Jun subunits form heterodimers with ATF2 or ATF7 in ABC DLBCL cell lines. (A) Lysates from the indicated ABC and GCB DLBCL cell lines were treated with the cross-linker BS3 or with solvent alone for 1 hour at 4°C. Cross-linked (open arrowheads) and non–cross-linked (filled arrowheads) proteins were revealed by western blot using anti-c-Jun, anti-JunB, and anti-JunD antibodies. (B) BS3-treated HBL-1 cell lysates were immunoprecipitated with anti-c-Jun beads or beads alone, separated by SDS-PAGE and stained with Coomassie. Proteins present in protein complexes of type I and II were analyzed by mass spectrometry (MS). ATF2 and ATF7 were identified to be part of the protein complex type I, whereas not enough material was present in protein complex type II for MS identification. h.c., heavy chain of c-Jun. (C) Protein expression in GCB and ABC cell lines was determined by western blot using anti-ATF2 and anti-ATF7 antibodies. (D) As in panel A, but proteins were revealed using anti-ATF2 and anti-ATF7 antibodies. Gray arrowhead indicates a third, unidentified type of complex. (E) Two ABC and 2 GCB cell lines were lysed and proteins were precipitated using anti-c-Jun, anti-JunB, anti-JunD, or anti-IRE1α antibodies (Ctr, control antibody). Proteins in lysates were analyzed by western blotting with anti-c-Jun, anti-JunB, and anti-JunD antibodies, and coprecipitating proteins (IP) with anti-ATF2 and anti-ATF7 antibodies, as indicated. (F) The composition of protein complexes of type I is schematically depicted. Data are representative of 3 (A,C) and 2 (D,E) independent experiments. *A nonspecific band recognized by anti-ATF7 (C,E) and migration of antibody heavy chains (E). WB, western blot.