Structural characterization of the PIT-1/ETS-1 interaction: PIT-1 phosphorylation regulates PIT-1/ETS-1 binding.
The POU-domain transcription factor Pit-1 and Ets-1, a member of the ETS family of transcription factors, can associate in solution and synergistically activate the prolactin promoter by binding to a composite response element in the prolactin promoter. We mapped the minimal region of Ets-1 required for the interaction with the Pit-1 POU-homeodomain. Here, we describe a detailed NMR study of the interaction between the POU-homeodomain of Pit-1 and the minimal interacting region of Ets-1. By using heteronuclear single quantum coherence titration experiments, we were able to map exact residues on the POU-homeodomain that are involved in the interaction with this minimal Ets-1 interaction domain. By using our NMR data, we generated point mutants in the POU-homeodomain and tested their effect on the interaction with Ets-1. Our results show that phosphorylation of Pit-1 can regulate the interaction with Ets-1.
Augustijn
Kevin D
KD
Department of Physiological Chemistry, University Medical Center Utrecht and Center for Biomedical Genetics, Universiteitsweg 100, 3584 CG, Utrecht, The Netherlands.
Duval
Dawn L
DL
Wechselberger
Rainer
R
Kaptein
Rob
R
Gutierrez-Hartmann
Arthur
A
van der Vliet
Peter C
PC
D000818:Animals; D002645:Chickens; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D010766:Phosphorylation; D010957:Plasmids / Q000378:metabolism; D017354:Point Mutation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D050784:Proto-Oncogene Protein c-ets-1; D011518:Proto-Oncogene Proteins / Q000737:chemistry* / Q000378:metabolism; D050783:Proto-Oncogene Proteins c-ets; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013997:Time Factors; D050817:Transcription Factor Pit-1; D014157:Transcription Factors / Q000737:chemistry* / Q000378:metabolism
D004268:DNA-Binding Proteins; C494041:ETS1 protein, human; C494187:POU1F1 protein, human; D050784:Proto-Oncogene Protein c-ets-1; D011518:Proto-Oncogene Proteins; D050783:Proto-Oncogene Proteins c-ets; D011993:Recombinant Fusion Proteins; D050817:Transcription Factor Pit-1; D014157:Transcription Factors; D005982:Glutathione Transferase