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Caspase inhibitors block oxidative stress-induced changes in LANA.

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posted on 28.07.2015, 03:59 authored by David A. Davis, Nicole E. Naiman, Victoria Wang, Prabha Shrestha, Muzammel Haque, Duosha Hu, Holda A. Anagho, Robert F. Carey, Katharine S. Davidoff, Robert Yarchoan

BCBL-1 (A, B) or BC-3 (C, D) cells were treated with PBS vehicle control or 100 μM H2O2 in the presence of 50 μM Z-FA-FMK (- ctrl), a negative control peptide (lanes 1 and 2), ZVAD-FMK (ZVAD), a pan-caspase inhibitor (lanes 3 and 4), a specific inhibitor of caspase-1 (C1 inh) (lanes 5 and 6) or a specific inhibitor of caspase-3/7 (C3/7 inh) (lanes 7 and 8). Nuclear (A, C) and cytoplasmic (B, D) extracts were harvested and LANA expression was analyzed by western blot and probed with antibody to FLAG, anti-rabbit secondary antibody conjugated to alkaline phosphatase and visualized using stabilized Western Blue substrate (Promega). Results are representative of 3 separate experiments for controls and ZVAD and 2 separate experiments for the C1 and C3/7 inhibitors. Molecular weight markers are shown to the left. To the right of the blots the different forms of LANA are indicated and include the location for full length LANA (LANA-fl) and three forms of LANA migrating below full length LANA with presumptive designations as LANAcp3, LANAcp2 and LANAcp5 based on their mobility.