ABSTRACT
Friend of GATA (FOG)-2 is a multi-zinc finger transcriptional corepressor protein that binds specifically to GATA4. Gene targeting studies have demonstrated that FOG-2 is required for normal cardiac morphogenesis, including the development of the coronary vasculature, left ventricular compact zone, and heart valves. To better understand the molecular mechanisms by which FOG-2 regulates these cardiac developmental programs, we screened a mouse day 11 embryo library using a yeast two-hybrid interaction trap with the fifth and sixth zinc fingers of FOG-2 as bait. Using this approach, we isolated clones encoding the orphan nuclear receptors chicken ovalbumin upstream promoter-transcription factor (COUP-TF) 2 and COUP-TF3. COUP-TF2-null embryos die during embryonic development with defective angiogenesis and cardiac defects, a pattern that partly resembles the FOG-2-null phenotype. The interaction between COUP-TF2 and FOG-2 in mammalian cells was confirmed by co-immunoprecipitation of these proteins from transfected COS-7 cells. The sites of binding interaction between COUP-TF2 and FOG-2 were mapped to zinc fingers 5 and 6 and fingers 7 and 8 of FOG-2 and to the carboxyl terminus of the COUP-TF proteins. Binding to COUP-TF2 was specific because FOG-2 did not interact with the ligand-binding domains of retinoid X receptor alpha, glucocorticoid receptor, and peroxisome proliferating antigen receptor gamma, which are related to the COUP-TF proteins. Full-length FOG-2 markedly enhanced transcriptional repression by GAL4-COUP-TF2(117-414), but not by a COUP-TF2 repression domain mutant. Moreover, FOG-2 repressed COUP-TF2dependent synergistic activation of the atrial natriuretic factor promoter by both GATA4 and the FOG-2-independent mutant GATA4-E215K. Taken together, these findings suggest that FOG-2 functions as a corepressor for both GATA and COUP-TF proteins.
Subject(s)
Atrial Natriuretic Factor/genetics , DNA-Binding Proteins/chemistry , Promoter Regions, Genetic , Receptors, Steroid , Transcription Factors/chemistry , 3T3 Cells , Animals , COS Cells , COUP Transcription Factor I , COUP Transcription Factors , Cell Line , DNA, Complementary/metabolism , DNA-Binding Proteins/metabolism , GATA4 Transcription Factor , Gene Library , Glutathione Transferase/metabolism , Ligands , Mice , Mutation , Phenotype , Precipitin Tests , Protein Binding , Protein Structure, Tertiary , Receptors, Cytoplasmic and Nuclear/chemistry , Receptors, Glucocorticoid/chemistry , Receptors, Retinoic Acid/chemistry , Recombinant Proteins/metabolism , Retinoid X Receptors , Transcription Factors/metabolism , Transcription, Genetic , Transfection , Two-Hybrid System Techniques , Zinc FingersABSTRACT
We report here the isolation of Tel-2, a novel member of the Ets transcription factor family, with high homology to Tel/ETV-6. Tel-2 is the second mammalian member of the Tel Ets family subclass whose prototype Tel is involved in various chromosomal translocations in human cancers. Six differentially expressed alternative splice products of Tel-2 were characterized encoding different Tel-2 isoforms which either contain or lack the amino-terminal Pointed domain and also vary at the carboxyl terminus. In contrast to Tel, which is highly expressed in several different cell types and tissues, Tel-2 is only weakly expressed in a variety of tissues and cell types, including placenta, prostate, spleen, liver, and lung. Tel-2 binds to functionally relevant Ets-binding sites of several genes and only the Tel-2 isoform containing the Pointed domain and the DNA-binding domain acts as a strong repressor of transcription. The retinoic acid receptor alpha and bone morphogenetic protein-6B (BMP-6) genes are specifically repressed by Tel-2 indicating a function for Tel-2 as an inhibitor of differentiation. Due to the important involvement of Tel in human cancer and the location of Tel-2 within the MHC cluster region, Tel-2 might be involved in chromosomal translocations in human cancer as well.