Figure 8.

Reduced steady state level of the aberrant HLA-A2 mRNA in 624MEL28 melanoma cells. 624MEL28 cells were incubated at 37°C for 72 h with IFN-γ (100 U/ml). Control cells were incubated under the same experimental conditions, but without IFN-γ. At the end of the incubation, total cellular RNA was extracted from cells and hybridized with a ³²P-labeled HLA-A2 riboprobe (496 bases) and with a ³²P-labeled human β-actin riboprobe (250 bases). After hybridization, RNA was digested by RNase, and the generated fragments were size fractionated by 5% PAGE. Gels were autoradiographed for 1 or 7 d. RNA from 624MEL38 and Raji cells were used as HLA-A2–positive and –negative controls, respectively. (A) RNA synthesized in vitro by an HLA-A2 genomic fragment containing exon 2, intron 2, and exon 3 (RNA) was used to identify the fragments generated by RNase digestion from the probe protected by the HLA-A2 mRNA containing intron 2. (B) The fragments generated by RNase digestion from the probe protected by a wild-type HLA-A2 mRNA, by HLA-A2 mRNA with exon 2 skipping, and by HLA-A2 mRNA with intron 2 retention are shown diagrammatically.