Mutational analysis of acute-phase response factor/Stat3 activation and dimerization.
Signal transducer and transcription (STAT) factors are activated by tyrosine phosphorylation in response to a variety of cytokines, growth factors, and hormones. Tyrosine phosphorylation triggers dimerization and nuclear translocation of these transcription factors. In this study, the functional role of carboxy-terminal portions of the STAT family member acute-phase response factor/Stat3 in activation, dimerization, and transactivating potential was analyzed. We demonstrate that truncation of 55 carboxy-terminal amino acids causes constitutive activation of Stat3 in COS-7 cells, as is known for the Stat3 isoform Stat3beta. By the use of deletion and point mutants, it is shown that both carboxy- and amino-terminal portions of Stat3 are involved in this phenomenon. Dimerization of Stat3 was blocked by point mutations affecting residues both in the vicinity of the tyrosine phosphorylation site (Y705) and more distant from this site, suggesting that multiple interactions are involved in dimer formation. Furthermore, by reporter gene assays we demonstrate that carboxy-terminally truncated Stat3 proteins are incapable of transactivating an interleukin-6-responsive promoter in COS-7 cells. In HepG2 hepatoma cells, however, these truncated Stat3 forms transmit signals from the interleukin-6 signal transducer gp130 equally well as does full-length Stat3. We conclude that, dependent on the cell type, different mechanisms allow Stat3 to regulate target gene transcription either with or without involvement of its putative carboxy-terminal transactivation domain.
Sasse
J
J
Institute of Biochemistry, Rheinisch-Westfälische Technische Hochschule Aachen, Germany.
Hemmann
U
U
Schwartz
C
C
Schniertshauer
U
U
Heesel
B
B
Landgraf
C
C
Schneider-Mergener
J
J
Heinrich
P C
PC
Horn
F
F
D000209:Acute-Phase Proteins / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D006528:Carcinoma, Hepatocellular; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D019281:Dimerization; D006801:Humans; D015850:Interleukin-6 / Q000378:metabolism; D008113:Liver Neoplasms; D008969:Molecular Sequence Data; D010766:Phosphorylation; D017354:Point Mutation; D011401:Promoter Regions, Genetic / Q000235:genetics; D050794:STAT1 Transcription Factor; D050796:STAT3 Transcription Factor; D017384:Sequence Deletion; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*; D015533:Transcriptional Activation / Q000502:physiology*; D014407:Tumor Cells, Cultured; D014443:Tyrosine / Q000378:metabolism; D018909:src Homology Domains
D000209:Acute-Phase Proteins; D004268:DNA-Binding Proteins; D015850:Interleukin-6; D050794:STAT1 Transcription Factor; C494079:STAT1 protein, human; D050796:STAT3 Transcription Factor; C494086:STAT3 protein, human; D015534:Trans-Activators; D014443:Tyrosine; D004247:DNA