ID: 188: The two sides of the STAT1 N-domain have opposite effects on binding to GAS elements

Abstract

Recently, it has been shown that substitution of alanine for phenylalanine 77 in the N-domain of STAT1 (signal transducer and activator of transcription (1) prevents cooperative binding of tetramers on DNA and severely impairs transcriptional responses upon stimulation of cells with interferon-gamma (IFN-gamma). Cooperative DNA binding is the basis of efficient switching between non-occupied and occupied promoter states. In this study, we generated N-terminal mutants of STAT1 which showed improved tetramer stability on DNA. We identified two negatively charged N-terminal residues in each protomer of dimeric STAT1 which are required for the dissociation of higher-order oligomers on DNA. Similarly to the STAT1 mutant with impaired tetramerization, these N-terminal mutants showed elevated tyrosine-phosphorylation levels and prolonged nuclear accumulation upon stimulation of cells with IFN-gamma. Unlike the global impairment of IFN-gamma signalling in the tetramerization-deficient mutant, improved tetramer stability of the N-terminal mutants affected transcription in a promoter-specific manner and resulted in a distinct gene expression pattern. In summary, using these mutants we have gained a new mechanistic insight into how protein-DNA interactions regulate STAT1-mediated target gene recognition. While one side of the N-terminal dimer is crucial for the formation of tetrameric complexes on IFN-gamma-regulated promoters, the other side localized in close contact to the longitudinal axis of DNA has a rather inhibitory effect on the formation of higher-order oligomers, simply by disrupting cooperative DNA binding.