Recombinant interferon-γ 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-γ). In fact, in clone 9229/18, rINF-γ 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 behaviour was investigated and DQA gene regulaton 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-γ 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 cyptoplasmatic mRNA. To study wether 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-γ treatment. In the absence of protein synthesis, the t( 1/2 ) 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-γ in a melanoma clone such as 9229/5.