PMC

Global comparison of gene expression in HPV31 and NHKs. Each dot corresponds to the Cy3 (x axis) and Cy5 (y axis) fluorescent intensity of one single element on the microarray. Twofold, 5-fold, and 10-fold changes in expression are indicated as parallel lines. Dots that represent more than 11.5-fold changes are internal controls for hybridization.

The induction of Stat-1 protein by IFN-γ is impaired in HPV31 cells. (A) Western blot analysis of equal amounts of protein lysates from NHKs and LKP31 cells treated with IFN-γ (1,000 U/ml) to detect Stat-1 protein level. The image in panel A was quantified by a densitometer, and the results are shown graphically in panel B.

Level of expression in genes suppressed in HPV31 cells demonstrated by microarray analysis is confirmed by Northern blot analysis. Total RNAs (20 μg) from NHKs or LKP31 cells were separated by formaldehyde agarose electrophoresis, transferred to a membrane, and hybridized with various cDNA probes. The slightly faster mobility of p21 and Stat-1 signals in LKP31 cells is likely due to electrophoretic artifact since it was not observed in separate experiments.

Induction of MxA RNA by IFN-α is impaired in HPV31 cells. (A) Northern blot analysis of 8 μg of total RNA from NHK or LKP31 cells treated with IFN-α (100 or 500 U/ml) with MxA cDNA as a probe. MxA RNA signals in panel A were quantified by a PhosphorImager, and the results are shown graphically in panel B. The induction of MxA RNA by IFN-α reaches comparable levels in LKP31 cells and NHKs at a higher dose (1,000 U/ml) and longer time exposure. (C) Northern blot analysis of 10 μg of total RNA from NHKs and LKP31 cells treated with IFN-α with MxA cDNA as a probe. MxA RNA signals in panel C were quantified by a PhosphorImager, and the results are shown graphically in panel D.

The induction of Stat-1 protein by IFN-α is impaired in HPV31 cells. (A) Western blot analysis of equal amounts of protein lysates from NHKs and LKP31 cells treated with IFN-α (100 or 500 U/ml) to detect Stat-1 protein level. Stat-1 protein migrates in SDS-PAGE as a doublet with molecular weights of 91,000 and 84,000. Quantitative estimation of the results in panel A was done with a densitometer and shown graphically in panel B. The induction of Stat-1 protein by IFN-α reaches comparable levels in LKP31 cells and NHKs with a higher dose (1,000 U/ml) and longer time. (C) Western blot analysis of equal amount of protein lysates from NHKs or LKP31 cells treated with IFN-α to detect Stat-1 protein. Quantitative estimation of the results in panel C was done with a densitometer and shown graphically in panel D.