Characterization of an IL-12 p40/p35 Truncated Fusion Protein That can Inhibit the Action of IL-12.
Interleukin-12 (IL-12), a potent inducer of interferon gamma (IFNγ), is a heterodimeric protein consisting of p40 and p35 subunits whose expression is regulated independently. IL-12 is part of a cytokine family (currently consisting of IL-12, IL-23, IL-27, and IL-35) that can have profoundly different immunologic effects, despite sharing subunits. In constructing a single-chain fusion of p40 and p35, we discovered an insert corresponding to an intron in the gene encoding the p35 subunit that would result in a truncated form of p35 if translated. To test its possible role, we constructed, expressed, and analyzed fusions of p40 with the full-length or the truncated form of p35. The fusion protein containing the truncated p35 did not stimulate the proliferation of the IL-12-responsive cell line CTLL-2 nor did it induce IFNγ or the chemokine IFNγ-inducible protein 10 (IP-10, CXCL10) or monokine induced by IFNγ (MIG, CXCL9) from spleen cells. In striking contrast, the full-length IL-12 p40/p35 fusion induced robust responses in both assays. Moreover, the truncated IL-12 fusion protein inhibited the action of the full-length IL-12 p40/p35 fusion in the proliferation assay and also blocked the induction of IFNγ. These findings raise the possibility that alternative splicing may provide an additional regulatory mechanism for IL-12.
Department of Microbiology and Immunology, University of Rochester Medical Center , Rochester, New York.
Denise
D
Skrombolas
Department of Microbiology and Immunology, University of Rochester Medical Center , Rochester, New York.
Isabel
I
Wylie
Department of Microbiology and Immunology, University of Rochester Medical Center , Rochester, New York.
Shivana
S
Maharaj
Department of Microbiology and Immunology, University of Rochester Medical Center , Rochester, New York.
John G
JG
Frelinger
D000595:Amino Acid Sequence; D000818:Animals; D002478:Cells, Cultured; D054357:Chemokine CXCL10 / Q000378:metabolism; D054370:Chemokine CXCL9 / Q000378:metabolism; D007371:Interferon-gamma / Q000378:metabolism; D018664:Interleukin-12 / Q000378:metabolism*; D053765:Interleukin-12 Subunit p35 / Q000378:metabolism*; D053762:Interleukin-12 Subunit p40 / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D021122:Protein Subunits / Q000378:metabolism
D054357:Chemokine CXCL10; D054370:Chemokine CXCL9; C485433:Cxcl10 protein, mouse; C485969:Cxcl9 protein, mouse; D053765:Interleukin-12 Subunit p35; D053762:Interleukin-12 Subunit p40; D021122:Protein Subunits; D018664:Interleukin-12; D007371:Interferon-gamma