ABSTRACT
SDF-1/CXCR4 plays an important role in promoting survival, expansion, and differentiation of T cell progenitors. The present study investigates the mechanism by which estrogen inhibits SDF-1alpha expression in mouse thymus. Mouse estrogen enhanced transcript (mEET) is endogenously expressed in a mouse thymus epithelial cell line 1 (MTEC1). In MTEC1 cells that express the transfected sense mEET, the SDF-1alpha transcription and its chemotactic activity were profoundly inhibited. Conversely, in MTEC1 that express the transfected anti-sense mEET, the SDF-1alpha transcription and its chemotactic activity were substantially augmented. Moreover, we disclosed that mEET inhibited the production of SDF-1alpha by its suppression of NF-kappaB translocation into nucleus. Using a combinatorial induction of doxycycline (Dox) and 17beta-estradiol (E2) on the sense and anti-sense mEET transfectants, it was demonstrated that an increase of mEET expression enhanced E2-induced inhibition of SDF-1alpha production, while a blockade of mEET expression alleviated E2-induced inhibition of SDF-1alpha production. In conclusion, the E2-imposed suppression of SDF-1alpha production is partly mediated by mEET involved signaling pathway.
Subject(s)
Chemokines, CXC/antagonists & inhibitors , Chemokines, CXC/biosynthesis , Estradiol/pharmacology , Phosphoproteins/physiology , Thymus Gland/metabolism , Animals , Cell Line , Chemokine CXCL12 , Chemokines, CXC/immunology , Chemotaxis/immunology , Doxycycline/immunology , Doxycycline/pharmacology , Epithelial Cells/immunology , Epithelial Cells/metabolism , Estradiol/immunology , Female , Gene Expression Regulation/immunology , Male , Mice , Mice, Inbred BALB C , NF-kappa B/immunology , Phosphoproteins/biosynthesis , Phosphoproteins/genetics , Phosphoproteins/immunology , RNA, Messenger/biosynthesis , RNA, Messenger/genetics , Reverse Transcriptase Polymerase Chain Reaction , Thymus Gland/cytology , Thymus Gland/drug effects , Thymus Gland/immunology , Transcription, Genetic/immunologyABSTRACT
A novel gene containing two typical estrogen responsive elements (ERE) was cloned from MTEC1 cells, a mouse thymus epithelial cell line that produces constitutively many chemokines. This gene is a homologue of the rat estrogen-enhanced transcript (EET) gene, and is called the mEET gene. mEET protein is expressed in cytoplasm. Addition of 17 beta-estradiol (E2) to the MTEC1 cell cultures enhanced mEET mRNA expression and, meanwhile, significantly inhibited monocyte chemoattractant protein-1 (MCP-1) production. To analyze the functional links between the expression of mEET and of MCP-1, we transfected MTEC1 cells with ERE-deleted antisense- or sense-mEET complementary (c)DNA construct and activated the transfected mEET cDNA in stable MTEC1 transfectants with doxycycline (Dox). Dox-induced activation of the mEET gene profoundly inhibited MCP-1 expression at both mRNA and protein levels and alleviated its chemotactic activity. Conversely, inactivation of the mEET gene substantially augmented MCP-1 expression. Activation of the mEET gene markedly attenuated activity of nuclear NF-kappaB. In summary, we have first demonstrated that estrogen-imposed inhibition of MCP-1 expression occurs through the activation of the mEET gene, its product suppresses nuclear NF-kappaB and negatively regulates MCP-1 gene activation.
