. Tamoxifen, an ER antagonist, partially reverses E2-mediated Fhl1 down-regulation in L6GNR4 cells. Furthermore, luciferase assay and EMSA identified a novel promoter region of Fhl1 that directly interacts with ERβ. ChIP of the ERβ- Fhl1 promoter complex from L6GNR4 cells confirmed that endogenous ERβ interacts with this region. These data indicate that E2 down-regulates Fhl1 expression through its binding to the ERβ. This is the first report of a small molecule that can affect Fhl1 expression. E2 may therefore be useful in developing new strategies for regulating Fhl1 expression and understanding the influence of estrogen on muscle growth and development.