Figure 1.

BimS does not induce apoptosis in bax^−/−bak^−/− MEF. (A) MEF of different genotypes were infected with pBabe–IRES–GFP (vector) or pBabe–BimS–IRES–GFP (BimS). Twenty-four hours after infection, cells were stained with DAPI, and photographed under phase contrast or DAPI filter. (B) Cells from A were collected and subjected to flow cytometric analysis (FACS). The percentage of dead cells was determined by the ratio of DAPI-positive cells to GFP-positive cells. Data shown is the average of three independent experiments. (C) Western blot with anti-Bim of lysates obtained from MEF with different genotypes infected with BimS. Ectopically expressed BimS and endogenous BimEL and BimL are indicated. (D) BimS interacts with Bcl-2 in bax^−/−bak^−/− cells. bax^−/−bak^−/− MEF infected with control vector or BimS were lysed. Immunoprecipitation and immunoblotting as described in Materials and Methods were performed with an anti-Bcl-2 antibody. A cell lysate from wild-type MEF was loaded as a control for Bcl-2 migration. The migration of immunoglobulin light chain (Ig) is also indicated. (E) BimS interacts with Bcl-x_L in bax^−/−bak^−/− cells. MEF infected with BimS or control vector were lysed. Immunoprecipitation and immunoblotting were performed as described in D by use of an anti-Bcl-x_L antibody. A cell lysate from wild-type MEF was loaded as a control for Bcl-x_L migration. The migration of immunoglobulin light chain (Ig) is also indicated. (F) Levels of Bcl-2 and Bcl-x_L are not altered in bax^−/−bak^−/− MEF. A Western blot of MEF lysates from different genotypes was probed with antibodies against Bcl-2, Bcl-x_L, and actin.