Stat3 recruitment by two distinct ligand-induced, tyrosine-phosphorylated docking sites in the interleukin-10 receptor intracellular domain.
Recent work has shown that IL-10 induces activation of the JAK-STAT signaling pathway. To define the mechanism underlying signal transducer and activator of transcription (STAT) protein recruitment to the interleukin 10 (IL-10) receptor, the STAT proteins activated by IL-10 in different cell populations were first defined using electrophoretic mobility shift assays. In all cells tested, IL-10 activated Stat1 and Stat3 and induced the formation of three distinct DNA binding complexes that contained different combinations of these two transcription factors. IL-10 also activated Stat5 in Ba/F3 cells that stably expressed the murine IL-10 receptor. Using a structure-function mutagenesis approach, two tyrosine residues (Tyr427 and Tyr477) in the intracellular domain of the murine IL-10 receptor were found to be redundantly required for receptor function and for activation of Stat3 but not for Stat1 or Stat5. Twelve amino acid peptides encompassing either of these two tyrosine residues in phosphorylated form coprecipitated Stat3 but not Stat1 and blocked IL-10-induced Stat3 phosphorylation in a cell-free system. In contrast, tyrosine-phosphorylated peptides containing Tyr374 or Tyr396 did not interact with Stat3 or block Stat3 activation. These data demonstrate that Stat3 but not Stat1 or Stat5 is directly recruited to the ligand-activated IL-10 receptor by binding to specific but redundant receptor intracellular domain sequences containing phosphotyrosine. This study thus supports the concept that utilization of distinct STAT proteins by different cytokine receptors is dependent on the expression of particular ligand-activatable, tyrosine-containing STAT docking sites in receptor intracellular domains.
Center for Immunology, Department of Pathology, Washington University School of Medicine, St. Louis, Missouri 63110, USA. schreiber@immunology.wustl.edu
R M
RM
Weber-Nordt
J K
JK
Riley
A C
AC
Greenlund
K W
KW
Moore
J E
JE
Darnell
R D
RD
Schreiber
D000209:Acute-Phase Proteins / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002474:Cell-Free System; D004268:DNA-Binding Proteins / Q000378:metabolism*; D006801:Humans; D016753:Interleukin-10 / Q000494:pharmacology*; D008024:Ligands; D008214:Lymphocytes / Q000276:immunology; D017737:Macrophages, Peritoneal / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D015345:Oligonucleotide Probes; D010446:Peptide Fragments / Q000737:chemistry; D019000:Phosphotyrosine; D018123:Receptors, Interleukin / Q000737:chemistry* / Q000378:metabolism*; D053701:Receptors, Interleukin-10; D050794:STAT1 Transcription Factor; D050796:STAT3 Transcription Factor; D015534:Trans-Activators / Q000378:metabolism*
D000209:Acute-Phase Proteins; D004268:DNA-Binding Proteins; D008024:Ligands; D015345:Oligonucleotide Probes; D010446:Peptide Fragments; D018123:Receptors, Interleukin; D053701:Receptors, Interleukin-10; D050794:STAT1 Transcription Factor; C494079:STAT1 protein, human; D050796:STAT3 Transcription Factor; C494086:STAT3 protein, human; C494080:Stat1 protein, mouse; C494087:Stat3 protein, mouse; D015534:Trans-Activators; D016753:Interleukin-10; D019000:Phosphotyrosine