Recombinant interferon-gamma differently affects DQA and DRA gene expression in two human melanoma clones: transcriptional and posttranscriptional regulation.
In a previous study, two human melanoma HLA class-II-negative clones have been shown to respond differently to recombinant interferon gamma (rINF-gamma). In fact, in clone 9229/18, rINF-gamma treatment led to a coordinate expression of both DR and DQ genes, whereas in clone 9229/5, a high increase in steady-state mRNA levels and cell surface antigen expression could be observed for DR but not for DQ genes. The molecular mechanism underlying such a different behavior was investigated and DQA gene regulation was studied both at the transcriptional and posttranscriptional level. Nuclear run-on experiments were performed on 9229/5 and 9229/18 clones. Treatment with rINF-gamma at 1,000 U/ml for 24 h led to a coordinate transcriptional activation of DRA and DQA genes in 9229/18 clone, whereas in clone 9229/5 it strongly augmented the rate of transcription of DRA but not DQA genes. For all class II genes studied, both melanoma clones showed a basal rate of transcription that never led to a mature cytoplasmatic mRNA. To study whether posttranscriptional mechanisms could affect DQA mRNA stability, a comparison was performed between mRNA turnover in 9229/18 cells treated with actinomycin D and actinomycin D plus cyclohexamide following rINF-gamma treatment. In the absence of protein synthesis, the t1/2 of specific DQA mRNA was largely reduced, showing that a short-lived protein is required to stabilize human DQA mRNA in melanoma cells. Our results indicate that DQA gene is subjected to a tight regulation, acting both at the transcriptional and posttranscriptional levels and that DQA and DRA genes can be differentially regulated at the transcriptional level by rINF-gamma in a melanoma clone such as 9229/5.
Castelli
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C
Division of Experimental Oncology D, Istituto Nazionale per lo Studio e la Cura dei Tumori, Milan, Italy.
Sensi
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M
Parmiani
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G
D001483:Base Sequence; D015342:DNA Probes; D015972:Gene Expression Regulation, Neoplastic; D006683:HLA-DQ Antigens / Q000235:genetics*; D006684:HLA-DR Antigens / Q000235:genetics*; D006801:Humans; D007371:Interferon-gamma / Q000494:pharmacology*; D008545:Melanoma / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D008969:Molecular Sequence Data; D012323:RNA Processing, Post-Transcriptional; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured / Q000276:immunology / Q000378:metabolism
D015342:DNA Probes; D006683:HLA-DQ Antigens; D006684:HLA-DR Antigens; D012333:RNA, Messenger; D007371:Interferon-gamma