Figure 9
IN Q168A failed to accumulate in a nuclear low molecular weight complex. (A) IN Q168A-HA levels rapidly decreased following viral infection. SupT1 cells were infected with HIV-1IN Q168A-HA for 2 h. At indicated time post infection, cells were lysed and equivalent amounts of each sample (100 μg of protein) were analyzed using Western blotting with antibodies against HA and MA. Input represents 0.1& of the amount of virus used to infect the cells. (B) SupT1 cells were infected with HIV-1 viruses encoding WT IN, IN-HA or INQ168A-HA. IN-HA was immunoprecipitated with the anti-HA affinity matrix and analyzed by Western blotting using antibodies against HA and LEDGF/p75. Inputs represent 5& of the immunoprecipitated material. (C) SupT1 cells were infected with HIV-1IN D116A-HA and WCE prepared at 2 h and 6 h p.i. were subjected to gel filtration. Fractions were collected and analyzed by Western blotting using antibodies against HA. Inputs represent 3.5& of WCE load. (D) Cytosolic and nuclear extracts (CE and NE, respectively) were prepared at 2 h p.i., subjected to gel filtration and collected fractions were analyzed by Western blotting using antibody against HA. Inputs represent 3,5& of WCE load.