Fig. 5. Analysis of ribosome distribution in unstressed and stressed HeLa cells. (A) Upper panel, polysome profiles of mock- (left) or tunicamycin-treated (90 minutes; right) HeLa cells. Absorbance at 254 nm (y axis, indicating RNA concentration) is plotted against migration in the 10-50% sucrose gradients (x axis). The positions of polysomes and ribosomal subunits are indicated. The monosomal 80S peak is not resolved from the 60S subunit peak in these gradients. (B) Fractions corresponding to the profile shown in A were pooled as indicated and analyzed by northern blot using a BiP-specific probe (upper panel) that detected endogenous human BiP mRNA (hBiP) or an actin-specific probe (lower panel). Note that after 90 minutes of tunicamycin treatment, increase in BiP mRNA was still below a factor of 1.5.