Fig 9.

TAK1 overexpression promotes cell death and counterselection of DLBCL lines. (A) The ABC-DLBCL line OCI-Ly10 and the GCB-DLBCL line OCI-Ly7 were transduced to express the secreted NF-κB-dependent GLuc reporter. Cells were analyzed for GLuc activity in culture supernatants after 0 and 4 h. −, media only. (B and C) OCI-Ly10 and OCI-Ly7 cells were transduced with LVs (MOI, 27) expressing either HA-TAK1 2A-GFP, HA-TAK1-AAYA 2A-GFP, or GFP only (vector). (B) To assess for counterselection, the GFP⁺ fraction at each time point is compared to GFP marking at day 1 (set as 100%). (C) Cell death was analyzed at days 3 and 9 posttransduction by DAPI incorporation into GFP⁺ cells and represented as fold difference over vector (set as 1). Graphs show means ± SEM from six (A) or three (B and C) independent experiments. **, P > 0.01; *, P > 0.05; ns, not significant, unpaired Student's t test. (D) OCI-Ly7 cells were transduced with LVs expressing TAK1 2A-GFP, TAK1-AAYA 2A-GFP, or GFP only (vector). GFP⁺ cells were sorted at 48 h. Left, GFP expression in untransduced cells or cells expressing TAK1, TAK1-AAYA, or GFP only (vector). Right, cells were incubated with 140 μM CHX for 0 to 6 h, and BCL10, TAK1, and actin levels were analyzed by immunoblotting. BCL10 levels, relative to that of actin, are shown below the blots.