Lck and proteins in the RANK signaling pathway are significantly up-regulated in Bfl-1ΔC/p53DD tumor-derived cells. (A) Western blots show increased levels of Lck and phospho-Lck in splenocytes from mice injected with FL5.12 cells expressing GFP-Bfl-1ΔC/p53DD versus parental FL5.12 GFP-Bfl-1ΔC/p53DD cells. The blot was probed with anti–phospho-Src/Lck, anti-Lck, or anti–β-actin. FL5.12 cells and FL5.12 cells expressing p53DD alone were negative controls; Jurkat T-cells are a positive control for Lck (1/5 input). (B) Immunoprecipitation of GFP-Bfl-1ΔC/p53DD-tumor-derived cells from mice 1, 2, 3, and 7 with anti-Lck, followed by Western blot with anti–phospho-Src-Lck. (C) Western blot showing elevated levels of MIP-1γ, DAP12, Gab1, and Rab27a in splenocytes derived from the mice analyzed in panel A. (D) Western blots show activation of the Akt, ERK, and IKK signaling pathways in splenocytes derived from the mice analyzed in panel A, as seen with anti-phospho p38, JNK, Akt, ERK, or IKK.