Embryo

The Role of Homeobox Protein Distal-Less 3 and Its Interaction with ETS2 in Regulating Bovine Interferon-Tau Gene Expression-Synergistic Transcriptional Activation with ETS2

Abstract
Distal-less 3 (DLX3), a homeodomain transcription factor required for placental development in the mouse, modestly transactivates hCG-alpha subunit gene (hCGA) expression in human choriocarcinoma cells. As hCG and interferon-tau (IFNT) are expressed in trophectoderm of primates and ruminants, respectively, we have tested the hypothesis that DLX3 regulates the genes for IFNT (IFNTs). A bovine IFNT1 promoter (-457 to +66), linked to a luciferase (luc) reporter, was transactivated about 20-fold by over expressing DLX3 in human JAr cells. Elimination of a potential DLX3-binding site (-54 GATAATGAG -46) by either truncation or mutagenesis abolished this effect. A sequence (-59 to -44) encompassing this site bound recombinant DLX3 specifically. Co-expression of DLX3 and ETS2, which is known to be a key regulator of IFNT expression, increased reporter activity >250-fold, while deletion of the established ETS2 site (-79 to -70) eliminated the ability of DLX3 to transactivate the gene. Conversely, mutation of the DLX3 site significantly reduced the transactivational effects of ETS2. DLX3 and ETS2 are co-expressed in JAr cells and in an IFNT-producing, bovine trophoblast cell line, CT-1. The two can be immunoprecipitated together as a complex from CT-1 cells, and RNAi-mediated, partial knockdown of DLX3 expression reduced the production of IFNT by about 50%. Together these results suggest that DLX3 has a central role in controlling IFNT gene expression by associating with ETS2 on the IFNT promoter.

Key words: Embryo • Gene regulation • Trophoblast • Interferon-Tau • Transcription factor