Identification of a functionally important sequence in the C terminus of the interferon-gamma receptor.
We have previously shown that the intracellular domain of the interferon-gamma (IFN-gamma) receptor plays an obligate role in receptor-mediated signal transduction. Moreover, we have specifically identified two regions within the human IFN-gamma receptor's intracellular domain required for functional activity: the membrane-proximal 48 amino acids required for both functional activity and receptor-mediated ligand internalization and the C-terminal 39 amino acids required exclusively for biologic response induction. Herein we report the identification of the 3 amino acids within the C-terminal region of the receptor that are obligatorily required for receptor function. By using a set of overlapping truncation mutants, the minimal functional sequence within the C-terminal region was localized to residues 434-444 (APTSFGYD-KPH). By mutating each individual residue within this sequence to alanine, three residues (Tyr-440, Asp-441, and His-444) were identified as being critical for IFN-gamma-dependent (i) upregulation of major histocompatibility complex class I proteins, (ii) activation of the IFN regulatory factor 1 gene, and (iii) stimulation of cells to produce nitric oxide. The more conservative Tyr-440-->Phe substitution also resulted in a nonfunctional receptor. Subsequent mutational analysis of all five of the IFN-gamma receptor's intracellular tyrosine residues revealed that Tyr-440 was the sole tyrosine required for receptor activity. These results thus identify a unique sequence in the IFN-gamma receptor that is required for initiation of IFN-gamma-dependent biologic responses and highlight the importance of the hydroxyl side chain of Tyr-440 in this process.
Farrar
M A
MA
Department of Pathology, Washington University School of Medicine, St. Louis, MO 63110.
Campbell
J D
JD
Schreiber
R D
RD
D000595:Amino Acid Sequence; D000818:Animals; D004252:DNA Mutational Analysis; D004268:DNA-Binding Proteins / Q000235:genetics; D005786:Gene Expression Regulation; D005805:Genes, MHC Class I; D006801:Humans; D050856:Interferon Regulatory Factor-1; D007371:Interferon-gamma / Q000502:physiology*; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009569:Nitric Oxide / Q000378:metabolism; D010750:Phosphoproteins / Q000235:genetics; D017471:Receptors, Interferon / Q000737:chemistry*; D011994:Recombinant Proteins; D015398:Signal Transduction; D014443:Tyrosine / Q000737:chemistry
D004268:DNA-Binding Proteins; C494257:IRF1 protein, human; D050856:Interferon Regulatory Factor-1; C494258:Irf1 protein, mouse; D010750:Phosphoproteins; D017471:Receptors, Interferon; D011994:Recombinant Proteins; D009569:Nitric Oxide; D014443:Tyrosine; D007371:Interferon-gamma