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. 2004 Jan;78(2):821–833. doi: 10.1128/JVI.78.2.821-833.2004

FIG. 5.

FIG. 5.

FIG. 5.

Effects of 16E1∧E4 mutants on keratin association and keratin dynamics. (A) Twenty-four hours after infection with rAd16E1∧E4ΔLLKLL or rAd16E1∧E4Δ87-92, SiHa cells were fixed, permeabilized, and stained with anti-16E1∧E4 antibody (red), anti-K8/K18 antibody (green), and DAPI (blue). The images were taken using 40× (upper panels) or 60× (lower panels) objectives. Whereas 16E1∧E4Δ87-92 showed evidence of keratin association, 16E1∧E4ΔLLKLL did not. (B) The 16E1∧E4 mutants, 16E1∧E4ΔLLKLL and 16E1∧E4Δ87-92, did not disturb keratin dynamics. G2/M-enriched SiHa cells expressing 16E1∧E4wt (E4wt), 16E1∧E4ΔLLKLL (ΔLLKLL), 16E1∧E4Δ87-92 (Δ87-92), or β-Gal were harvested and fractionated as described in the legend to Fig. 4. Equal volumes of the four cellular fractions were loaded and Western blotted to detect 16E1∧E4, K18, and 14-3-3 (all isoforms). In contrast to wt 16E1∧E4, in which the soluble keratin pool was almost completely lost, in cells expressing β-Gal or the mutant 16E1∧E4 proteins, keratin dynamics were unaltered. Emp, Empigen.