Aims. Chimeric T cells directed to the γ-subunit of the fetal acetylcholine receptor (fAChR) produce large amounts of interferon-γ (IFNγ) on coculture with fAChR-expressing rhabdomyosarcoma (RMS) cells prior to RMS cell death. The aim of this study was to elucidate whether IFNγ blocks proliferation and survival of RMS cells and modulates expression of genes with relevance for cytotoxicity of chimeric T cells. Methods. Expression levels of IFNγ receptor (IFNGR), AChR, MHCI, MHCII, and CIITA (class II transactivator) by RMS were checked by flow cytometry, qRT-PCR, and western blot. Proliferation and cell survival were investigated by annexin V and propidium iodide staining and MTT (thiazolyl-blue-tetrazolium-bromide) assay. Key phosphorylation and binding sites of IFNGRs were checked by DNA sequencing. Results. IFNγ treatment blocked proliferation in 3 of 6 RMS cell lines, but reduced survival in only one. IFNGR was expressed at levels comparable to controls and binding sites for JAK and STAT1 were intact. Induction of several target genes (e.g., AChR, MHCI, and MHCII) by IFNγ was detected on the RNA level but not protein level. Conclusions. IFNγ does not significantly contribute to the killing of RMS cells by fAChR directed chimeric T cells. Signalling downstream of the IFNR receptor, including the posttranscriptional level, is impaired in most RMS cell lines. 1. Introduction Interferon gamma (IFNγ) plays a crucial role in tumor formation and protects host against growth of spontaneous or transplanted tumors [1, 2]. Besides its immunomodulatory effects, IFNγ has an influence on proliferation and induces apoptosis in vitro in many primary tumor cells and established tumor cell lines [3–6]. IFNγ is the only member of the type II interferon family and is mainly produced by activated NK-cells and NKT cells [7], as well as CD4+ T-cells and cytotoxic CD8+ lymphocytes [8]. The active form of the cytokine is a dimer which binds to a heterodimeric receptor complex that consists of IFNGR1 and IFNGR2 subunits and is associated with two Janus kinase family members, Jak1 and Jak2. Changes in confirmation of receptor subunits after IFNγ binding activate Jak1 and Jak2, which in turn phosphorylate IFNGR1 and generate a binding site for recruitment, phosphorylation, and dimerization of signal transducer and activator of transcription 1 (STAT1). After translocation of STAT1 homodimers to the nucleus and binding to GAS (IFNγ activated sites) promotor elements, transcription of target genes is initiated [9–11], including MHC class I and II genes with immunomodulation
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