International Immunology, Vol. 13, No. 4, 529-539,
April 2001
© 2001 Japanese Society for Immunology
CTLA-4–Fas ligand functions as a trans signal converter protein in bridging antigen-presenting cells and T cells
Department of Pathology and Laboratory Medicine, University of Pennsylvania, 6 Gates Pavilion, 3400 Spruce Street, Philadelphia, PA 19104-4283, USA
Correspondence to: M. L. Tykocinski
Co-stimulator blockade and trans inhibitory signaling, using agents such as CTLA-4–Ig and Fas ligand (FasL) respectively have been invoked as alternative strategies for suppressing pathogenic T cells. This study describes a novel hetero-bifunctional fusion protein, CTLA-4–FasL, designed to combine within a single protein both co-stimulator blocking and trans inhibitory signaling potentials. A chimeric expression cassette, in which the ectodomain coding sequences for CTLA-4 and FasL were linked in-frame, was used to produce a CTLA-4–FasL fusion protein. CTLA-4–FasL binding to both B7-1/B7-2-expressing Daudi B cells and Fas-expressing Jurkat T cells was documented by immunofluorescence and flow cytometry. The capacity of CTLA-4–FasL to induce apoptosis in Jurkat targets was markedly enhanced by the addition of Daudi and other B7-1/B7-2+ B cell lines, which provided a membrane platform for the otherwise soluble CTLA-4-fusion protein. Moreover, in dual-chamber experiments, Daudi cells pre-coated with CTLA-4–FasL demonstrated Jurkat inhibitory activity that was cell-contact dependent. Significantly, when used to inhibit in vitro cellular proliferation of peripheral blood mononuclear cells, CTLA-4–FasL was ~1000-fold more potent than the extensively characterized CTLA-4–Ig fusion protein. Furthermore, the degree of inhibition induced by CTLA-4–FasL substantially surpassed that observed for CTLA-4–Ig and a soluble FasL when used in combination. CTLA-4–FasL represents the first of a novel class of fusion proteins, designated here as `trans signal converter proteins', that combine trans signal masking and direct trans signaling functions.
Keywords: apoptosis, cell-cell interactions, human, T lymphocytes
Transmitting editor: J. Allison
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