The activities of E2F transcription factors are inhibited by interactions with members of the retinoblastoma (RB) tumor suppressor family, p105RB, p107 and p130. In cycling cells p107 and p130 also interact with heterodimers comprised of Cdk2 and either A or E cyclins. We characterized E2F complexes present in C2C12 and P19 mouse cells induced to differentiate into muscle and neuronal cells, respectively. In both undifferentiated C2C12 and P19 cells, in addition to free species, E2F was found in complexes containing p107 or p130 and Cdk2. No E2F-pRB complexes were detected by electrophoretic mobility shift assays even though such cells were shown to contain pRB and E2F species capable of interacting in vitro. These results suggested that although present, pRB was unable to interact with E2F. Following differentiation of C2C12 cells into myotubes, E2F was present in at least two complexes which contained p130, but not in those containing p107 or Cdk2. Low levels of E2F-pRB complexes were also detected in fully differentiated C2C12 myotubes and in freshly isolated skeletal muscle. In the case of differentiated P19 neuronal cells, E2F was found in complexes containing pRB, p107 and p130. However, such cells may not be representative of fully differentiated neurons, as studies with rodent brain extracts indicated that only pRB-E2F complexes and those recognized by a p130-specific serum were present. These results suggested that in both muscle and neurons, pRB and p130 may play specific roles in the development or maintenance of terminal differentiation.