Deletion of the carboxyl-terminal transactivation domain of MGF-Stat5 results in sustained DNA binding and a dominant negative phenotype.
The Stat (signal transducer and activator of transcription) factors transmit cytokine, growth factor, and hormone responses. Seven members of the Stat gene family are known. MGF-Stat5a has been discovered as a mediator of the prolactin response in mammary epithelial cells. Two closely related variants of Stat5, Stat5a and Stat5b, are encoded by distinct genes. We examined the functional properties of the carboxyl termini of these molecules. Wild-type Stat5a (794 amino acids) and the carboxyl-terminal deletion mutant Stat5a delta 772 supported prolactin-induced transcription of a beta-casein promoter-reporter construct in COS7 cells; Stat5a delta 750 did not. Upon prolactin activation, tyrosine phosphorylation and the specificity of DNA binding were indistinguishable among the three Stat5a variants. Tyrosine dephosphorylation and the downregulation of the DNA-binding activity were delayed in the Stat5a delta 750 mutant. The carboxyl-terminal transactivation domain of Stat5a, amino acids 722 to 794, can be conferred to the DNA-binding domain of the yeast transcription factor GAL4. Coexpression of Stat5a or Stat5b and of the carboxyl-terminal deletion mutants resulted in the suppression of transcriptional induction in COS or Ba/F3 cells. We propose that Stat5a delta 750 and Stat5b delta 754 are lacking functional transactivation domains and exert their dominant negative effects by blocking the DNA-binding site in Stat5-responsive gene promoters.
Moriggl
R
R
Tumor Biology Center, Institute for Experimental Cancer Research, Freiburg, Germany.
Gouilleux-Gruart
V
V
Jähne
R
R
Berchtold
S
S
Gartmann
C
C
Liu
X
X
Hennighausen
L
L
Sotiropoulos
A
A
Groner
B
B
Gouilleux
F
F
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D019556:COS Cells; D002364:Caseins / Q000235:genetics; D002417:Cattle; D002467:Cell Nucleus / Q000378:metabolism; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*; D005656:Fungal Proteins / Q000096:biosynthesis / Q000378:metabolism; D017930:Genes, Reporter; D006801:Humans; D007700:Kinetics; D008156:Luciferases / Q000096:biosynthesis; D008894:Milk Proteins; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009838:Oligodeoxyribonucleotides; D010641:Phenotype; D019000:Phosphotyrosine / Q000032:analysis; D011401:Promoter Regions, Genetic; D017433:Protein Structure, Secondary; D050799:STAT5 Transcription Factor; D029701:Saccharomyces cerevisiae Proteins; D017384:Sequence Deletion; D017386:Sequence Homology, Amino Acid; D012756:Sheep; D015534:Trans-Activators / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*; D014157:Transcription Factors; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014162:Transfection; D025521:Tumor Suppressor Proteins
D002364:Caseins; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C052130:GAL4 protein, S cerevisiae; D008894:Milk Proteins; D009838:Oligodeoxyribonucleotides; D050799:STAT5 Transcription Factor; C494092:STAT5A protein, human; C494095:STAT5B protein, human; D029701:Saccharomyces cerevisiae Proteins; D015534:Trans-Activators; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; D019000:Phosphotyrosine; D004247:DNA; D008156:Luciferases