FIG. 1.

Deubiquitinating activity in Ad-infected cells. (a) Decline of the Ub conjugates pool in Ad2-infected cells. HeLa cells were infected with Ad2 (multiplicity of infection = 100), whereupon cells were harvested at the indicated time p.i. and whole cell, nuclear, and AS nuclear extracts were prepared. The same amount of the protein from each sample was resolved by SDS-PAGE and analyzed by Western blotting with anti-Ub antibodies. Lane control, extracts from mock-infected cells. Arrows indicate the Ub conjugates significantly diminished during Ad2 infection. The uH2A band was also recognized by anti-uH2A MAb E6C5 (not shown). (b) Ad2-infected cells demonstrate increased DUB activity. Whole cell lysates from Ad2- and mock-infected cells were incubated with zLRGG-AMC fluorogenic substrate (100 μM, 1 h) and the fluorescence of liberated AMC was measured at 345 nm/440 nm. Values are means ± standard deviations (error bars) from three separate experiments