The coiled-coil domain of Stat3 is essential for its SH2 domain-mediated receptor binding and subsequent activation induced by epidermal growth factor and interleukin-6.
STAT proteins are a family of latent transcription factors that mediate the response to various cytokines and growth factors. Upon stimulation by cytokines, STAT proteins are recruited to the receptors via their SH2 domains, phosphorylated on a specific tyrosine, dimerized, and translocated into the nucleus, where they bind specific DNA sequences and activate the target gene transcription. STATs share highly conserved structures, including an N-domain, a coiled-coil domain, a DNA-binding domain, a linker domain, and an SH2 domain. To investigate the role of the coiled-coil domain, we performed a systematic deletion analysis of the N-domain and each of the alpha-helices and mutagenesis of conserved residues in the coiled-coil region of Stat3. Our results indicate that the coiled-coil domain is essential for Stat3 recruitment to the receptor and the subsequent tyrosine phosphorylation and tyrosine phosphorylation-dependent activities, such as dimer formation, nuclear translocation, and DNA binding, stimulated by epidermal growth factor (EGF) or interleukin-6 (IL-6). Single mutation of Asp170 or, to a lesser extent, Lys177 in alpha-helix 1 diminishes both receptor binding and tyrosine phosphorylation. Furthermore, the Asp170 mutant retains its ability to bind to DNA when phosphorylated on Tyr705 by Src kinase in vitro, implying a functional SH2 domain. Finally, we demonstrate a direct binding of Stat3 to the receptor. Taken together, our data reveal a novel role for the coiled-coil domain that regulates the early events in Stat3 activation and function.
Zhang
T
T
Institute of Molecular and Cell Biology, Singapore 117609, Singapore.
Kee
W H
WH
Seow
K T
KT
Fung
W
W
Cao
X
X
D000818:Animals; D015703:Antigens, CD / Q000378:metabolism; D019556:COS Cells; D006528:Carcinoma, Hepatocellular / Q000473:pathology; D050822:Cytokine Receptor gp130; D004247:DNA / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000502:physiology; D019281:Dimerization; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D005786:Gene Expression Regulation; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D015850:Interleukin-6 / Q000494:pharmacology*; D008113:Liver Neoplasms / Q000473:pathology; D008562:Membrane Glycoproteins / Q000378:metabolism; D051379:Mice; D009363:Neoplasm Proteins / Q000378:metabolism; D010766:Phosphorylation; D017354:Point Mutation; D011485:Protein Binding; D011499:Protein Processing, Post-Translational; D017433:Protein Structure, Secondary; D019947:Receptors, Interleukin-6 / Q000187:drug effects / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000502:physiology; D050796:STAT3 Transcription Factor; D013329:Structure-Activity Relationship; D015534:Trans-Activators / Q000737:chemistry* / Q000502:physiology; D014158:Transcription, Genetic / Q000187:drug effects*; D014162:Transfection; D014407:Tumor Cells, Cultured; D018909:src Homology Domains
D015703:Antigens, CD; D004268:DNA-Binding Proteins; C494199:IL6ST protein, human; C494200:Il6st protein, mouse; D015850:Interleukin-6; D008562:Membrane Glycoproteins; D009363:Neoplasm Proteins; D019947:Receptors, Interleukin-6; D011993:Recombinant Fusion Proteins; D050796:STAT3 Transcription Factor; C494086:STAT3 protein, human; C494087:Stat3 protein, mouse; D015534:Trans-Activators; D050822:Cytokine Receptor gp130; D004815:Epidermal Growth Factor; D004247:DNA